Chapter 89: Chronic Myelogenous Leukemia and Related Disorders

Chronic myelogenous leukemia (CML) is a multipotential hematopoietic stem cell disease characterized by anemia, extreme blood granulocytosis and granulocytic immaturity, basophilia, often thrombocytosis, and splenomegaly. The hematopoietic cells contain a reciprocal translocation between chromosomes 9 and 22 in more than 95 percent of patients with classic morphologic findings, which leads to an overtly foreshortened long arm of one of the pair of chromosome 22 (i.e., 22,22q−), referred to as the Philadelphia (Ph) chromosome. A rearrangement of the breakpoint cluster region gene (BCR) on the long arm of chromosome 22 defines this form of CML and is present even in the 10 percent of patients without an overt 22q abnormality by Giemsa chromosome banding. The natural history of the disease is to undergo clonal evolution into an accelerated phase and/or a rapidly progressive blast phase, an acute leukemia, highly refractory to therapy, which had been a frequent event prior to the introduction of tyrosine kinase inhibitors (TKIs) in 2001.

In 1845, Bennett¹ in Scotland and Virchow² in Germany described patients with splenic enlargement, severe anemia, and enormous concentrations of leukocytes in their blood at autopsy. Bennett initially favored an extreme pyemia as the explanation, but Virchow argued against suppuration as a cause. Additional cases were reported by Craige³ and others, and in 1847 Virchow⁴ introduced the designation weisses Blut and leukämie (leukemia). In 1878, Neumann⁵ proposed that the marrow not only was the site of normal blood cell production, but also was the site from which leukemia originated and used the term myelogene (myelogenous) leukemia. Subsequent observations amplified the clinical and laboratory features of the disease, but few fundamental insights were gained until the discovery by Nowell and Hungerford,⁶ who reported in 1960 that two patients with the disease had an apparent loss of the long arm of chromosome 21 or 22, an abnormality that was quickly confirmed^7,8,9 and designated the Ph chromosome.⁷ Advanced cytogenetic techniques confirmed that it was chromosome 22. This observation led to a new approach to diagnosis, a marker to study the pathogenesis of the disease, and a focus for future studies of the molecular pathology of the disease. The availability of banding techniques to define the fine structure of chromosomes^10,11 led to the discovery by Rowley¹² that the apparent lost chromosomal material on chromosome 22 was part of a reciprocal translocation between chromosomes 9 and 22. The discovery that the cellular oncogene ABL1 on chromosome 9 and a segment of chromosome 22, the BCR, fuse as a result of the translocation provided a basis for the study of the molecular cause of the disease.^13,14 The appreciation that the fusion gene encoded a constitutively active tyrosine kinase (BCR-ABL1) that was capable of inducing the disease in mice established the fusion gene product as the proximate cause of the malignant transformation. The search for, identification of, and clinical development of a small molecule inhibitor of the mutant tyrosine kinase provided a specific agent, imatinib mesylate, with which to inhibit the molecule that incites the disease.¹⁵ Several more potent congeners have also been synthesized (see “Etiology and Pathogenesis” below). Thomas and colleagues established that allogeneic hematopoietic stem cell transplantation could cure the disease.¹⁶ An engaging monograph on the discoveries and the scientists involved from the identification of the Ph chromosome to the development of imatinib has been published.¹⁷

CML accounts for approximately 15 percent of all cases of leukemia, or approximately 6500 new cases in the United States in 2015. The age-adjusted incidence rate in the United States is approximately 2.3 per 100,000 persons for men and approximately 1.2 per 100,000 persons for women. The incidence around the world varies by a factor of approximately twofold. The lowest incidence is in Sweden and China (approximately 0.7 per 100,000 persons), and the highest incidence is in Switzerland and the United States (approximately 1.5 per 100,000 persons).¹⁸ The age-specific incidence rate for CML in the United States increases logarithmically with age, from approximately 0.2 per 100,000 per year in persons younger than 20 years to a rate of approximately 10.0 per 100,000 in octogenarians (Fig. 89–1). Although CML occurs in children and adolescents, less than 10 percent of all cases occur in persons between 1 and 20 years old. CML represents approximately 3 percent of all childhood leukemias. Multiple occurrences of CML in families are rare. There is no concordance of the disease between identical twins. There is no analytical epidemiologic evidence for a familial predisposition to CML in Swedish databases.¹⁹ There is some evidence that overweightness and obesity can increase the incidence of CML.²⁰

ENVIRONMENTAL LEUKEMOGENS

Exposure to very high doses of ionizing radiation can increase the occurrence of CML above the expected frequency in comparable populations. Three major populations—the Japanese exposed to the radiation released by the atomic bomb detonations at Nagasaki and Hiroshima,²¹ British patients with ankylosing spondylitis treated with spine irradiation,²² and women with uterine cervical carcinoma who received radiation therapy²³—had a frequency of CML (as well as acute leukemia) significantly above the frequency expected in comparable unexposed groups. The median latent period was approximately 4 years in irradiated spondylitics, among whom approximately 20 percent of the leukemia cases were CML; 9 years in the uterine cervical cancer patients, of whom approximately 30 percent had CML; and 11 years in the Japanese survivors of the atomic bombs, of whom approximately 30 percent of the leukemia patients had CML.²⁴ Chemical leukemogens, such as benzene and alkylating agents, are not causative agents of CML, presumably because of their inability to induce the specific chromosome translocation required to cause the disease.^25,26,27

ORIGIN FROM A MUTANT HEMATOPOIETIC STEM CELL

CML results from the malignant transformation of a single hematopoietic stem cell. The disease is acquired (somatic mutation), given that the identical twin of patients with CML and the offspring of mothers with the disease neither carry the Ph chromosome nor develop the disease.²⁸ The origin of CML from a single hematopoietic stem cell is supported by the following lines of evidence:

1. Involvement of erythropoiesis, neutrophilopoiesis, eosinophilopoiesis, basophilopoiesis, monocytopoiesis, and thrombopoiesis in chronic phase CML.²⁹

2. Presence of the Ph chromosome (22q−) in erythroblasts; neutrophilic, eosinophilic, and basophilic granulocytes; macrophages; and megakaryocytes.³⁰

3. Presence of a single glucose-6-phosphate dehydrogenase isoenzyme in red cells, neutrophils, eosinophils, basophils, monocytes, and platelets, but not in fibroblasts or other somatic cells in women with CML who are heterozygotes for isoenzymes A and B.^31,32,33

4. Presence of the Ph translocation only on a structurally anomalous chromosome 9 or 22 of each chromosome pair in every cell analyzed in occasional patients with a structurally dissimilar 9 or 22 chromosome within the pair.^34,35,36

5. Presence of the Ph chromosome in one, but not the other, cell lineage of patients who are a mosaic for sex chromosomes, as in Turner syndrome (45X/46XX)³⁷ and Klinefelter syndrome (46XY/47XXY).³⁸

6. Molecular studies showing variation in the breakpoint of chromosome 22 among different patients with CML but precisely the same breakpoint among cells within a single patient with CML.^39,40

7. Combined DNA hybridization-methylation analysis of women who have restriction fragment length polymorphisms at the X-linked locus for hypoxanthine phosphoribosyltransferase (HPRT), which enables distinction of the two alleles of the HPRT gene in heterozygous females, coupled with methylation-sensitive restriction-enzyme cleavage patterns, which permits delineation of whether cells contain either the maternally derived or the paternally derived copy of the gene.⁴¹

The foregoing observations place the parent cell of the clone at least at the level of the hematopoietic multipotential cell.

THE CHRONIC MYELOGENOUS LEUKEMIA STEM CELL

Acquisition of the BCR-ABL1 fusion gene as a result of the t(9;22) (q34;q11.2) in a single primitive multipotential hematopoietic cell (possibly the pluripotential stem cell) results in the CML stem cell, necessary for the initiation and maintenance of the chronic phase of CML.^42,43 The phenotype of the CML stem cell is not fully defined but they are among the CD34+CD33−Lin−Thy1+ KIT− fraction of CML cells.⁴³ A proportion of CML stem cells is in the G₀ phase of the cell cycle and is resistant to therapy with BCR-ABL1 inhibitors. These cells represent a pool for the regrowth of the tumor in most patients, if suppressive therapy is interrupted. The leukemia stem cell is resistant to TKI therapy, but a pan-BCL2 inhibitor has been found to sensitize marrow leukemia stem cells to tyrosine kinase inhibition.⁴⁴ N-cadherin and WNT-β-catenin signaling are also thought to mediate microenvironmental protection of CML stem cells from TKIs.⁴⁵ The acquisition of genetic and epigenetic events in a derivative BCR-ABL1–positive cell can result in evolution to accelerated phase and blastic transformation⁴⁶ (see “Accelerated Phase and Blast Crisis of Chronic Myelogenous Leukemia” below).

PLURIPOTENTIAL STEM CELL LESION

Some patients in chronic phase CML have lymphocytes that are derived from the primordial malignant cell. Evidence for this finding includes the following: A single isoenzyme for glucose-6-phosphate dehydrogenase has been found in some T and B lymphocytes in women with CML who are heterozygous for isoenzymes A and B⁴⁷; blood cells from patients with CML induced to proliferate with Epstein-Barr virus (presumptive B lymphocytes) are of the same glucose-6-phosphate dehydrogenase isoenzyme type, have cytoplasmic immunoglobulin heavy and light chains, and contain the Ph chromosome⁴⁸; blood lymphocytes stimulated with B lymphocyte mitogens contain the Ph chromosome^49,50; purified B lymphocytes from the blood in chronic phase CML contain an abnormal, elongated phosphoprotein coded for by the chimeric gene resulting from the t(9;22)⁵¹; and fluorescence in situ hybridization (FISH) has detected the BCR-ABL1 fusion gene in approximately 25 percent of B lymphocytes in some, but not all, patients in chronic phase.^52,53 These findings suggest that B lymphocytes are derived from the malignant clone, placing the lesion closer to, if not in, the pluripotential lymphohematopoietic stem cell.^{47,48,49,50,51} Previous studies have found that the B lymphocyte pool is a mosaic, containing both Ph chromosome– and BCR-ABL1–positive cells and Ph chromosome– or BCR-ABL1–negative cells. Results of studies examining the derivation of T lymphocytes from the malignant clone are more ambiguous but indicate that T lymphocytes are derived from the malignant clone in some patients.^{47,49,54,55,56,57,58,59,60,61,62,63} Natural killer (NK) cells isolated from patients with chronic phase CML do not contain the BCR-ABL1 fusion gene.⁶⁴ It is possible that myelopoiesis is invariably clonal and lymphopoiesis is an unpredictable mosaic derived largely from normal residual stem cells. This conclusion is supported by the finding that progenitors of T, B, and NK lymphocytes contain the Ph chromosome and BCR-ABL1, but most B-cell and all T-cell progenitors derived from the leukemic clone undergo apoptosis, leaving unaffected cells in the blood.^65,66,67,68

The cell in which the mutation occurs may be even more primitive in that some endothelial cells generated in vitro express the BCR-ABL1 fusion gene, as do some cells in the patient’s vascular endothelium.⁶⁹

ETIOLOGIC ROLE OF THE Ph CHROMOSOME

Early studies indicated that the Ph chromosome may appear after the initial leukemogenic event.^70,71,72,73 Patients with CML have developed the Ph chromosome during the course of the disease, have experienced periods of the disease when the Ph chromosome disappeared,⁷⁴ or have had Ph chromosome–positive and Ph chromosome–negative cells concurrently.^{75,76,77,78,79}

Nearly all, if not all, patients with CML have an abnormality of chromosome 22 at a molecular level (BCR rearrangement). Thus, earlier studies indicating an absence of a Ph chromosome was not a valid measure of the normality of chromosome 22. The molecular abnormality in CML involving the ABL1 gene on chromosome 9 and the BCR gene on chromosome 22 has been established as being the proximate cause of the chronic phase of the disease (see “Molecular Pathology” below).

COEXISTENCE OF NORMAL STEM CELLS

Most, if not all, patients with CML have hematopoietic stem cells that, after treatment^80,81,82 or culture in vitro,^83,84,85 use of special cell isolation techniques,^81,82 or use of cell transfer to nonobese diabetic (NOD)/severe combined immunodeficiency (SCID) mice⁸⁸ do not have the Ph chromosome^89,90 or the BCR-ABL1 fusion gene.^{91,92,93,94,95} The switch to Ph chromosome–negative cells in vitro is associated with a loss of monoclonal glucose-6-phosphate dehydrogenase isoenzyme patterns, indicating the persistence and reemergence of normal polyclonal hematopoiesis rather than reversion to a Ph chromosome–negative clone.⁹⁶ In confirmation, BCR-ABL1+, CD34+, human leukocyte antigen (HLA)-DR− cells isolated from women with early phase CML are polyclonal using the human androgen receptor assay (HUMARA) to assess X chromosome inactivation patterns.⁹⁷ Very primitive hematopoietic cells, the long-term culture–initiating cells (LTC-ICs), are present in Ph chromosome–negative cytapheresis samples collected during early recovery after chemotherapy for CML.⁹⁸ These LTC-ICs are most commonly present when samples are collected within 3 months of diagnosis.⁹⁹ Variable levels of BCR-ABL1–negative progenitors are found in the CD34+DR− population, but low levels are found in the CD34+CD38− population.^95,100 Preprogenitors for the CD34+DR− cells are predominantly BCR-ABL1–negative in both marrow and blood at diagnosis.¹⁰¹ However, some cells with surface marker characteristics of very primitive normal hematopoietic cells do express the BCR-ABL1 gene.¹⁰² Both normal and leukemic SCID-repopulating cells coexist in the marrow and blood from CML patients in chronic phase, whereas only leukemic SCID-repopulating cells are detected in blast crisis.^103,104

PROGENITOR CELL CHARACTERISTICS

Progenitor Cell Dysfunction

The leukemic transformation resulting from the BCR-ABL1 fusion oncogene is maintained by a relatively small number of BCR-ABL1 stem cells that favor differentiation over self-renewal.¹⁰⁵ This predisposition to differentiation and progenitor cell expansion is mediated by an autocrine interleukin (IL)-3–granulocyte colony-stimulating factor (G-CSF) loop.¹⁰⁵ The earliest progenitors have the capacity to undergo marked expansion of erythroid, granulocytic, and megakaryocytic cell populations, and have a decreased sensitivity to regulation.^105,106,107 This expansion is especially dramatic in the more mature progenitor cell compartment.^105,108 The proliferative capacity of individual granulocytic progenitors is decreased compared to normal cells. Thus, the progenitor cell population in marrow and blood expands proportionately more than the increase in granulopoiesis.¹⁰⁹ BCR-ABL1 reduces growth factor dependence of progenitor cells.

Erythroid progenitors are expanded, erythroid precursor maturation is blocked at the basophilic erythroblast stage, and the extent of erythropoiesis is inversely proportional to the total white cell count.¹¹⁰

Progenitor Cell Characterization

Phenotypic differences of stem and progenitor cells in CML patients compared to normal subjects have been identified.¹¹¹ For example, a greater proportion of the circulating leukemic colony-forming unit–granulocyte-monocytes (CFU-GMs) express high levels of the adhesion receptor CD44¹¹² and low levels of L-selectin¹¹³ in contrast to normal cells. Leukemic CD34+ cells overexpress the P glycoprotein that determines the multidrug resistance phenotype.¹¹⁴

BCR-ABL1–positive progenitors survive less well in long-term culture than do their normal counterparts. Leukemic CFU-GM colonies, unlike normal colonies, decrease in long-term cultures that are deficient in KIT ligand,¹¹⁵ whereas their proliferation is favored in the presence of KIT ligand.¹¹⁶ Macrophage inflammatory protein (MIP)-1α, renamed CCL3, does not inhibit growth factor-mediated proliferation of CD34+ cells from CML patients, as it does CD34+ cells from normal subjects, even though the CCL3 receptor is expressed.¹¹⁷ Another chemokine, monocyte chemotactic protein (MCP)-1 or CCL2, unlike CCL3, is an endogenous chemokine that cooperates with transforming growth factor beta (TGF-β) to inhibit the cycling of primitive normal, but not CML, progenitors in long-term human marrow cultures.¹¹⁸ Leukemic progenitors are less sensitive than normal progenitors to the antiproliferative effects of TGF-β.¹¹⁹

Effects of BCR-ABL1 on Cell Adhesion

Primitive progenitors and blast colony-forming cells from patients with CML have decreased adherence to marrow stromal cells.¹²⁰ This defect is normalized if stromal cells are treated with interferon (IFN)-α.¹²¹ As a result, BCR-ABL1–negative progenitors are enriched in the adherent fraction of circulating CD34+ cells in chronic phase CML patients. The most primitive BCR-ABL1–positive cells in the blood of patients with CML differ from their normal counterparts. They are increased in frequency and are activated, such that signals that block cell mitosis are bypassed.¹²²

Ph chromosome–positive colony-forming cells adhere less to fibronectin (and to marrow stroma) than do their normal counterparts. Adhesion is fostered as a result of restoration of cooperation between activated β₁ integrins and the altered epitopes of CD44.^123,124 CML granulocytes have reduced and altered binding to P-selectin because of modification in the CD15 antigens.¹²⁵ BCR-ABL1–induced defects in integrin function may underlie the abnormal circulation and proliferation of progenitors^126,127 because growth signaling can occur through the fibronectin receptor.¹²⁸ IFN-α restores normal integrin-mediated inhibition of hematopoietic progenitor proliferation by the marrow microenvironment.¹²⁹ There are conflicting data regarding the effects of TKI effects on adhesion of CML cells to stroma.^130,131

BCR-ABL1–encoded fusion protein p210^BCR-ABL binds to actin, and several cytoskeletal proteins are thereby phosphorylated. The p210^BCR-ABL interacts with actin filaments through an actin-binding domain. BCR-ABL1 transfection is associated with increased spontaneous motility, membrane ruffling, formation of long actin extensions (filopodia), and accelerated rate of protrusion and retraction of pseudopodia on fibronectin-coated surfaces. IFN-α treatment slowly converts the abnormal motility phenotype of BCR-ABL1–transformed cells toward normal.¹³² Integrins regulate the c-ABL–encoded tyrosine kinase activity and its cytoplasmic nuclear transport.¹³³ The p210^BCR-ABL1 abrogates the anchorage requirement but not the growth factor requirement for proliferation.¹³⁴

In normal cells exposed to IL-3, paxillin tyrosine residues are phosphorylated. In cells transformed by p210^BCR-ABL1, the tyrosines of paxillin, vinculin, p125^FAK, talin, and tensin are constitutively phosphorylated. Pseudopodia enriched in focal adhesion proteins^134,135 are present in cells expressing p210^BCR-ABL1.

The sum of evidence suggests that defects in adhesion (contact and anchoring) of CML primitive cells remove them from their controlling signals normally received from microenvironmental cells via cytokine messages. These signals retain the balance among cell survival, cell death, cell proliferation, and cell differentiation. Inappropriate phosphorylation of cytoskeletal proteins, possibly independent of the mutant tyrosine kinase, is thought to be the key factor in disturbed integrin function of CML cells.

MOLECULAR PATHOLOGY

Ph Chromosome

The genetic disturbance became evident with the knowledge that CML was derived from a primitive cell containing a 22q− abnormality.^6,11 The abnormal chromosome contained only 60 percent of the DNA in other G-group chromosomes.¹³⁶ Cytogenetic analysis indicated the G-group chromosome involved was different from the extra G-group chromosome in Down syndrome, which had been assigned number 21. Thus, the former was assigned number 22—even though it proved to be slightly longer than the chromosome involved in Down syndrome.^11,137 The Paris Conference on Nomenclature decided not to undo the concept that Down syndrome is trisomy 21 and assigned the Ph chromosome and its normal counterpart, 22.¹³⁸ Using quinacrine (Q) and Giemsa (G) banding, Rowley¹² reported in 1973 that the material missing from chromosome 22 was not lost (deleted) from the cell, but was translocated to the distal portion of the long arm of chromosome 9. The amount of material translocated to chromosome 9 was approximately equivalent to that lost from chromosome 22, and the translocation was predicted to be balanced.¹² Moreover, the breaks were localized to band 34 on the long arm of chromosome 9 and band 11 on the long arm of chromosome 22. Therefore, the classic Ph chromosome is t(9;22)(q34;q11), abbreviated t(Ph) (Fig. 89–2). The Ph chromosome can develop on either the maternal or the paternal member of the pair.¹³⁹

Mutation of ABL1 and BCR Genes

Mutations of the ABL1 gene on chromosome 9 and of the BCR gene on chromosome 22 are central to the development of CML (Fig. 89–3).^140,141,142

In 1982, the human cellular homologue ABL1 of the transforming sequence of the Abelson murine leukemia virus was localized to human chromosome 9.¹⁴³ In 1983, ABL1 was shown to be on the segment of chromosome 9 that is translocated to chromosome 22¹⁴⁴ by demonstrating reaction to hybridization probes for ABL1 only in somatic cell hybrids of human CML cells containing 22q− but not those containing 9q+. v-abl is the viral oncogenic homologue of the normal cellular ABL1 gene. This gene (v-abl) can induce malignant transformation of cells in culture and can induce leukemia in susceptible mice.¹⁴⁵

The ABL1 gene is rearranged and amplified in cell lines from patients with CML.¹⁴⁶ Cell lines and fresh isolates of CML cells contain an abnormal, elongated 8-kb RNA transcript,^{147,148,149,150} which is transcribed from the new chimeric gene produced by the fusion of the 5′ portion of the BCR gene left on chromosome 22 with the 3′ portion of the ABL1 gene translocated from chromosome 9 (Fig. 89–4).¹⁴⁴ The fusion mRNA leads to the translation of a unique tyrosine phosphoprotein kinase of 210 kDa (p210^BCR-ABL), which can phosphorylate tyrosine residues on cellular proteins similar to the action of the v-abl protein product.^{151,152,153,154,155} The ABL1 locus contains at least two alleles, one having a 500-bp deletion.¹⁵⁷ In normal cells, the ABL1 protooncogene codes for a tyrosine kinase of molecular weight 145,000, which is translated only in trace quantities and lacks any in vitro kinase activity.¹⁵² The fusion product expressed by the BCR-ABL1 gene is hypothesized to lead to malignant transformation because of the abnormally regulated enzymatic activity of the chimeric tyrosine protein kinase.^{153,154,158,159} Construction of BCR-ABL1 fusion genes indicated that BCR sequences could also activate a microfilament-binding function, but the tyrosine kinase and microfilament-binding functions were not linked. Nevertheless, tyrosine kinase modification of actin filament function has been proposed as a step in leukemogenesis.¹⁶⁰

p210^BCR-ABL Fusion Protein

The breakpoints on chromosome 9 are not narrowly clustered, ranging from approximately 15 to more than 40 kb upstream from the most proximate region (first exon) of the ABL1 gene.^143,144,161 The breakpoints on chromosome 22 occur over a very short, approximately 5 to 6 kb, stretch of DNA referred to as the breakpoint cluster region (M-bcr),^162,163 which is part of a much longer BCR^164,165 gene (see Fig. 89–4). Three main BCRs have been characterized on chromosome 22: major (M-bcr), minor (m-bcr), and micro (μ-bcr). The three different breakpoints result in a p210, p190, and p230 fusion protein, respectively (see Fig. 89–3). The overwhelming majority of CML patients have a BCR-ABL1 fusion gene that encodes a fusion protein of 210 kDa (p210^BCR-ABL1), for which mRNA transcripts have e14a2 or a e13a2 fusion junction (see Fig. 89–3).¹⁶⁶ The “e” represents the BCR exon and “a” the ABL1 exon sites involved in the translocation. A BCR-ABL1 with an e1a2 type of junction has been identified in approximately 50 percent of the Ph chromosome–positive acute lymphoblastic leukemia cases and results in the production of a BCR-ABL1 protein of 190 kDa (p190^BCR-ABL). Almost all CML cases at diagnosis that encode a p210^BCR-ABL also express BCR-ABL transcripts for p190.¹⁶⁷ The biologic or clinical significance of these dual transcripts is not known. Transgenic mice expressing p210^BCR-ABL develop acute lymphoblastic leukemia in the founder mice, but all transgenic progeny have a myeloproliferative disorder resembling CML.¹⁶⁸

The BCR gene encodes a 160-kDa serine-threonine kinase, which, when it oligomerizes, autophosphorylates and transphosphorylates several protein substrates.¹⁶⁹ Aberrant methylation of the M-bcr in CML occurs.¹⁶⁶ The first exon sequences of the BCR gene potentiate the tyrosine kinase of ABL when they fuse as a result of the translocation.¹⁷⁰ The central portion of BCR has homology to DBL, a gene involved in the control of cell division after the S phase of the cell cycle. The C-terminus of BCR has a guanosine triphosphatase (GTPase)-activating protein for p21^rac, a member of the RAS family of guanosine triphosphate (GTP)-binding proteins.¹⁷¹ A reciprocal hybrid gene ABL-BCR1 is formed on chromosome 9q+ when BCR-ABL1 fuses on chromosome 22. The ABL-BCR1 fusion gene actively transcribes in most patients with CML.¹⁷²

Variations in breakpoints involving smaller stretches of chromosome 9 and rearrangements outside the M-bcr of chromosome 22 can occur.³⁷ In a few cases of CML with no evident elongation of chromosome 9, molecular probes have shown that ABL1 still is translocated to chromosome 22.¹⁷³ In occasional patients with Ph chromosome–positive CML, the break in chromosome 22 is outside the M-bcr, and transcription of a fusion RNA of the usual type fails or a fusion RNA is transcribed that does not hybridize with the classic M-bcr complementary DNA (cDNA) probe.¹⁷⁴

In cases in which the Ph chromosome is not found, BCR-ABL1 still may be located on chromosome 9 (a masked Ph chromosome).¹⁷⁵ The BCR gene can recombine with genomically distinct sites on band 11q13 in complex translocations in a region rich in Alu repeat elements.¹⁷⁶ ETV6/ABL1 fusion genes have also been found in BCR-ABL1–negative CML.¹⁷⁷

The BCR breakpoint site has been examined as a factor in disease prognosis. Some studies have shown no correlation between CML chronicity and breakpoint site, although thrombocytosis may be more common with 3′ breakpoint sites and basophilia with 5′ breakpoint sites.¹⁷⁸ No difference in response to IFN-α therapy was noted, and survival was not significantly different, although patients with 3′ deletions tended to have shorter survival.¹⁷⁹ Others have observed a better response to IFN-α in patients with a 3′ rearrangement, which is being examined with imatinib mesylate therapy.¹⁸⁰

CML patients with m-bcr breakpoints develop a blast crisis with monocytosis and an absence of splenomegaly and basophilia.¹⁸¹ The p230 (e19a2 RNA junction) encoded by μ-bcr is rarely expressed but has been associated with neutrophilic CML or thrombocytosis (see “Special Clinical Features” below). Other rare breakpoints have been described.¹⁸² For example, a case with a 12-bp insert between BCR and ABL1 resulted in a BCR-ABL1–negative (false-negative), Ph chromosome–positive CML with thrombocythemia.¹⁸³ Another novel BCR-ABL1 fusion gene (e6a2) in a patient with Ph chromosome–negative CML encoded an oncoprotein of 185 kDa.¹⁸⁴ Typical CML also has been associated with an e19a2 junction BCR-ABL1 transcript.¹⁸⁵

Experimental support for the hypothesis that p210^BCR-ABL1 tyrosine phosphoprotein kinase is transforming is provided by a retroviral gene transfer system that permits expression of the protein. Mouse marrow cells transfected with BCR-ABL1 develop clonal outgrowths of immature cells expressing the p210^BCR-ABL1 tyrosine kinase. Some clones progress to a malignant phenotype, can be transplanted, and can induce tumors in syngeneic mice.¹⁸⁶ Similar studies suggest that the p210^BCR-ABL can transform 3T3 murine fibroblasts if the gag gene sequence from a helper virus cooperates.¹⁸⁷ The BCR-ABL1 gene from a retroviral vector has been expressed in an IL-3–dependent cell line. Clones derived from the infected line transform over months to IL-3 independency, are capable of increased proliferation, and develop chromosomal abnormalities.¹⁸⁸

A series of mouse models in which the BCR-ABL1 was used to induce leukemogenesis have been described.^{189,190,191,192,193,194,195,196,197} Lethally irradiated mice have been reconstituted with marrow enriched for cycling stem cells infected with a BCR-ABL1–bearing retrovirus. Fatal diseases with abnormal accumulations of macrophagic, erythroid, mast, and lymphoid cells develop.¹⁸⁸ Classic CML did not occur, and complete transformation was not documented. The cell lines from spleen and marrow from mice with a BCR-ABL1 retrovirus infection were predominantly mast cells; however, in some cases these cell lines spontaneously switched to either erythroid and megakaryocytic, erythroid, or granulocytic lineages displaying maturation. They were transplantable (transformed) and contained the same proviral inserts as the original mast cell line.¹⁹⁸ Murine marrow also has been infected with a retrovirus encoding p210^BCR-ABL and transplanted into irradiated syngeneic recipients.¹⁸⁹ Although several types of hematologic malignancies developed, a syndrome mimicking human CML also occurred. Mice transgenic for a p190^BCR-ABL develop an acute lymphocytic leukemia (ALL) lymphoma syndrome¹⁹⁰ that resembles human Ph chromosome–positive ALL. When a p210^BCR-ABL transcript is introduced into a mouse germline (one-cell fertilized eggs), the p210 founder and progeny transgenic animals developed leukemia of B or T lymphoid or of myeloid origin after a relatively long latency period. In contrast, p190 transgenic mice exclusively developed leukemia of B-cell origin, with a relatively short period of latency. This finding was believed to be consistent with the apparent indolent nature of human CML during the chronic phase.¹⁹¹ When transgenic mice express p210^BCR-ABL, the transgenes develop ALL, whereas the progeny develop a myeloproliferative disorder.¹⁹²

Mouse models remain important for exploring the pathogenesis of the acute and chronic BCR-ABL1–mediated leukemias in vivo and in examining the potential effects of new drugs targeted at BCR-ABL1.¹⁹⁹

BCR-ABL IN HEALTHY SUBJECTS

BCR-ABL1 fusion genes can be found in the leukocytes of some normal individuals using a two-step reverse transcriptase polymerase chain reaction assay. Thus, although BCR-ABL1 may be expressed relatively frequently at very low levels in hematopoietic cells, only infrequently do the cells acquire the additional changes necessary to produce leukemia. This may be a dosage effect.²⁰⁰

BCR-ABL1 AND SIGNAL TRANSDUCTION

The tyrosine phosphoprotein kinase activity of p210^BCR-ABL1 has been causally linked to the development of Ph chromosome–positive leukemia in man.^201-212 p210^BCR-ABL1 is, unlike the ABL1 protein that is located principally in the nucleus, located in the cytoplasm making it accessible to a large number of interactions, especially components of signal transduction pathways.^205,206,213 It binds and/or phosphorylates more than 20 cellular proteins in its role as an oncoprotein.²⁰⁶ A subunit of phosphatidylinositol 3′-kinase (PI3K) associates with p210^BCR-ABL; this interaction is required for the proliferation of BCR-ABL1–dependent cell lines and primary CML cells. Wortmannin, a nonspecific inhibitor of the p110 subunit of the kinase, inhibits growth of these cells.²⁰⁷

The pathways and interactions invoked by BCR-ABL1 acting on mitogen-activated protein kinases are multiple and complex.^214,215 A RAF-encoded serine-threonine kinase activity is regulated by p210^BCR-ABL. Downregulation of RAF expression inhibits both BCR-ABL1–dependent growth of CML cells and growth factor–dependent proliferation of normal hematopoietic progenitors.²⁰⁸ The efficiency of cell transformation by BCR-ABL1 is affected by an adaptor protein that can relate tyrosine kinase signals to RAS. This involves growth factor receptor–bound protein-2 (GRB2). p210^BCR-ABL also activates multiple alternative pathways of RAS.²⁰⁹ PI3K is constitutively activated by BCR-ABL1, generates inositol lipids, and is dysregulated through the downregulation by BCR-ABL1 of polyinositol phosphate tumor suppressors, such as PTEN and SHIP1.²¹³ Figure 89–5 demonstrates interaction of p210^BCR-ABL with various mediators of signal transduction.

Reactive oxygen species are increased in BCR-ABL1–transformed cells and may act as a second messenger to modulate enzymes regulated by the reduction-oxidation (redox) equilibrium. An increase in these reactive oxygen products is postulated to play a role in the acquisition of additional mutations as a result of production of reactive oxygen species through the chronic phase, contributing to the progression to accelerated phase.^213,216

The adaptor molecule CRKL is a major in vivo substrate for p210^BCR-ABL, and it acts to relate p210^BCR-ABL to downstream effectors. CRKL is a linker protein that has homology to the v-crk oncogene product. Antibodies to CRKL can immunoprecipitate paxillin. Paxillin is a focal adhesion protein²¹⁰ that is phosphorylated by p210^BCR-ABL. The p210^BCR-ABL may be physically linked to paxillin by CRKL. CRKL binds to CBL, an oncogene product that induces B cell and myeloid leukemias in mice.²¹¹ The Src homology 3 domains of CRKL do not bind to CBL, but they do bind BCR-ABL. Therefore, CRKL mediates the oncogenic signal of BCR-ABL to CBL. The p120^CBL and the adaptor proteins CRKL and c-CRK also link c-abl, p190^BCR-ABL, and p210^BCR-ABL to the PI3K pathway.²¹² The p120^CBL also coprecipitates with the p85 subunit of PI3K, CRKL, and c-CRK. The p210^BCR-ABL may, therefore, induce the formation of multimeric complexes of signaling proteins.²¹⁷ These complexes contain paxillin and talin and may explain some of the adhesive defects of CML cells.²¹⁸

Hef2 also binds to CRKL in leukemic tissues of p190^BCR-ABL transgenic mice. Hef2 is involved in the integrin signaling pathway²¹⁹ and encodes a protein that accelerates GTP hydrolysis of RAS-encoded proteins and neurofibromin. The latter negatively regulates granulocyte-monocyte colony-stimulating factor (GM-CSF) signaling through RAS in hematopoietic cells.²²⁰ p62^DOK, a constitutively tyrosine-phosphorylated, p120^RAS GAP-associated protein, which is rapidly tyrosine phosphorylated upon activation of the c-kit receptor,²²¹ is also associated with ABL1.²²²

Nuclear factor (NF)-κB activation is also required for p210^BCR-ABL-mediated transformation.²²³ Expression of p210^BCR-ABL leads to activation of NF-κB–dependent transcription via nuclear translocation.²²⁴

Cell lines that express p210^BCR-ABL also demonstrate constitutive activation of Janus kinases (JAKs) and signal transducers and activators of transcription (STATs), usually STAT5.²²⁵ STAT5 is also activated in primary mouse marrow cells acutely transformed by the BCR-ABL1²²⁶; p210^BCR-ABL1 coimmunoprecipitates with and constitutively phosphorylates the common β subunit of the IL-3 and GM-CSF receptors and JAK2.²²⁷ Both ABL1 and BCR are also multifunctional regulators of the GTP-binding protein family Rho^228,229 and the growth factor-binding protein GRB2, which links tyrosine kinases to RAS and forms a complex with BCR-ABL1 and the nucleotide exchange factor Sos that leads to activation of RAS.²³⁰

The p210^BCR-ABL1 also activates Jun kinase and requires Jun for transformation.²³¹ In some CML cell lines, p210^BCR-ABL1 is associated with the retinoblastoma (Rb) protein.²³² Loss of the neurofibromatosis (NF1) tumor-suppressor gene, a RAS GTPase-activating protein, also is sufficient to produce a myeloproliferative neoplasm in mice akin to human CML resulting from RAS-mediated hypersensitivity to GM-CSF.²³³

EFFECTS OF BCR-ABL ON APOPTOSIS

Whether p210^BCR-ABL1 influences the expansion of the malignant clone in CML by inhibiting apoptosis is uncertain. In one study, the survival of normal and CML progenitors was the same after in vitro incubation in serum-deprived conditions and after treatment with X-irradiation or glucocorticoids.²³⁴ p210^BCR-ABL1 inhibits apoptosis by delaying the G₂/M transition of the cell cycle after DNA damage.²³⁵ The p210^BCR-ABL1 also may exert an antiapoptotic effect in factor-dependent hematopoietic cells.^236,237

p210^BCR-ABL1 does not prevent apoptotic death induced by human NK or lymphokine-activated killer cells directed against CML or normal cells.²³⁸ In accelerated and blast phases, apoptosis rates were lower in CML neutrophils. G-CSF and GM-CSF considerably decreased the rate of apoptosis in CML neutrophils.²³⁹

TELOMERE LENGTH

Patients with CML present with a somewhat shortened mean telomere length in granulocytic cells but not blood T lymphocytes at diagnosis, but considerable overlap exists in the distribution of telomere length with healthy individuals.^240,241,242 The rate of shortening of telomere length during the chronic phase is correlated with a more rapid onset of accelerated phase.^240,242 Telomerase reverse transcriptase (TERT) is the catalytic subunit, expression of which is closely correlated with telomerase activity. In CML CD34+ cells containing BCR-ABL1, the expression of TERT is significantly lower than in normal CD34+ cells, consistent with accelerated shortening of telomeres in CML cells.²⁴³ A further significant decrease in telomere length occurs in the accelerated phase of CML. Telomerase activity is increased in the accelerated phase.²⁴⁴ When therapy permits restoration of Ph chromosome–negative cells in the blood, these cells have telomere length comparable to that in matched healthy controls.²⁴⁵

SIGNS AND SYMPTOMS

In the 70 percent of patients who are symptomatic at diagnosis, the most frequent complaints include easy fatigability, loss of sense of well-being, decreased tolerance to exertion, anorexia, abdominal discomfort, early satiety (related to splenic enlargement), weight loss, and excessive sweating.^246,247,248 The symptoms are vague, nonspecific, and gradual in onset (weeks to months). A physical examination may detect pallor and splenomegaly. The latter was present in approximately 90 percent of patients at diagnosis, but with medical care being sought earlier, the presence of splenomegaly at the time of diagnosis is decreasing in frequency.²⁴⁷ Sternal tenderness, especially the lower portion, is common; occasionally, patients notice it themselves.

Uncommon presenting symptoms include those of dramatic hypermetabolism (night sweats, heat intolerance, weight loss) simulating thyrotoxicosis; acute gouty arthritis, presumably related in part to hyperuricemia; priapism, tinnitus, or stupor from the leukostasis associated with greatly exaggerated blood leukocyte count elevations^249,250,251; left upper quadrant and left shoulder pain as a consequence of splenic infarction and perisplenitis; vasopressin-responsive diabetes insipidus^252,253; and acne urticata associated with hyperhistaminemia.²⁵⁴ Acute febrile neutrophilic dermatosis (Sweet syndrome), a perivascular infiltrate of neutrophils in the dermis, can occur. In the latter situation, fever accompanied by painful maculonodular violaceous lesions on the trunk, arms, legs, and face are characteristic.^255,256 Spontaneous rupture of the spleen is a rare event.^257,258 Digital necrosis has been reported as a rare paraneoplastic event.^259,260

In an increasing proportion of patients, the disease is discovered, coincidentally, when blood cell counts are measured at a periodic medical examination.

CHILDHOOD PRESENTATION

Hyperleukocytosis and symptoms or signs therefrom are a more common feature in patients who present with CML before the age of 20 years. The white cell counts at diagnosis are on average more than twice that in adults, the fraction of blood blasts, promyelocytes, and myelocytes is significantly higher, and clinical manifestations of hyperleukocytosis are far more frequent in children than adults.²⁵⁰

LABORATORY FINDINGS

Blood

The presumptive diagnosis of CML can be made from the results of the blood cell counts and examination of the blood film.^26,246,247 The blood hemoglobin concentration is decreased in most patients at the time of diagnosis. Red cells usually are only slightly altered, with an increase in variation from small to large size and only occasional misshapen (elliptical or irregular) erythrocytes. Small numbers of nucleated red cells are commonly present. The reticulocyte count is normal or slightly elevated, but clinically significant hemolysis is rare.^246,261,262 Rare cases of mild erythrocytosis^263,264 or erythroid aplasia^265,266 have been documented.

The total leukocyte count is always elevated at the time of diagnosis and is nearly always greater than 25 × 10⁹/L; at least half the patients have total white counts greater than 100 × 10⁹/L (Fig. 89–6).^26,246,247 The total leukocyte count rises progressively in untreated patients. Rare patients may have dramatic cyclic variations in white cell counts as much as an order of magnitude with cycle intervals of approximately 60 days.^267,268 Granulocytes at all stages of development are present in the blood and are generally normal in appearance (Fig. 89–7). In this series, the mean blast cell prevalence was approximately 3 percent but can range from 0 to 10 percent; progranulocyte prevalence was approximately 4 percent; myelocytes, metamyelocytes, and bands accounted for approximately 40 percent; and segmented neutrophils accounted for approximately 35 percent of total leukocytes (Table 89–1). Often, there is a “myelocyte bulge” in which the differential count shows an exaggerated proportion of myelocytes compared to the proportion observed in normal persons. Hypersegmented neutrophils are commonly present.

Neutrophil alkaline phosphatase activity is low or absent in more than 90 percent of patients with CML.^269,270,271 The mRNA for alkaline phosphatase is undetectable in neutrophils of patients with CML.²⁷² The activity increases toward or to normal in the presence of intense inflammation or infection and when the total leukocytic count is decreased to or near normal with treatment.^271,273 CML neutrophils regain alkaline phosphatase activity after infusion into leukopenic recipients, suggesting the effect of regulators or factors extrinsic to the neutrophils. With the availability of specific markers, BCR-ABL1 in CML and JAK2 mutations in polycythemia, leukocyte alkaline phosphatase is no longer used for diagnostic purposes.

The proportion of eosinophils usually is not increased, but the absolute eosinophil count nearly always is increased. Rarely, eosinophils are so prominent that they dominate the granulocytic cells and lead to the designation Ph chromosome–positive eosinophilic CML. An absolute increase in the basophil concentration is present in almost all patients, and this finding can be useful in preliminary consideration of the differential diagnosis.^26,278 Basophilic progenitor cells are increased in the blood.²⁷⁹ The proportion of basophils usually is not greater than 10 to 15 percent during the chronic phase but may, in rare patients, represent 30 to 80 percent of the total leukocyte count during chronic phase and lead to the designation of Ph chromosome–positive basophilic CML.²⁸⁰ Flow cytometry using anti-CD203c provides very accurate assessment of the basophil frequency. Basophils may be hypogranulated or have an immature phenotype and may be left uncounted in an optical differential white cell count. Anti-CD203c recognizes these cells as basophils.²⁸¹ Granules of basophils in patients with CML, unlike normal basophils, contain mast cell α-tryptase.^281,282 Granulocytes containing both eosinophilic and basophilic granules (mixed granulation) are commonly present.²⁸³

The total absolute lymphocyte count is increased (mean: approximately 15 × 10⁹/L) in patients with CML at the time of diagnosis²⁸⁴ as a result of the balanced increase in T-helper and T-suppressor cells.²⁸⁵ B lymphocytes are not increased.²⁸⁸ T lymphocytes also are increased in the spleen.²⁸⁶ NK cell activity is defective in CML patients as a result of decreased maturation of these cells in vivo^287,288 and a decrease in the absolute number of circulating NK cells in patients with CML. The latter change can perhaps be related to increased apoptosis.²⁸⁹ The CD56 bright subset of NK cells is particularly decreased. These cells are reduced more as CML progresses, and they respond less to stimuli that recruit clonogenic NK cells compared to NK cells from normal subjects.²⁹⁰

The platelet count is elevated in approximately 50 percent of patients at the time of diagnosis and is normal in most of the rest.²⁹¹ The median value in patients at diagnosis is approximately 400 × 10⁹ cells/L. The platelet count may increase during the course of the chronic phase. Platelet counts greater than 1000 × 10⁹/L are not unusual, and platelet counts as high as 5000 to 7000 × 10⁹/L have occurred. Thrombohemorrhagic complications of thrombocytosis are infrequent. Occasionally, the platelet count may be below normal at the time of diagnosis, but this finding usually signals an impending progression to the accelerated phase of the disease (see “Accelerated Phase and Blast Crisis of Chronic Myelogenous Leukemia” below) and may also occur in with massive splenomegaly.

Functional abnormalities of neutrophils (adhesion, emigration, phagocytosis) are mild; are compensated for by high neutrophil concentrations; and do not predispose patients in chronic phase to infections by either usual or opportunistic organisms.^292,293,294 Platelet dysfunction can occur but is not associated with spontaneous or exaggerated bleeding. A decrease in the second wave of epinephrine-induced platelet aggregation is the most common abnormality and is associated with a deficiency of adenine nucleotides in the storage pool.^295,296

Marrow

Morphology The marrow is markedly hypercellular, and hematopoietic tissue takes up 75 to 90 percent of the marrow volume, with fat markedly reduced (see Fig. 89–7).^297,298 Granulopoiesis is dominant, with a granulocytic-to-erythroid ratio between 10:1 and 30:1, rather than the normal 2:1 to 4:1. Erythropoiesis usually is decreased, and megakaryocytes are normal or increased in number. Eosinophils and basophils may be increased, usually in proportion to their increase in the blood. Mitotic figures are increased in number. Mast cells are often seen, and uncommonly a juxtamembrane domain mutant of KIT coincides with BCR-ABL1 in CML.²⁹⁹ Rare reports of marrow mastocytosis have been explained by a KIT mutation as an additional genetic abnormality or by dual clones in the marrow.^300,301 Macrophages that mimic Gaucher cells in appearance are sometimes seen. This finding is a result of the inability of normal cellular glucocerebrosidase activity to degrade the increased glucocerebroside load associated with markedly increased cell turnover.³⁰² Macrophages also can become engorged with lipids, which, when oxidized and polymerized, yield ceroid pigment. This pigment imparts a granular and bluish cast to the cells after polychrome staining; such cells have been referred to as sea-blue histiocytes.³⁰²

Collagen type III (reticulin fibrosis), which takes the silver impregnation stain, is commonly increased at the time of diagnosis in nearly half the patients,³⁰³ and is correlated with the proportion of megakaryocytes in the marrow.^304,305 Increased fibrosis also is correlated with larger spleen size, more severe anemia, and a higher proportion of marrow and blood blast cells.

The marrows of CML patients have a mean doubling of microvessel density compared to healthy controls and have more angiogenesis in marrow than other forms of leukemia.^306,307,308 This increased marrow vascularity decreases to normal after treatment.³⁰⁹

Progenitor Cell Growth Cells that form colonies of neutrophils and macrophages or eosinophils (CFUs) are increased in the marrow and blood. The increase in CFUs in marrow is approximately 20-fold normal and in blood approximately 500-fold normal. The CFUs are of lighter buoyant density than those in normal marrow.¹⁰⁰ More primitive progenitors that can initiate long-term cultures of hematopoiesis also are markedly increased.³¹¹ Spontaneous blood-derived granulocyte-macrophage colony growth is common, although CFUs also respond to growth factor stimulation.

Cytogenetics The marrow and nucleated blood cells of more than 90 percent of patients with clinical and laboratory signs that fall within the criteria for the diagnosis of CML contain the Ph chromosome (22q−) as measured by G-banding, and virtually all patients have the t(9;22)(q34;q11)(BCR-ABL1) by FISH. The Ph chromosome is present in all blood cell lineages (erythroblasts, granulocytes, monocytes, megakaryocytes, T- and B-cell progenitors) but is not present in the majority of blood B lymphocytes or in most T lymphocytes.^54,56 Approximately 70 percent of patients in the chronic phase have the classic Ph chromosome in their cells.³¹² The remaining 20 percent also have a missing Y chromosome [t(Ph),−Y]; an additional C-group chromosome, usually number 8 [t(Ph),+8]; an additional chromosome 22q− but without the 9q+ [t(Ph), 22q−]; or t(Ph) plus either another stable translocation or another minor clone. These variations have not been shown to affect the duration of the chronic phase. Deletion of the Y chromosome occurs in approximately 10 percent of healthy men older than 60 years.^313,314

Variant Ph chromosome translocations occur in approximately 5 percent of subjects with CML and involve complex rearrangements (three chromosomes), and every chromosome except the Y chromosome can be involved.^{315,316,317,318,319} The Ph chromosome, that is, 22q−, is present, but the gross exchange of chromosomal material involves a chromosome other than 9 (simple variant) or involves exchange of material among chromosomes 9 and 22 and a third or more chromosomes (complex variant; Fig. 89–8). High-resolution techniques have indicated that 9q34-qter is transposed to 22q11 in simple and in complex translocations.^320,321 Thus, the fusion of 9q34 with 22q11 seems to occur in the cells of most patients with CML.³²³ Complex translocations involving chromosome 3 have been notable.^322,323,324 In rare cases, a reciprocal translocation with a chromosome other than 9 to chromosome 22 is larger than usual, and the posttranslocation shortening of the long arms of 22 is not apparent. This circumstance has been referred to as a masked Ph chromosome or masked translocation because the 22q− is not evident by microscopic examination,^325,326 although t(9;22) may occur as judged by banding techniques or molecular probes.³²⁷

Approximately 10 percent of patients have a deletion of the derivative 9 chromosome adjacent to the chromosome breakpoint. Although this deletion is thought to be an important factor in resistance to drug effects with IFN therapy, it does not appear to be significant with the use of imatinib.²¹⁴

Molecular Probes In a small proportion of patients with a clinical disease analogous to CML, cytogenetic studies do not disclose a classic, variant, or masked Ph chromosome. In these cases, use of a panel of restriction enzymes and Southern blot analyses with a molecular probe for the breakpoint cluster region on chromosome 22 nearly always detects rearrangement of fragments. This finding has led to the conclusion that almost all cases of CML have an abnormality of the long arm of chromosome number 22 (BCR rearrangement).^{328,329,330,331,332} Ph chromosome–negative CML cells with BCR rearrangement can express p210^BCR-ABL1, and such patients have a clinical course similar to Ph chromosome–positive CML.^{328,333,334,335,336}

The ability to identify the molecular consequences of the t(9;22), that is, BCR rearrangement, mRNA transcripts of the mutant fusion gene, and p210^BCR-ABL1, has resulted in diagnostic tests supplementary to cytogenetic analysis.³³² These tests include Southern blot analysis of BCR rearrangement,^{334,335,336,337,338} polymerase chain reaction (PCR) amplification of the abnormal mRNA,³³⁹ and a less complex variation on the latter, a hybridization protection assay.³⁴⁰

PCR can achieve a sensitivity of one positive cell in approximately 500,000 to one million cells. This extreme sensitivity requires special care in analysis and the inclusion of negative controls.^{341,342,343,344} Fusions e13a3, e14a3, and e19a2 are not detectable with standard PCR primers.^344A

A multicolor FISH method to detect the BCR-ABL1 fusion in patients with CML is a rapid and sensitive alternative to Southern blot and PCR-dependent methods.³⁴⁵ For diagnostic purposes, FISH is simple, accurate, and sensitive, and can detect the various molecular fusions (e.g., e13a2, e14a2, e1a2).^{346,347,348,349,350} Interphase FISH is faster and more sensitive than cytogenetics in identifying the Ph chromosome. If the concentration of CML cells is very low, interphase FISH may not detect BCR-ABL1, so it has limited use for detecting minimal residual disease.³⁵¹ Hypermetaphase FISH allows analysis of up to 500 metaphases per sample in 1 day. Several factors influence the false-positive and false-negative rates of FISH identification of BCR-ABL1, including definition of a fusion signal, nuclear size, and the genomic position of the ABL1 breakpoint.³⁵² Double BCR-ABL fusion signals (double-fusion [D]-FISH) have been proposed as being more accurate than the fusion signal used in dual color (single-fusion) S-FISH, because in the latter case a small percentage of the normal BCR and ABL1 signals overlap.³⁵³

The frequency of cytogenetic analysis can be reduced if patients are monitored by molecular methods such as competitive reverse transcriptase (RT)-PCR. Molecular analyses can be performed on blood samples and therefore are much easier to use than cytogenetic analysis of marrow cell metaphases. Quantitative RT-PCR is the method of choice for monitoring patients for residual disease or reappearance of disease after marrow transplantation and for following response to TKIs once routine cytogenetics and FISH are negative for the Ph chromosome. Competitive PCR can detect reappearance of or increasing levels of BCR-ABL1 RNA transcripts prior to clinical relapse in patients after transplantation.^354,355,356

Chemical Abnormalities

Uric Acid An increased production of uric acid with hyperuricemia and hyperuricosuria occurs in untreated CML.³⁵⁷ Uric acid excretion often is two to three times normal in patients with CML. If aggressive therapy leads to rapid cell lysis, excretion of the additional purine load may produce urinary tract blockage from uric acid precipitates. Formation of urinary urate stones is common in patients with CML, and some patients with latent gout may develop acute gouty arthritis or uric acid nephropathy.³⁵⁸ The likelihood of complications from urate overproduction is greatly increased by starvation, acidosis, renal disease, or diuretic drug therapy.

Serum Vitamin B₁₂–Binding Proteins and Vitamin B₁₂ Neutrophils contain vitamin B₁₂–binding proteins, including transcobalamins I and III (synonym: R-type B₁₂-binding protein or cobalophilin).^{359,360,361,362} Patients with myeloproliferative neoplasms have an increased serum level of vitamin B₁₂–binding capacity, and the source of the protein is principally mature neutrophilic granulocytes.^359,360 The increase in transcobalamin level and the resultant increase in vitamin B₁₂ concentration are particularly notable in CML, although any increase in the number of neutrophilic granulocytes, as in leukemoid reactions, can be accompanied by an increase in serum vitamin B₁₂–binding protein levels and vitamin B₁₂ concentration.³⁶² The serum vitamin B₁₂ level in CML patients is increased on average to more than 10 times normal.³⁶³ The increase is proportional to the total leukocyte count in untreated patients and falls toward normal levels with treatment, although increased vitamin B₁₂ levels commonly persist even after the white cell count is lowered to near normal with therapy.

Pernicious anemia and CML may rarely coexist. In this situation, the tissues are vitamin B₁₂ deficient, but the serum vitamin B₁₂ level may be normal because of the elevated level of transcobalamin I, a binder with a very high affinity for vitamin B₁₂.³⁶³

Whole Blood Histamine Mean histamine levels are markedly increased in patients in chronic phase (median: approximately 5000 ng/mL) compared to healthy individuals (median: approximately 50 ng/mL); and, this elevation is correlated with the blood basophil count.³⁶⁴ Cases of exaggerated basophilia and disabling pruritus, urticaria, and gastric hyperacidity have occurred, associated with enormous increases (several hundredfold) of blood histamine concentration.^365,366

Serum Lactic Dehydrogenase, Potassium, Calcium, and Cholesterol The level of serum lactic acid dehydrogenase (LDH) is elevated in CML.³⁶⁷ Pseudohyperkalemia resulting from the release of potassium from white cells during clotting³⁶⁸ and spurious hypoxemia or pseudohypoglycemia from in vitro utilization of oxygen or glucose by granulocytes can occur. Hypercalcemia³⁶⁹ or hypokalemia³⁷⁰ has occurred during the chronic phase of the disease, but such complications are very rare until the disorder transforms to acute leukemia. Elevated serum and urinary lysozyme levels are features of leukemia with greater monocytic components and are not features of CML.³⁷¹ Serum cholesterol is decreased in patients with CML.^372,373

Serum Angiogenic Factors Angiogenin, endoglin (CD105), vascular endothelial growth factor (VEGF), β-fibroblast growth factor, and hepatocyte growth factor are increased strikingly in the serum of CML patients.^{307,308,374,375}

BCR-ABL1–POSITIVE THROMBOCYTHEMIA

Either of two syndromes—thrombocythemia with the Ph chromosome and BCR-ABL1 rearrangement or thrombocythemia without a Ph chromosome but with the BCR-ABL1 rearrangement—may precede the overt signs of CML or its accelerated phase.^{376,377,378,379} In general, the disease closely mimics classic essential thrombocythemia initially: marked platelet elevation, extreme megakaryocytic hyperplasia, normal or mildly elevated white cell count, no or very slight myeloid immaturity in the blood, and minimal anemia. Minor bleeding, such as epistaxis, erythromelalgia, or signs of thrombosis, such as cerebral or limb ischemia, are occasionally present. In some cases, the absolute basophil count is mildly elevated. Approximately 5 percent of patients with apparent essential thrombocythemia have a Ph chromosome.³⁷⁶ In another study, two of 121 patients with essential thrombocythemia had BCR-ABL1 transcripts, and one of these patients also had a Ph chromosome in the marrow cells, whereas in a different study, four of 32 patients with thrombocythemia had low levels of BCR-ABL1 transcripts in blood cells. Approximately one in 20 patients with CML present with the features of essential thrombocythemia.^377,378 Evolution to blast crisis may occur.^380,381

NEUTROPHILIC CHRONIC MYELOGENOUS LEUKEMIA

A rare variant of BCR-ABL1–positive CML has been described in which the elevated white cell count is composed principally of mature neutrophils.^382,383 The white cell count is lower (on average: 30 to 50 × 10⁹/L) at the time of diagnosis than is the case with classic CML (median: 100 to 150 × 10⁹/L). Moreover, patients with neutrophilic CML usually do not have basophilia, notable myeloid immaturity in the blood, prominent splenomegaly, or low leukocyte alkaline phosphatase scores. The cells of these patients have the Ph chromosome but have an unusual BCR-ABL1 fusion gene in that the breakpoint in the BCR gene is between exons 19 and 20. This breakpoint location results in fusion of most of the BCR gene with ABL1 (e19a2 type BCR-ABL1), which leads to a larger fusion protein (230 kDa) compared to the fusion protein in classic CML (210 kDa; see Fig. 89–3). This correlation between genotype and phenotype has not been observed in all cases.³⁸⁴ This variant usually has an indolent course, which may be the result of very low levels of mRNA for p230 and the undetectable or barely detectable p230 protein in cells.³⁸⁵

MINOR-BCR BREAKPOINT–POSITIVE CHRONIC MYELOGENOUS LEUKEMIA

A small portion of patients with BCR-ABL1–positive CML have the breakpoint on the BCR gene in the first intron (m-bcr), resulting in a 190-kDa fusion protein instead of the classic 210-kDa protein observed in most patients with CML (see Fig. 89–3). The m-bcr molecular lesion is similar to that observed in approximately 60 percent of patients with BCR rearrangement-positive ALL. In patients with m-bcr CML, monocytes are more prominent, the white cell count is lower on average, and basophilia and splenomegaly are less prominent than in disease with classic BCR breakpoint (M-bcr). The few reported cases had a short interval before either myeloid or lymphoid blast transformation developed.^386,387

HYPERLEUKOCYTOSIS

Approximately 15 percent of patients present with symptoms or signs referable to leukostasis as a result of the intravascular flow-impeding effects of white cell counts greater than 300 × 10⁹/L.²⁴⁹ Hyperleukocytosis is more prevalent in children with Ph chromosome–positive CML.²⁵⁰ The effects of total leukocyte counts from 300 to 800 × 10⁹/L include impaired circulation of the lung, central nervous system, special sensory organs, and penis, resulting in some combination of tachypnea, dyspnea, cyanosis, dizziness, slurred speech, delirium, stupor, visual blurring, diplopia, retinal vein distention, retinal hemorrhages, papilledema, tinnitus, impaired hearing, and priapism.²⁵¹ In asymptomatic patients with hyperleukocytosis, initial treatment with hydration and hydroxyurea usually can be used to decrease the white cell count. Hydroxyurea treatment should be designed to accomplish a gradual decrease in white cell count over a few days so as to avoid the tumor lysis syndrome. If signs of hyperleukocytosis are present, hydration, leukapheresis, and hydroxyurea can be used simultaneously; hydroxyurea dose should be selected to avoid exaggerated tumor lysis.

CONCURRENCE OF LYMPHOID MALIGNANCIES

CML may emerge in patients with established chronic lymphocytic leukemia (CLL).^388,389,390 A few patients have presented with simultaneous occurrence of the two diseases.^391,392 A single case of lymphocytic leukemoid reaction simulating CLL that regressed as CML emerged has been reported.³⁹³ In some cases, the CLL lymphocytes did not contain the Ph chromosome, whereas the CML cells did, suggesting the presence of two independent clonal disorders.^{388,389,393,394} In other cases, the Ph chromosome was present in the myeloid and lymphoid cells, indicating a common origin.³⁹² Patients may present with Ph chromosome–positive acute lymphoblastic leukemia and, following chemotherapy-induced remission, develop the features of typical CML.³⁹⁵

DISEASES MIMICKING CHRONIC MYELOGENOUS LEUKEMIA

The diagnosis of CML is made based on the characteristic granulocytosis, white cell differential count, increased absolute basophil count, and splenomegaly coupled with the presence of the Ph chromosome or its variants (90 percent of patients) or a BCR rearrangement on chromosome 22 (>95 percent of patients).

Patients with other chronic hematopoietic stem cell diseases, such as polycythemia vera, essential thrombocythemia, or primary myelofibrosis, only occasionally have closely overlapping features. For example, the total white cell count is greater than 30 × 10⁹/L in more than 90 percent of patients with CML and increases inexorably over weeks or months of observation, whereas the total white cell count is less than 30 × 10⁹/L in more than 90 percent of patients with the three other classic chronic clonal myeloid diseases and usually does not change significantly over months to years. Polycythemia vera is associated with increased red cell mass and hemoglobin concentration and displays clinical signs of plethora; CML does not have these features. Patients with primary myelofibrosis invariably have marked teardrop poikilocytes and other severe red cell shape, size, and chromicity changes, as well as prominent nucleated red cells in the blood; CML rarely has these features. Patients with essential thrombocythemia have a platelet count greater than 450 × 10⁹/L and usually only mild neutrophilia (<20 × 10⁹/L); the slight neutrophilia distinguishes it from the proportion (approximately 25 percent) of CML patients with platelet counts greater than 450 × 10⁹/L, who at the time of diagnosis have white cell counts above 25 × 10⁹/L. In addition, patients with the clinical features of polycythemia vera or primary myelofibrosis do not have the Ph chromosome or BCR rearrangement in their blood and marrow cells, except in extremely rare cases. A very small proportion of patients with apparent essential thrombocythemia has BCR-ABL1 transcripts in their marrow and blood cells, and occasionally a Ph chromosome and may represent an atypical initial phase of CML (see “BCR-ABL1–Positive Thrombocythemia” above). The presence of a mutation in the JAK2 gene in more than 95 percent of patients with polycythemia vera is an important distinguishing feature (Chap. 84). The blood cells of approximately 50 percent of patients with primary myelofibrosis or essential thrombocythemia carry the JAK2 gene mutation and in those with primary myelofibrosis who do not, a significant proportion have a mutation in the calreticulin or the c-MPL gene (Chap. 86).

Increased awareness of the features of related disorders, such as chronic myelomonocytic leukemia (CMML) and chronic neutrophilic leukemia, and an appreciation that older patients are prone to atypical clonal myeloid diseases, have minimized the inappropriate diagnosis of Ph chromosome–negative CML, which should be avoided unless the clinical features are characteristic of classic CML and a masked Ph chromosome or BCR rearrangement is not found.

Reactive leukocytosis can occur with absolute neutrophil counts of 30 to 100 × 10⁹/L. Usually these leukemoid reactions occur in the setting of an overt inflammatory disease (e.g., pancreatitis), cancer (e.g., lung), or infection (e.g., pneumococcal pneumonia). If the incitant is not apparent, the absence of granulocytic immaturity, basophilia, or splenomegaly, and the absence of BCR/ABL1 in blood cells virtually eliminates classic CML as a consideration.

The precise diagnosis of CML is helpful in estimating the patient’s prognosis, identifying the utility of TKIs, and assessing the timing of special therapies, such as allogeneic hematopoietic stem cell transplantation.

Ph CHROMOSOME–POSITIVE CLONAL MYELOID DISEASES AND APLASTIC ANEMIA

The Ph chromosome has been found rarely in patients with apparent polycythemia vera,³⁹⁶ polycythemia vera that later evolves into Ph chromosome–positive CML,^397,398,399 primary myelofibrosis,^400,401 and myelodysplastic syndrome (MDS).^402,403 Molecular studies to determine the presence of the BCR-ABL1 were not performed in cases reported before 1985. Essential thrombocythemia with a Ph chromosome and/or BCR-ABL1 rearrangement in blood cells was discussed earlier (see “Special Clinical Features” above). Rare cases of aplastic anemia have presented with BCR-ABL1–positive cells or have evolved into BCR-ABL1 CML.^404,405

HYPERURICEMIA

Hyperuricemia and hyperuricosuria are frequent features of CML at diagnosis or in relapse.⁴⁰⁶ The need for treatment of hyperuricemia is a function of the elevated pretreatment serum uric acid concentration, blood white cell concentration, spleen size, and dose of cytolytic therapy planned. If these variables suggest a high risk for a significant amount of cell lysis, allopurinol 300 mg/day orally and adequate hydration to maintain a good urine flow should be instituted prior to therapy. Allopurinol is associated with a high frequency of allergic skin reactions and should be discontinued after the blood leukocyte count and spleen size have decreased and the risk of exaggerated cell lysis has passed. If hyperuricemia is extreme, usually over 9 mg/dL, rasburicase can be administered.⁴⁰⁷ Rasburicase is a recombinant urate oxidase that converts uric acid to allantoin. Rasburicase, unlike allopurinol, reduces the uric acid pool very rapidly, does not result in the accumulation of xanthine or hypoxanthine, and does not require alkalinization of urine facilitating phosphate excretion.⁴⁰⁸ Although the manufacturer recommends a dose every day for 5 days, several reports have indicated that one injection will produce a rapid and sustained decrease in serum uric acid, significantly decreasing the cost of therapy.⁴⁰⁹ Another alternative is to use allopurinol for a few days after one injection of rasburicase. A dose of 0.2 mg/kg of ideal body weight of rasburicase intravenously has been used.⁴¹⁰

INITIAL CYTOREDUCTION THERAPY

A TKI is now used as initial therapy in patients with CML. In cases where the white cell count is markedly elevated, hydroxyurea can be used prior to or in conjunction with a TKI. If rapid cytoreduction is required because of signs of the hyperleukocytic syndrome, leukapheresis and hydroxyurea often are combined.

Leukapheresis

Leukapheresis can control CML only temporarily. For this reason, it is rarely used in chronic phase CML and is useful in only two types of patients: the hyperleukocytic patient in whom rapid cytoreduction can reverse symptoms and signs of leukostasis (e.g., stupor, hypoxia, tinnitus, papilledema, priapism),^249,250,251 and in the pregnant patient with CML who can be controlled by leukapheresis treatment without other therapy either during the early months of pregnancy when therapy poses a higher risk to the fetus or, in some cases, throughout the pregnancy.^411,412 Because of the large body burden of leukocytes in marrow, blood, and spleen, and the high proliferative rate in CML, leukocyte reduction by apheresis is less efficient than in other types of leukemia.^249,251 Leukapheresis reduces the burden of tumor cells subject to chemotherapeutically induced cytolysis and thus the production and the excretion of uric acid. In hyperleukocytic nonpregnant patients, leukapheresis is best used in conjunction with hydroxyurea to ensure rapid and optimal reduction in white cell count.

Hydroxyurea

Hydroxyurea 1 to 6 g/day orally, depending on the height of the white cell count, can be used to initiate elective therapy.⁴¹³ Urgent treatment of extraordinary total white cell counts may require higher doses. The dose of hydroxyurea should be decreased as the total white cell count decreases and usually is given at 1 to 2 g/day when the total white cell count reaches 20 × 10⁹/L. The drug should be temporarily discontinued if the white cell count drops below 5 × 10⁹/L. If hydroxyurea is being used in combination with a TKI, it is usually tapered and discontinued once a hematologic response to the TKI is observed.

Anagrelide

Anagrelide can be used for platelet reduction in patients who present with elevated platelet counts. This agent acts directly to decrease megakaryocyte mass, and it can lead to a precipitous fall in platelet counts. In occasional patients who still have significant thrombocythemia after a TKI is initiated, combination with anagrelide is associated with a normalization of platelet counts.⁴¹⁴

INITIAL THERAPY WITH A TYROSINE KINASE INHIBITOR

Imatinib mesylate (imatinib) was the first TKI developed, and it was approved by the FDA for initial therapy of CML in 2002. Subsequently, two second-generation TKIs, nilotinib and dasatinib, were approved for initial therapy in 2010. This approval was based upon superior cytogenetic and molecular response rates with nilotinib and dasatinib at benchmark time points and lower rates of conversion to accelerated or blast phase as compared with imatinib. Thus far, an overall survival advantage of dasatinib or nilotinib compared with imatinib has not been shown.⁴¹⁵ Third-generation TKIs, bosutinib and ponatinib, are under study. Table 89–2 compares these inhibitors.

Imatinib Patients with newly diagnosed, chronic phase CML can be started on imatinib, 400 mg/day by mouth. The goal of imatinib therapy is to decrease the cells bearing the t(9;22) translocation (leukemic cells) to the lowest levels possible, during which process normal (polyclonal) hematopoiesis is restored. The efficacy of imatinib is judged by measuring three benchmarks: hematologic response, cytogenetic response, and molecular response as defined in Table 89–3.^416,417 These benchmarks are used to determine its maximal therapeutic effect. The time to achieve a maximal effect is variable, but as long as a patient is having a continued reduction in the size of the leukemic clone as judged by cytogenetic or PCR measurements, and has met response benchmarks, the drug is continued at 400 mg/day. If the patient stops responding before a complete cytogenetic remission or complete molecular remission is achieved, the dose can be increased to 600 mg/day or to 800 mg/day (400 mg every 12 hours), if tolerated. Alternatively, another TKI can be used. About two-thirds of patients who do not have a significant hematologic response or who relapse while receiving imatinib at a dose of 400 mg/day achieve a complete or partial hematologic response with higher doses, but few cytogenetic responses occur.⁴¹⁸ Some patients without a cytogenetic response can enter a partial or complete cytogenetic response(CCyR) with higher doses of imatinib. Unfortunately, the responses to higher doses of imatinib in patients lacking a hematologic or cytogenetic response at 400 mg/day usually are transient.^419,420

Several studies have examined use of imatinib at doses higher than 400 mg/day. Patients with newly diagnosed chronic phase CML treated with imatinib, 800 mg/day, administered in two 400-mg doses, every 12 hours, had a frequency of 90 percent CCyR, and 96 percent had at least a major cytogenetic response (MCyR). At a median of 15 months, no patients had progressed and 63 percent showed blood BCR-ABL1/ABL1 percentage ratios of less than 0.05 percent. Twenty-eight percent of patients had undetectable BCR-ABL1 blood levels.⁴²¹ In one trial, major molecular remission (MMR) at 12 and 24 months was higher in those receiving doses of imatinib greater than 600 mg/day.⁴²² In another trial, patients receiving 400 mg twice per day had a MCyR of 90 and 96 percent at 12 and 18 months, respectively. MMR rates were 48 percent and 54 percent at 6 and 12 months, respectively. These results compared favorably to historical data in the IRIS (International Randomized Study of Interferon) trial studying 400 mg/day of imatinib, but myelosuppression, rash, fatigue, and musculoskeletal symptoms were greater with these higher doses. Responses were, also, more rapid with the higher doses. More edema, gastrointestinal symptoms, rash, and myelosuppression occurred at the higher doses.⁴²³ Despite these reports the current starting dose is customarily 400 mg/day, balancing both effectiveness and tolerability in newly diagnosed patients. Moreover, the more rapid response with higher doses of imatinib may not translate into better long-term results.^424,425 For example, in another trial, MMR and CCyR at 12 months were not significantly different between standard and high-dose patients, although patients in higher-risk categories based on Sokal scores, fared better with high-dose imatinib.⁴²⁴ (See “Course and Prognosis” below for an explanation of the Sokal Score.)

Doses of imatinib lower than 400 mg/day result in fewer CCyR and a shorter duration of that response. Patients who are older and who have lower body weight may only tolerate a lower dose, but they are less likely to achieve a CCyR.⁴²⁶ If however, a patient is on a lower dose (e.g., 300 mg/day) for a special reason (body size or tolerance level) and achieves a complete hematologic response (CHR) and CCyR within 12 months of onset of therapy, acceptable outcomes without excess toxicity can result.⁴²⁷

Some patients have been followed for up to 8 years on imatinib in the IRIS trial (IFN vs. STI571).⁴²⁸ With median followup of 60 months, the best observed MCyR and CCyR rates were 89 percent and 82 percent, respectively. Only 7 percent had progressed to accelerated or blast phase, and overall survival rate was 89 percent. The best MMR rate was 86 percent during the 8-year followup. No patient with an MMR at 12 months progressed to accelerated or blast phase. By 8 years of follow up on the IRIS trial, 22 percent of patients had discontinued imatinib treatment because of an unsatisfactory response or toxicity, and only 55 percent remained on study. The 5-year probability of remaining in MCyR while receiving imatinib was approximately 60 percent. Achieving a CCyR correlated with progression-free survival, but achieving a MMR conferred no further survival benefit.⁴²⁹

Use of Imatinib in Patients with Variant Chromosomal Translocations or Breakpoints

Patients with variant Ph chromosome translocations have a similar prognosis to that of patients with classic Ph chromosome translocations who are treated with imatinib.⁴³⁰ (See Fig. 89–3 for a diagram of breakpoints.) Patients with the e13a2, p210^BCR-ABL translocation respond well to imatinib, with similar rates of complete cytogenetic remission.⁴³¹ The e13a2 transcript may be more sensitive to imatinib than the e14a2 transcript.⁴³² In a patient with both e1a2 and e14a2 fusion transcripts, only the p210 e14a2 transcript disappeared, whereas the e1a2 transcript persisted during progression to blast phase. No mutation in the kinase domain of ABL1 was found.⁴³³ Thus, different clones in a patient may have a different sensitivity to imatinib. Deletions of the derivative chromosome 9 do not influence the response and outcomes in CML chronic phase when using imatinib.⁴³⁴

Response to Imatinib in Children and Older Patients

More than 80 percent of children with chronic phase CML who are treated with imatinib, 260 to 570 mg/m², enter a complete cytogenetic remission. Imatinib is now approved for use in pediatric patients. Weight gain is the most common side effect of imatinib.⁴³⁵ In patients who were older than age 60 years, similar cytogenetic response rates and survival rates were noted as in younger patients in the late chronic phase who were treated concurrently, suggesting that age is not usually a factor in response.^436,437

Side Effects and Special Treatment Considerations

Imatinib is usually tolerated. Most adverse effects are manageable and seldom require permanent cessation of therapy. Reduction to subtherapeutic doses is not recommended; it is better to interrupt therapy for a time.⁴³⁸

Myelosuppression is common, especially at treatment onset when the CML clone accounts for most of the blood cells. Dose reduction to less than 300 mg/day is not advisable for myelosuppression. The drug should be stopped until blood counts recover. G-CSF or GM-CSF can prevent or treat neutropenia.^439,440 Platelet transfusion may be used for severe thrombocytopenia. Patients with imatinib-induced chronic cytopenias have inferior responses.⁴⁴¹ Myelosuppression is an independent adverse factor for achieving cytogenetic responses with imatinib.⁴⁴² Erythropoiesis-stimulating agents may be used to raise hemoglobin levels, and their use does not appear to affect CML outcomes, but may increase the risk of thrombosis.⁴⁴³ Severe irreversible marrow aplasias after imatinib exposure can occur.⁴⁴⁴

The main side effects noted with imatinib include fatigue, edema, nausea, diarrhea, muscle cramps, and rash.⁴⁴⁵ Elevated hepatic transaminases can occur. Mild transaminase elevations often respond to glucocorticoid use.⁴⁴⁶ Hepatotoxicity is uncommon, occurring in approximately 3 percent of patients, usually within 6 months of onset of imatinib use. Acute liver failure has been described.⁴⁴⁷ The severe periorbital edema occasionally observed is postulated to be a drug effect on the function of platelet-derived growth factor receptor (PDGFR) and KIT expressed by dermal dendrocytes. Surgical decompression of severe edema rarely has been required.⁴⁴⁸ Although no effects on spermatogenesis have been reported, women of childbearing age are at risk of teratogenic effects on their fetus.⁴⁴⁸

Weight gain is associated with imatinib use.⁴⁴⁹ Patients with renal impairment require lower doses of imatinib.⁴⁵⁰ Hypophosphatemia⁴⁵¹ and altered bone and mineral metabolism have occurred.^452,453 Cutaneous reactions with imatinib therapy occur in approximately 15 percent of patients.⁴⁵⁴ Except for severe reactions (approximately 5 percent of patients), such as Stevens-Johnson syndrome, exfoliative dermatitis, and erythema multiforme, cutaneous reactions rarely require permanent discontinuation of therapy. With milder reactions, concomitant glucocorticoid therapy or brief discontinuation of imatinib with gradual reintroduction at a lower dose and then a gradual increase in dose can be accomplished.^456,457 With very mild cases, concurrent treatment with antihistamine or other symptomatic therapy may be successful. Oral desensitization regimens have been described that allow some patients to continue imatinib therapy. Hair depigmentation⁴⁵⁸ and hypopigmentation of the skin,⁴⁵⁹ probably related to the inhibition of the KIT receptor tyrosine kinase by imatinib, have been reported.

Other Effects of Imatinib

Imatinib has been found to cause regression of marrow fibrosis.⁴⁶⁰ One study found that the extent of marrow fibrosis in CML is not a prognostic factor with imatinib therapy,⁴⁶¹ whereas another study observed that although imatinib reverses marrow fibrosis in patients with CML, it does not change the unfavorable prognosis associated with fibrosis.⁴⁶²

Imatinib reverses exaggerated VEGF secretion in patients with CML,⁴⁶³ and it may reverse exaggerated marrow angiogenesis.⁴⁶⁴ It can reduce marrow cellularity and normalize morphologic features regardless of cytogenetic response. BCR-ABL1–positive cells persist in patients despite prolonged treatment responses with imatinib.⁴⁶⁵

Pharmacokinetic Considerations During Imatinib Therapy

Mean plasma trough concentration of imatinib and its metabolite, CGP74588, obtained at about 1 month (presumptive steady-state) was 979 ± 530 ng/mL. The rate of CCyR and MMR was higher within the highest quartiles of imatinib trough levels.⁴⁶⁶ Some therapists suggest that imatinib plasma levels be checked in cases of suboptimal response in order to adjust the dose, but access to this monitoring is not routinely available.⁴⁶⁷ Comedications and population covariates, such as body weight and white cell count, had no, or minimal, effect on imatinib clearance.⁴⁶⁸ Patients with CML on hemodialysis have been successfully treated with imatinib.⁴⁶⁹ Therapy interruptions and nonadherence with oral imatinib usage are common, and patient education and close monitoring are important to ensure compliance.⁴⁷⁰

Initiation of Therapy with Newer Tyrosine Kinase Inhibitors

Dasatinib Dasatinib is a second-generation oral BCR-ABL1 inhibitor with dual inhibition of ABL1 and SRC.⁴⁷¹ It can bind to both the active and inactive conformation of the ABL1 kinase domain, so it may be affected by mutations resulting in resistance.⁴⁷¹

Dasatinib was first studied for use in initial therapy in a phase II trial that accrued 62 patients. Ninety-eight percent achieved a CCyR, and the median time to CCyR was 3 months. The MMR rate was 82 percent. Responses were durable, and the recommended treatment schedule based on a safety profile was 100 mg, once daily.⁴⁷² A randomized, phase III trial compared the efficacy of dasatinib to imatinib.⁴⁷³ In this study, 259 patients received dasatinib, 100 mg/day, and 260 received imatinib, 400 mg/day. After 12 months of followup, the rates of CCyR by 3, 6, and 9 months were 54, 73, and 78 percent, respectively, for patients on dasatinib as compared to 31, 59, and 67 percent, respectively, for those on imatinib. The 24-month CCyR was 80 percent for patients using dasatinib, as compared to 74 percent for those on imatinib. The MMR showed a similar trend, and the median time to MMR was 15 months for those using dasatinib, compared to 36 months for those using imatinib.

Long-term followup data have confirmed faster and deeper responses to dasatinib as compared to imatinib.⁴⁷⁴ At 4 years, 76 percent of dasatinib-treated patients had attained a MMR (BCR/ABL1 <0.1 percent) as compared to 63 percent using imatinib. At 3, 6, and 12 months, more patients achieved molecular responses using dasatinib than those using imatinib. There were fewer patients who progressed to accelerated or blast phase using dasatinib as compared to imatinib. To date, there is no difference in progression-free survival or overall survival between the two groups.⁴⁷⁴ In a randomized study, with a minimal followup of 3 years, the proportion of patients with BCR-ABL1 transcript levels less than 10 percent was higher in those using dasatinib as compared to those using imatinib.⁴⁷⁵ Better responses were observed at 3, 6, and 12 months in the patients using dasatinib. The achievement of an early molecular response was predictive of improved progression-free and overall survival.⁴⁷⁶

Toxicity of Dasatinib Most grade 3 or 4 adverse events with dasatinib are hematologic and all cell lines can be affected. Some patients may have bleeding from inhibition of platelet aggregation.⁴⁷⁷ Adverse events noted less frequently with dasatinib than imatinib include nausea, vomiting, myalgia, rash, and fluid retention, including superficial edema. Pleural effusions can be seen with dasatinib use. In one trial, 14 percent of patients had grade 1 or 2 pleural effusions at 24 months, but grade 3 or 4 effusions occurred in only 2 patients. This toxicity did not affect drug efficacy. Dasatinib may also increase the risk of pulmonary arterial hypertension at any time, and this is an indication to discontinue dasatinib.⁴⁷⁸ Dasatinib may prolong the QTc interval, and it should be used with caution in those who have long QT syndrome or those taking drugs that may lengthen the QT interval.⁴⁷⁴ Hypophosphatemia was found in 7 percent of cases.⁴⁷⁴

Dasatinib is metabolized primarily by hepatic cytochrome P450 (CYP) 3A4 enzymes, so inducers of this enzyme may decrease the effective dose, and inhibitors may increase the effective dose. Increases or decreases in administered dose may be needed to compensate for these effects. Antacids can also reduce dasatinib effects.⁴⁷⁹ Lymphocytosis from the clonal expansion of NK/T cells has occurred during dasatinib treatment.⁴⁸⁰ In a population of dasatinib-treated patients with large granular lymphocyte expansion, 90 percent had T-cell receptor delta rearrangements, the functional significance of which is unknown.⁴⁸¹ Lymph node follicular hyperplasia has been noted on dasatinib therapy.⁴⁸² Unlike the case with imatinib, dasatinib cellular uptake is not affected by octamer-binding protein-1 (OCT-1) activity, which is a substrate of the efflux proteins, ABCB1 and ABCG2. Resistance to dasatinib is often found with point mutations in ABL1 at residue 315 or 317.

Nilotinib Unlike dasatinib, nilotinib is a selective, orally bioavailable, ATP-competitive inhibitor of BCR-ABL1 which is 20 to 50 times more potent than imatinib in vitro.⁴⁸³ Like imatinib, it does not induce apoptosis in CD34+ CML cells.⁴⁸⁴ As with dasatinib, nilotinib was first tested as initial therapy in phase II trials,⁴⁸⁵ which were followed by a randomized phase III trial. In one phase II trial, 51 patients received nilotinib 400 mg, twice a day, and 98 percent entered CCyR and 76 percent had a MMR by 6 months.⁴⁸⁶ The phase III trial compared nilotinib, 300 mg twice daily, 400 mg twice daily, and imatinib 400 mg daily.⁴⁸⁷ At 12 months, the MMR which had been chosen as the primary end point, was 44 percent (nilotinib 300 mg dose), 43 percent (nilotinib 400 mg dose), and 22 percent (imatinib 400 mg daily). The CCyR rates were 15 percent higher with nilotinib than imatinib. The rate of progression to accelerated or blast phase was 4 percent at 1 year with imatinib and less than 1 percent with nilotinib. These improvements were observed in each prognostic group based on Sokal risk groups. The patients using either 300 or 400 mg doses had minimal differences, so in 2010, nilotinib was approved at a dose of 300 mg twice daily for initial CML therapy. At 4 years of followup, more patients using either dose of nilotinib achieved a MMR than those using imatinib (73 and 70 percent vs. 53 percent).⁴⁸⁸ The rates of progression to accelerated and blast phase were also lower for nilotinib than imatinib. The 4-year freedom-from-progression and overall-survival rates were not different between the two groups. MMR rates at 3 years were higher for those patients using nilotinib, 300 mg, twice per day, in the three risk groups of Sokal (low risk, 79 percent; intermediate risk, 76 percent; and high risk, 52 percent progression as compared to those using imatinib (65, 55, and 30 percent progression, respectively).⁴⁸⁸ There was a reduced incidence of BCR-ABL1 mutations in in patients using nilotinib compared to those using imatinib. Nilotinib use led to fewer (less than half as many) treatment-emergent BCR-ABL1 mutations than did imatinib treatment, and to reduced rates of progression to accelerated phase and blast crisis in patients with these mutations.⁴⁸⁹

Toxicity of Nilotinib Nilotinib is rarely associated with edema or muscle cramps. Grades 3 and 4 cytopenias were seen in 29 percent of cases in one trial.⁴⁸⁷ Grade 3 or 4 elevations in lipase, bilirubin, and hyperglycemia were observed in 17, 8, and 12 percent of patients, and hypophosphatemia was seen in 16 percent. QTc prolongation can occur, so one should monitor the electrocardiogram (EKG) readings for 7 days after starting therapy and with dose changes.⁴⁸³ Electrolyte abnormalities should be corrected at outset of treatment. Nilotinib may be associated with an increased risk of peripheral vascular disease, which may be arterial or venous.⁴⁹⁰ If thrombosis occurs, it should no longer be used for therapy.

Bosutinib and Ponatinib These TKIs are not approved for use in initial therapy. In the one trial, which compared efficacy for bosutinib, 500 mg once daily, with imatinib, 400 mg daily in newly diagnosed chronic phase patients, the primary end point of CCyR at 12 months was 70 percent for bosutinib and 63 percent for imatinib.^491,492 Results from followup of MMR rates, transformation rates, and durability of remission are not yet available.

Summary of Tyrosine Kinase Inhibitor Selection for Initial Therapy of Chronic Phase Chronic Myelogenous Leukemia

The goal of initial TKI therapy is to achieve a CCyR within 12 months or no later than 18 months of therapy, and to prevent progression to the accelerated or blast phase. How best to achieve these goals remains controversial. Hence, the National Comprehensive Cancer Network (NCCN) guidelines list imatinib, nilotinib, and dasatinib as all being acceptable TKIs for initial treatment of CML.⁴⁹² Many clinicians would choose a second-generation TKI, given the rapidity and depth of response and the lower rates of transformation to advanced phases of the disease, but others use imatinib because of the lack of proof of prolongation of survival with nilotinib or dasatinib. In those with intermediate- or high-risk disease as assessed by the Sokal and Hasford models (see “Course and Prognosis” below for details of these scores), nilotinib or dasatinib may be preferred over imatinib to achieve rapid, better responses—the “hit hard, hit early” approach.⁴⁹³ Thus, until survival data are available, any one of the three approved TKIs may be used for initial therapy. Choice of agent may be dependent upon cost considerations, ease of administration, patient risk scores or perceived risk,⁴⁹⁴ and the drug’s side-effect profile (see Table 89–2).

Defining a Response to Tyrosine Kinase Inhibitors

Table 89–3 contains definitions of hematologic, cytogenetic, and molecular responses. The guidelines for periodic monitoring patients who are in chronic phase and receiving TKI therapy are shown in Table 89–4. The median BCR-ABL1 levels for imatinib-treated patients can decrease over at least 5 years. Table 89–5⁴⁹⁵ lists the milestones at 3, 6, 12, and 18 months expected of patients as indicators of an appropriate response in patients treated initially with a TKI.⁴³⁴ There is variation in an individual patient’s time to maximal response. Consequently, if a patient has not met those precise milestones but shows a continued decrease in the proportion of Ph chromosome–positive cells on cytogenetic examination of marrow, or if in a CCyR, a continued decrease in the level of the PCR signal for BCR-ABL1 and is near the benchmark, the treatment can be continued. Only (1) failure to meet the benchmarks at 3, 6, or 12 months or (2) loss of response as defined as loss of a CHR or CCyR, (3) development of new cytogenetic abnormalities, (4) acquisition of a BCR-ABL1 mutation, or (5) an increase in the BCR-ABL1/ABL1 ratio of 1-log or more on serial RT-PCR testing or into the range associated with reappearance of the Ph chromosome on G-banding should generate a change in treatment to limit the risk of progression of the disease. Because of the variability in PCR testing, those changes should be confirmed within 1 month. Patients who have 100 percent Ph chromosome–positive cells after 6 months of therapy have a minimal chance of achieving a MCyR or CCyR and may be offered allogeneic stem cell transplantation, if applicable.^417,496

Achieving a cytogenetic response is associated with progression-free survival in patients treated with imatinib and is an important goal of therapy (97 percent in those with a CCyR vs. 81 percent in those without a MCyR).⁴⁹⁷ At 5 years of followup, of those patients who achieved CCyR on imatinib in the IRIS study, only 3 percent had progressed to accelerated or blast phase during treatment.⁴²⁸ A CCyR at 1 year may be the major predictor for overall survival and progression-free survival.⁴⁹⁸ In patients in chronic phase treated with imatinib, nilotinib, or dasatinib, early responses (at 3 months) in BCR-ABL1 transcript reduction or decrease of Ph chromosome frequency predicted for better outcomes as measured by event-free or overall survival. For example, patients with less than 10 percent BCR-ABL1 transcripts at 3 months had a 3-year event-free survival of 95 percent or greater, whereas those with greater than 10 percent transcript level at 3 months had a 61 percent event-free survival.⁴⁹⁹

Molecular response is determined by the decrease in BCR-ABL1 mRNA by PCR. This is the only means to measure the depth of the response once a CCyR is attained. The achievement of MMR after treatment with imatinib is associated with durable long-term CCyR and a lower rate of disease progression. Only 5 percent of patients achieving a MMR with imatinib lost a CCyR compared to 37 percent who did not achieve that degree of response.⁵⁰⁰ Also, the 5-year followup of the IRIS trial showed that no patient with a CCyR and MMR at 12 months had progressed to a more advanced phase of the disease.⁴⁹⁷ The IRIS study 7-year followup also showed that in those with a MMR at that point in time, progression was rare. The estimated event-free survival was 95 percent for those with MMR at 18 months compared with 86 percent in those without a MMR.⁵⁰¹ To date, there is no evidence that a change of therapy would improve survival in those with CCyR but not a MMR. In those with stable CCyR after treatment with a second-generation TKI, achievement of MMR may not have significance as a predictor of survival.⁴⁹⁹

The time taken to achieve MMR is also thought to have prognostic significance. In the IRIS study, the chance of disease progression was higher in those who failed to achieve a 1-log reduction in BCR/ABL1 transcripts by 3 months or a 2-log reduction by 6 months.⁵⁰² A BCR-ABL1 transcript level greater than 10 percent after 3 months is a significant predictor for long-term outcomes.⁵⁰³ In another analysis, chronic phase patients treated with imatinib 400 mg/day who had BCR-ABL1 transcripts of 9.84 percent or less at 3 months had better overall, progression-free, and event-free survival at 8 years than did those with values of 9.84 percent or greater.⁵⁰³ Also, there is evidence that early molecular response to initial therapy with dasatinib or nilotinib in patients with CML is a predictor of overall response. In one trial with dasatinib, patients with BCR-ABL1 transcripts of 10 percent or less at 3 months had significantly better 5-year progression-free (92 vs. 67 percent) and 4-year overall survival than did those with greater than 10 percent transcripts (95 vs. 83 percent).⁵⁰⁴ In another study with nilotinib, patients with BCR-ABL1 of 10 percent or less at 3 months also had improved 4-year progression-free survival compared to those with BCR-ABL1 greater than 10 percent at 3 months (95 vs. 85 percent).⁵⁰⁵ Progression was defined as transformation to accelerated or blast phase.

Rising BCR-ABL1 Transcript Levels Rising BCR-ABL1 transcripts can indicate a new mutation or cytogenetic relapse. A significant change has been defined as either a twofold increase, serially increasing levels, or a 1-log increase.^492,495 There are no current recommendations for therapy changes based on an increase in BCR-ABL1 transcripts. Serial increases or increases of 1-log or more should trigger ABL mutational analysis and frequent monitoring of BCR-ABL1 transcripts.

Suboptimal Therapeutic Responses The initial European Leukemia Network guidelines defined suboptimal response as no cytogenetic response at 3 months, less than a partial cytogenetic response (PCyR) at 6 months, a PCyR at 12 months, and less than a MMR at 18 months.^495,506 The significance of a suboptimal response is dependent on the cause; for example, this may be insignificant if the result of drug intolerance or noncompliance as opposed to drug resistance (see “Acquired Resistance” below). The significance also depends on time of suboptimal response with earlier time points indicating a worse prognosis. Currently, CCyR and PCyR at 3 months are considered optimal and suboptimal responses. Suboptimal responses are labeled as a “warning” response in the Network guidelines.⁴⁹⁵ These response levels may trigger ABL mutation analysis or closer monitoring.

Secondary Chromosomal Changes with Tyrosine Kinase Inhibitors

Because of its earlier development, most of these data are obtained with imatinib treatment. Clonal abnormalities in cells lacking a detectable Ph chromosome or BCR-ABL1 rearrangements have been detected in patients undergoing imatinib therapy who previously were treated with IFN-α.^507,508 These cytogenetic changes were noted in seven patients at a median of 13 months of imatinib therapy, and trisomy 8 was the most frequent abnormality. All of these patients had MCyRs to imatinib.⁵⁰⁷ The presence of additional chromosomal abnormalities is considered to be a feature of the accelerated phase of CML. In some patients, clonal evolution may be related to imatinib resistance.⁵⁰⁹ Clonal abnormalities may be present in up to 10 percent of patients taking imatinib.⁵¹⁰ Some of these cases may be associated with a MDS, especially in those patients with previous exposure to cytarabine and idarubicin. The antiproliferative effect of imatinib allows restoration of polyclonal hematopoiesis in CCyR, which could permit the manifestation of a Ph chromosome–negative disorder.⁵¹¹ Some investigators have found that, with the possible exception of +8, +Ph, and i(17), additional chromosomal abnormalities at diagnosis are not associated with an inferior outcome to imatinib therapy.^512,513 In contrast, another group found that development of trisomy 8 in patients taking imatinib, while associated with pancytopenia, did not result in signs of disease progression. In a series of 34 CML patients who developed Ph chromosome–negative clones while taking imatinib, the most common abnormalities were trisomy 8 and monosomy 7. In 11 of these patients, no archival evidence of these clones was present before imatinib therapy was initiated, and none of the patients developed myelodysplasia.⁵¹⁴ In patients treated at diagnosis with imatinib, 9 percent developed chromosomal abnormalities in Ph chromosome-negative metaphases. These appeared at a median of 18 months, and the most common abnormalities were −Y and +8. Most were temporary and disappeared within 5 months. Only one patient with −7 progressed to acute myelogenous leukemia (AML).⁵¹⁵ Cytogenetic clonal evolution may not be an important impediment to achieving a MCyR or CCyR with imatinib, but it is an independent poor prognostic factor for survival of patients in chronic and accelerated phases of CML.⁵¹⁶ Imatinib therapy may overcome the poor prognostic significance of derivative chromosome 9 in CML.⁵¹⁷

Adherence to Tyrosine Kinase Inhibitor Therapy

Noncompliance with therapy is associated with poorer outcomes. In one trial, patients with a suboptimal response had higher nonadherence (23 percent) than did those with optimal responses (7 percent).⁵¹⁸ In another study, adherence was the only independent predictor for achieving a complete molecular response (CMR) on imatinib. Patients with an adherence rate of 85 percent or less had a greater chance of losing their CCyR at 2 years (27 percent) than did those with better adherence (1.5 percent). They, also, had a lower chance of remaining on imatinib.⁵¹⁹ Adherence has also been correlated with level of molecular response. In patients using a TKI for about 5 years, median adherence was 98 percent (range: 24 to 100 percent). If adherence was greater than 90 percent, there was a higher probability for a 3-log reduction in BCR/ABL1 transcripts and a CMR. If adherence was less than 80 percent, no MMRs occurred.⁵²⁰ The poor adherence to second-generation TKIs has not been studied for a sufficient duration to determine its impact. Management of side effects is of importance in maintaining a high rate of adherence.

Development of Tyrosine Kinase Inhibitor Resistance

The development of resistance to imatinib is not surprising.^521,522 Its specificity and “snug fit” into the ABL1-kinase pocket provide the ideal circumstance for resistance.⁵²³ Some cases demonstrate primary resistance to imatinib, and gene profiling has demonstrated differential expression of about 46 genes in responders compared to nonresponders.⁵²⁴ Even in patients with CCyR, malignant progenitors at the LTC-IC stage persist. Chronic phase CML stem cells are resistant to imatinib and are genetically unstable.⁵²⁵ These cells have a high level of BCR-ABL1 transcription, and they are thought to express transporter proteins that result in abnormal imatinib flux.⁵²⁶ Mathematical models suggest that imatinib rapidly eliminates leukemic progenitors, but does not deplete CML stem cells. Such models predict the probability of developing resistant mutations and can estimate the time that resistance will emerge.⁵²⁷ Several potential mechanisms of resistance include BCR-ABL1 amplification in the presence of imatinib, P-glycoprotein–mediated drug efflux, altered drug metabolism, acquisition of BCR-ABL1–independent signaling characteristics, and point mutations in the ABL1 kinase domain that decrease imatinib binding. Each of these mechanisms of resistance may have clinical relevance.

Primary Resistance Primary resistance to imatinib is defined as lack of CHR at 6 months or failure to achieve any level of cytogenetic response at 6 months, a MCyR at 12 months, or a CCyR at 18 months. This may occur in 15 to 25 percent of patients. Primary resistance may often be the result of inadequate plasma concentration because of binding of the drug to proteins, such as albumin or α₁-acid glycoprotein. In an analysis of the IRIS study, plasma levels of imatinib following the first month of treatment proved to be a significant predictor for clinical response. Plasma levels are not available for clinical use, however, so these have minimal influence on treatment decisions when responses are not as expected. Only one gene, prostaglandin-endoperoxide synthase 1/cyclooxygenase1 (PTGS1/COX1) was found to differentiate primary imatinib resistance. Eleven genes were associated with secondary resistance after imatinib therapy in those without an ABL1 kinase domain mutation.⁵²⁸ Expression of the OCT-1, which mediates drug influx, is thought to be important for imatinib but not dasatinib effectiveness.^529,530 Many CML patients who have a suboptimal response to imatinib have low OCT-1 activity, but this can be overcome with higher doses of imatinib or use of dasatinib, which uptake is not dependent on OCT-1 expression.⁵³⁰ OCT-1 expression is associated with MMR at 12 and 24 months, and it is a predictor of the long-term risk of resistance and of transformation in patients treated with imatinib.⁵³¹ CML CD34+ cells overexpress the drug transporter ABCG2, and imatinib, dasatinib, and nilotinib are substrates for ABCB1 and ABCG2. Overexpression of MDR1 has been associated with decreased intracellular concentration of imatinib.⁵³²

Acquired Resistance Acquired resistance is that which occurs after exposure to TKIs or other treatments. Amplified gene expression and increased BCR-ABL1 protein expression are often reported in resistant patients. Duplication of the Ph chromosome and isodicentric chromosomes are a possible mechanism of resistance to imatinib.^533,534

Mutations in the ABL1 kinase domain are a frequent mechanism of resistance. Kinase domain mutations were the only independent predictor for the loss of CCyR and progression when compared to those without a mutation.⁵³⁵ Mutations in the ABL1 kinase domain may predate imatinib treatment,⁵³⁶ and several BCR-ABL1 kinase domain mutants associated with imatinib resistance remain sensitive to the drug, suggesting a need for characterization before a resistant phenotype can be attributed to the given mutation.⁵³⁷ The mutant clone does not always have a proliferative advantage.⁵³⁸ Some of these mutations may lie outside the kinase domain, and more than 40 such mutations have been described. Screening early phase CML patients for mutations before the start of imatinib therapy is not cost-effective because of their low incidence, but in patients with evidence of an increase in CML cells while on imatinib, mutation searches are indicated.⁵³⁹ BCR-ABL1 kinase domain point mutations are rare in those who have had good cytogenetic responses to imatinib, and when detected in that setting, their presence does not always predict relapse.⁵⁴⁰ Mutations in the ABL1 portion of the BCR-ABL1 oncogene are present in approximately 40 percent of patients who do not achieve a CHR or CCyR to imatinib. ABL1 mutations were found in those patients with both primary and acquired resistance. Amino acid substitutions in seven residues accounted for 85 percent of all mutations associated with resistance.⁵⁴¹ The mutations most associated with resistance are Thr315ILe, Gly250Glu, Glu255Lys, and Thr253His substitutions. Few of the described mutations directly affect imatinib binding.⁵⁴² Mutations in the ABL1–ATP phosphate-binding loop (P-loop) are most closely associated with a poor prognosis,⁵⁴³ and these P-loop mutations predict for disease progression. Overall survival is worse for P-loop and for T315I mutations, but not significantly different when other mutations are present.⁵⁴⁴

Ultra-deep sequencing approaches have shown that routine Sanger sequencing underestimates BCR-ABL1 mutation in 55 percent of samples where the missed mutations had low abundance.⁵⁴⁵ Mass spectrometry can detect a 0.05 to 0.5 percent level of mutations, as well.⁵⁴⁶ For many mutations, the concentration that inhibits 50 percent (IC₅₀) of various TKIs and response have not been documented.⁵⁴⁷

Second-generation BCR-ABL1 inhibitors (see “Dasatinib and Nilotinib” below) are able to overcome imatinib-resistant mutants, with the exception of the T315I mutations (Table 89–6). Mutations F317L and V299L are resistant to dasatinib and mutations Y253H, E255K, and F359I are resistant to nilotinib. Ponatinib was active against T315I and against other BCR-ABL1 mutations resistant to dasatinib or nilotinib.⁵⁴⁸ Ponatinib may be effective against individual ABL1 point mutations but may not overcome some compound mutations, which are two or more mutations in the same BCR/ABL1 molecule. Some mutations may be polyclonal as well.⁵⁴⁹ The T315I mutation results in steric hindrance, which precludes access of some TKIs to the ATP-binding pocket of the ABL1 kinase domain.⁵⁵⁰ In a series of 27 patients with T315I mutation, survival was dependent on stage of disease, with many of the chronic phase patients described as having an indolent course.⁵⁵¹ In addition to ponatinib, agents such as IFN-α and homoharringtonine have also been proposed as therapy for those with the T315I mutation.⁵⁵²

In some cases of resistance associated with imatinib, other signal pathways independent of BCR-ABL1 may become important in cell proliferation.⁵⁵³ These include heat shock protein (hsp) 70,⁵⁵⁴ survivin,⁵⁵⁵ LYN kinase,⁵⁵⁶ SRC,⁵⁵⁷ and GRB2, among others.⁵⁵⁸

Dose escalation, combination therapy, and treatment interruption have been proposed as means to overcome drug resistance.⁵⁵⁹ Combination therapy from the outset⁵⁶⁰ also has been proposed to prevent development of resistance. Treatment interruption to stop clonal selection of resistant cells has been proposed.⁵⁵⁷ Gene-expression profiles may be useful to predict the clinical effectiveness of imatinib for CML treatment, thereby allowing individualized therapy from the outset.⁵⁶⁰ In patients with relapse or resistance, alternative approaches include increasing the dose of imatinib or switching to dasatinib or nilotinib.⁵⁶¹ Allogeneic hematopoietic stem cell transplantation is not recommended unless patients have inadequate response or intolerance to multiple TKIs or have the T315I mutation. Mutational analysis is not recommended at diagnosis but is of help in selecting TKI therapy for patients with an inadequate initial response or loss of response to TKI therapy. Mutation type may dictate choice of the next TKI.⁵⁶²

Management of Resistance

Dose Escalation If the patient is on imatinib, 400 mg/day, dose escalation to 800 mg/day can be tried. This may be most efficacious in patients with cytogenetic relapse who had achieved a cytogenetic response with the initial dose of imatinib, but is not likely to benefit those who have not had a cytogenetic response with imatinib at 400 mg/day.

Second-Generation Tyrosine Kinase Inhibitor Therapy Several TKIs are approved for use in the case of imatinib resistance or intolerance. In general, outcome with each is comparable, so the choice of agent often depends on its side-effect profile or in some cases on mutation type (see Tables 89–2 and 89–6).⁵⁶²

The 3-month molecular response after initiation of a second TKI is also a predictor of overall and event-free survival for those patients in chronic phase when switched from imatinib to another TKI.⁵⁰⁵ A BCR-ABL1 level of 10 percent or less at 3 months is desirable. In patients treated with nilotinib after imatinib resistance or intolerance, the 4-year progression-free and overall survival was 85 and 95 percent, respectively, if BCR-ABL1 transcripts were 10 percent or less as compared to 42 and 71 percent for those with BCR-ABL1 transcripts greater than 10 percent at 3 months.⁵⁶³

Either dasatinib or nilotinib can be used in cases of imatinib resistance or intolerance. Dasatinib is 325-fold more potent than imatinib; and, as a dual inhibitor of SRC and ABL1 kinases, dasatinib is able to bind to BCR-ABL1 with less-stringent conformational requirements.⁵⁶⁴ In patients resistant to imatinib, dasatinib, 140 mg/day (70 mg q12h), resulted in a higher proportion of MCyR, CCyR, and MMR than did 800 mg/day (400 mg q12h) of imatinib. Treatment failure was decreased and progression-free survival was improved with dasatinib.⁵⁶⁵ Unlike imatinib, dasatinib penetrates the blood–brain barrier.⁵⁶⁶ In long-term followup of 670 patients with imatinib-resistant or intolerant CML, treated with various doses of dasatinib, 28 percent of patients remained on study treatment at 6 years. Survival was in the range of 76 to 83 percent and a MMR was achieved in 45 percent on a dose of 100 mg daily. Molecular and cytogenetic responses at 3 and 6 months were predictive of survival. For those with BCR-ABL1 transcripts less than 10 percent at 3 months, progression-free survival was 68 percent, whereas it was only 26 percent for those with BCR-ABL1 transcripts greater than 10 percent.⁵⁶⁷ Nilotinib in a dose of 400 mg every 12 hours induced approximately 40 percent of patients who were resistant to or intolerant of imatinib into a MCyR, and approximately 30 percent into a CCyR. Nilotinib, 400 mg BID, is the recommended dose for those intolerant or resistant to imatinib.

Bosutinib and Ponatinib In addition to dasatinib and nilotinib, bosutinib and ponatinib, third-generation TKIs, are active against many of the imatinib-resistant BCR-ABL1 kinase domain mutations and are effective treatment options for CML resistant to standard-dose imatinib. Both were FDA approved for this indication in 2012. Bosutinib is active with F317L, Y253H, and F359C/I/V mutations. Ponatinib is also effective against many mutations resistant to nilotinib or dasatinib and has activity in cases with a T315I mutation (see Table 89–6).

Bosutinib is a dual SRC-ABL1 TKI that resulted in a CCyR in 24 percent and CHR in 73 percent of patients who had used two prior TKIs.⁵⁶⁸ All mutations except T315I were responsive. It is approved for chronic phase, accelerated phase, or blast phase in those resistant or intolerant to prior TKI therapy. The dose is 500 mg/day, orally, with food. Diarrhea, nausea, decreased platelet count, other gastrointestinal complaints, rash, and anemia were the most common adverse events, and most of these were of low-grade. Bosutinib is not yet approved for use as an initial agent in CML, as it did not demonstrate significant improvement in CCyR rates at 1 year compared to imatinib. It was, however, associated with higher 1-year MMR rates, faster time to response, and less disease progression.⁴⁹¹

Ponatinib blocks native and mutated BCR-ABL1 including the gatekeeper mutation T315I. In a phase I dose escalation study, pancreatitis, rash, and myelosuppression were major toxicities. In 12 patients with T315I mutations, 92 percent had a MCyR. In those with accelerated or blast phase, or Ph chromosome–positive ALL, 36 percent had a major hematologic response and 32 percent had a MCyRs. Arterial thrombotic events occurred, however.⁵⁴⁸ Retrospective analyses of these incidents led to temporary withdrawal of this medication, which is now used primarily in cases of T315I mutation or failure of several other TKIs. If vascular occlusion occurs, therapy should be halted immediately.⁵⁶⁹ Ponatinib has a black box warning for vascular occlusion, heart failure, and hepatotoxicity.

A third-line TKI should be considered in patients who have failed two prior generations of TKI therapy, but responses tend to be infrequent and are usually not durable, so clinical trials, other agents, or allogeneic hematopoietic stem cell transplantation should be considered. A prior CCyR on either imatinib or subsequent nilotinib or dasatinib therapy was the only predictor of a cytogenetic remission on a third-line therapy.⁵⁷⁰

Non–Tyrosine Kinase Inhibitor Therapies

Omacetaxine Omacetaxine (homoharringtonine) is a Cephalotaxus alkaloid that has activity against the T315I mutation. Omacetaxine was approved on the basis of a study that recruited patients who had already been on two or more TKIs. In those enrolled in chronic phase, 67 percent had a CHR; a MCyR or a CCyR was achieved in 22 and 4 percent, respectively. Median overall survival was 30 months.⁵⁷⁰ In those with T315I mutations enrolled on a separate study, MMR was achieved in 17 percent of patients and the T315I clone was reduced in 61 percent of patients.⁵⁷¹ The most common side effects with this medication are cytopenias. This agent also has activity in accelerated phase CML, but little in blast phase. It was approved by the FDA in October 2012 for chronic and accelerated phase patients intolerant of other therapy or in cases not responding to two or more TKIs.

Several inhibitors of signal transduction mediators involved in the downstream effects of BCR-ABL have been proposed for use in imatinib-resistant CML. These inhibitors include the JAK2 inhibitor AG490,⁵⁷² SRC kinase inhibitors, mTOR (mammalian target of rapamycin) inhibitors, such as rapamycin,⁵⁷³ the proteasome inhibitor bortezomib,^574,575 histone deacetylators,^576,577 PI3K or MEK (mitogen-activated kinase) inhibitors and inhibitors of the WNT/β-catenin pathway thought to be active in CML stem cells. Imatinib resistance often is associated with restored activation of the BCR-ABL1 signal transduction pathway, suggesting that BCR-ABL1 remains a valid target to overcome resistance in these cases.⁵⁷⁸ BCR-ABL1 point mutations isolated from patients with imatinib-resistant CML are sensitive to inhibitors of the BCR-ABL1 chaperone hsp90, such as geldanamycin.⁵⁷⁹ Many of these agents have not yet entered clinical trials, but some are in early phase trials, such as inhibitors of the hedgehog pathway, which is activated in CML but not normal hematopoietic stem cells.⁵⁸⁰ Some are being used in conjunction with imatinib in resistant cases.

Combined Therapy Agents that have been proposed for use in combination to improve response rates or to overcome resistance to imatinib have included IFN-α, cytarabine, omacetaxine multiagent chemotherapy, arsenic trioxide, and decitabine, with some supporting in vitro data.^{581,582,583,584,585,586} Combining imatinib with chemotherapeutic agents is more myelosuppressive, and final effects on response rates and survival have yet to be determined. This approach is rarely used in chronic phase CML, but is used in accelerated or blast phase or in Ph chromosome–positive acute lymphoblastic leukemia.⁵⁸⁷

Disease Prognosis and Monitoring During Tyrosine Kinase Inhibitor Therapy

Treatment failure should lead to alterations in therapeutic strategy.⁵⁸⁸ For patients treated initially with imatinib, BCR-ABL1 expression in cytogenetic responders and nonresponders was similar. BCR-ABL1 expression became significantly different 3 months after treatment and became increasingly different between responders and nonresponders with continued therapy at 6, 9, and 12 months.⁵⁸⁹

One mode of monitoring patients undergoing TKI therapy is to measure blood counts at least once per month and to obtain marrow samples every 6 months until a complete cytogenetic remission is obtained.⁵⁹⁰ Thereafter, marrow samples are obtained no more often than yearly to monitor for other clonal abnormalities. Quantitative RT-PCR is performed every 3 months on blood or marrow. A 1-log increase in the level of BCR-ABL1 reactivity, confirmed on a repeat sample at least 1 month later, suggests a loss of response to treatment. In patients who do not have a CHR at 3 months, or a MCyR after 6 to 12 months, other therapeutic options are considered.⁵⁹¹ The molecular response after 2 to 3 months of therapy is a strong predictor of clinical and cytogenetic response.⁵⁹² Sequencing the BCR-ABL1 kinase domain can reveal emergence of resistant clones and is useful if there is an insufficient initial response to imatinib or a second-generation TKI (see Table 89–5) or any sign of loss of response, such as relapse to Ph chromosome–positive status, a 1-log increase in BCR/ABL1 transcript ratio, or loss of a MMR.⁴¹⁷

In patients receiving second-generation TKIs as second-line therapy, those who have no cytogenetic response at 3 to 6 months should be considered for allogeneic transplantation or switched to an alternative therapy in a clinical trial. After CCyR is attained, molecular monitoring is recommended every 3 months for 3 years and every 4 to 6 months thereafter.⁴⁹² After 12 months, those with a MCyR had a significant survival advantage over those with lesser responses.⁵⁹³ Those with BCR-ABL1 transcripts greater than 10 percent following initial imatinib treatment should be switched to dasatinib, nilotinib, or bosutinib. If this is not an option because of cost or availability, high-dose imatinib can be considered, but it also unlikely to have benefit for more than a few months. For those already on a second-generation TKI, a clinical trial or alternative TKI could be tried or patients may continue on the same TKI with very careful followup, as the impact of this benchmark on overall survival is not yet known.⁴⁹² The 6-month evaluation should identify patients with a poor outcome. For those who have MCyR or BCR-ABL1 of less than 10 percent at 6 months, overall survival was 100 percent as compared to 79 percent for those with no response.⁵⁹⁴ At 12 and 18 months, CCyR is the optimal response.⁴¹⁷ In those who have achieved CCyR, MMR may not be of prognostic significance.⁵⁹⁵

Rising BCR-ABL1 levels may be associated with an ABL1 mutation or relapse of disease. Those with more than a twofold rise in BCR-ABL1 are more likely to have a mutation.⁵⁹⁶ A serial rise in the BCR-ABL1 level may also indicate loss of response to therapy.⁴¹⁵

OTHER AGENTS USED IN TREATMENT OF CHRONIC PHASE

Interferon-α

IFN-α, formerly the most effective agent, is rarely used in the treatment of CML. A CCyR with IFN-α was uncommon (13 percent), but 10-year survival rates in responders were approximately 70 percent.⁵⁹⁷ CCyRs to IFN-α were stable and durable.⁵⁹⁸ Approximately 50 percent of complete responders become long-term survivors. Common toxicities of IFN-α use include fatigue, low-grade fever, weight loss, liver function test abnormalities, hematologic changes, and neuropsychiatric symptoms. Overall survival is improved in imatinib-treated patients compared with patients treated with IFN-α or IFN-α plus cytarabine.⁵⁹⁹ Nevertheless, among all patients who attained a major or CCyR at 12 months, the survival rate was comparable in either case. IFN-α has also been proposed as an immune stimulant to consolidate imatinib remissions because additive effects have been noted.^600,601 Conversely, those treated initially with INF-α who achieve a CCyR have an improved molecular response with imatinib.^602,603 Some patients intolerant to a TKI may be treated successfully with INF-α.

Use of Other Chemotherapeutic Agents in Chronic Phase

Hydroxyurea The major side effect of hydroxyurea is an extension of its pharmacologic effect, that is, reversible suppression of hematopoiesis, often with megaloblastic erythropoiesis. The median survival of patients with CML treated with hydroxyurea alone is approximately 5 years. Studies with high-dose hydroxyurea indicate that marrow metaphase cells in some patients lose the Ph chromosome either partially or completely after such therapy.⁶⁰⁴ Hydroxyurea often is used for initial cytoreduction, but it has few other indications in the TKI era of CML therapeutics. Chronic use of hydroxyurea is associated with leg ulcers.⁶⁰⁵

Cytarabine IFN-α_2b combined with cytarabine (20 mg/m² per day for 10 days per month) in the chronic phase was associated with a greater proportion of MCyRs at 12 months and with greater survival prolongation than was IFN alone.⁶⁰⁶ Toxicities with these drug combinations were greater, and this combination has been replaced by TKI therapy and is rarely used.

Busulfan Once the mainstay of treatment for the chronic phase, busulfan usage now is rare.⁶⁰⁷ It is used primarily as part of the preparative regimen for allografting or autografting. It may be used occasionally in older patients who do not tolerate TKIs.

Other Cytotoxic Agents Intensive multidrug regimens have been used in an attempt to eradicate the Ph chromosome–positive clone and, occasionally, have led to prolongation of remission or cure of the disease. This approach did not significantly increase population survival.⁶⁰⁸

Other Potential Therapeutic Agents in Chronic Phase Chronic Myelogenous Leukemia The farnesyltransferase inhibitors lonafarnib and tipifarnib have been combined with imatinib and have activity after imatinib failure.^609,610 The hypomethylation agent decitabine has activity in imatinib refractory CML. INNO-406, a dual BCR-ABL/LYN inhibitor, suppresses the growth of CML cells in the central nervous system.⁶¹¹ MicroRNA approaches may eventually play a role in CML treatment,⁶¹² and synthetic BCR-ABL1 small interfering ribonucleic acid (siRNA) has been used in a patient with resistant CML, postallografting with inhibition of BCR-ABL1 noted.⁶¹³ Ribozymes targeting BCR-ABL1 mRNA have been used as CML treatment,^614,615 and these approaches probably will have the most utility for in vitro purging of CML marrow cells before autotransplantation.^616,617

RADIOTHERAPY

Palliative splenic irradiation may be useful occasionally in subjects who have entered the accelerated or advanced chronic phase and are troubled with extreme splenomegaly with splenic pain, perisplenitis, and encroachment of the spleen on the gastrointestinal tract.⁶¹⁸ Splenic irradiation may palliate symptoms for a short time.⁶¹⁹ Spleen size associated with chronic phase disease usually is decreased with TKI therapy.

Palliative radiotherapy may be useful for extramedullary myeloid sarcomas, which may occur occasionally in bone or soft tissue during the late chronic or accelerated phase.

SPLENECTOMY

Splenectomy does not prolong the chronic phase of CML, delay the onset of the accelerated phase, enhance sensitivity to TKIs or chemotherapy, or prolong survival of patients.⁶²⁰ In carefully selected patients with symptomatic thrombocytopenia unresponsive to therapy, mechanical discomfort, hypercatabolic symptoms, and portal hypertension, splenectomy may be useful. Postoperative morbidity from infection, thrombosis, or hemorrhage has been high, with mortality rates up to 10 percent reported.⁶²¹ Splenectomy performed before allografting has not been found to influence the severity of graft-versus-host disease (GVHD) or survival after allogeneic hematopoietic stem cell transplantation.⁶²²

TREATMENT OF CHRONIC PHASE CHRONIC MYELOGENOUS LEUKEMIA DURING PREGNANCY

Treatment of chronic phase CML during pregnancy is sometimes needed to prevent placental insufficiency from hyperleukocytosis. Imatinib may be teratogenic. Normal newborns have been delivered by patients who conceived and ingested imatinib during early pregnancy.^{623,624,625,626} In 125 women exposed to imatinib during pregnancy, 50 percent delivered normal infants, and 25 percent underwent elective terminations, three of the latter following the identification of fetal abnormalities. Twelve other infants had abnormalities.⁶²⁷ The majority of patients who discontinue imatinib during pregnancy lose their complete hematologic remission and their cytogenetic responses.⁶²⁸ One fetal fatality during pregnancy as a result of a meningocele has been reported. Males treated with imatinib have fathered healthy infants.⁶²⁹ Current recommendations are to practice contraception during treatment with any TKI, or, if pregnant, at the onset of the disease, to consider IFN treatment until delivery.^626,630 Imatinib does appear in breast milk.⁶³¹

IFN can be used during pregnancy with minimum risk of teratogenicity. Eight patients treated with IFN from the first trimester have been described, and each of these pregnancies resulted in normal infants, except for one with mild neonatal thrombocytopenia. All infants had normal growth patterns.⁶³² Hydroxyurea may be useful during the second and third trimester but should be avoided in the first trimester.^626,633 Leukapheresis in the first trimester (or longer) also can be used to avoid fetal drug exposure early in pregnancy (see “Leukapheresis” above). It is important to use a TKI after delivery to achieve the best outcome. Further observation may show it to be safe later in pregnancy. Although controversial, stopping and later restarting TKI therapy may not result in as favorable an outcome of therapy as use of IFN or other approaches, initially.⁶²⁶ If conception is desired, attaining a MMR before the TKI therapy is discontinued and a 3-month washout period are recommended.⁴¹⁵ The patient should be made aware of the risk for CML progression during the pregnancy.

DISCONTINUATION OF TYROSINE KINASE INHIBITOR THERAPY

Patients responding to TKI therapy are likely to maintain their response. TKIs are associated with a significant symptom burden, however,⁶³⁴ and one-third have persistent moderate to severe symptoms. Persistent CMR is seen in only a minority of patients, and the vast majority of patients on TKIs have minimal residual disease even if their BCR-ABL1 transcripts are undetectable, and this may lead to relapse if the drug is stopped. Thus, in the absence of a clinical trial, life-long therapy with a TKI is recommended.

Some patients in a CMR for 2 years or more have been able to discontinue imatinib without relapse.^635,636 Discontinuation of imatinib in 12 patients who had undetectable disease for at least 2 years resulted in six patients having a molecular relapse within 1 to 5 months (imatinib was reintroduced with a response) and six others remaining in CMR for a median of 18 months.⁶³⁷ There are numerous anecdotes of patients relapsing when imatinib was stopped. In patients with intolerable side effects on imatinib, the dose may be reduced in some cases without the loss of a CMR.⁶³⁸ In occasional patients in whom imatinib is stopped, a cytogenetic response of up to 15 months has persisted.^{639,640,641,642} In a study of 100 patients with a CMR (>5-log reduction in BCR/ABL1 transcripts) for at least 2 years who stopped imatinib, 69 patients had at least 12 months of followup. Of these, 39 percent were stable and 61 percent relapsed, most within the first 6 months. The outcome of stopping was better for those with a lower Sokal risk score at diagnosis.⁶³⁶ In another study, 40 patients were evaluated, and treatment-free remission at 24 months was 47.1 percent for all patients and was higher if patients had prior IFN treatment.⁶³⁵ Female sex and early molecular response predict stable undetectable BCR-ABL1 transcripts in chronic phase CML, the criteria for early stopping.⁶⁴³ In a series of 423 CML patients treated with imatinib, the rate of undetectable BCR/ABL1 transcripts and stable MMR (4.5-log decrease) was 36.5 percent.⁶⁴³ A model to analyze the risk of molecular relapse after cessation of TKIs has been developed.⁶⁴⁴ Reports of discontinuations, reappearance of the clone, treatment with a second prolonged course of imatinib, discontinuation with retention of MMR at 32 months followup, and retention of CMR in 12.5 percent have been described.⁶⁴⁵

Discontinuation of nilotinib or dasatinib has not been reported in a significant series, and larger prospective studies will be needed to determine in which patients stopping treatment can be safely attempted. Because early, quiescent Ph chromosome–positive cells (CD34+Lin−) are insensitive to imatinib in vitro,⁵⁴⁶ at present it is advisable to maintain treatment indefinitely until the criteria for cessation, if any, can be established in clinical trials. Intermittent imatinib dosing has been explored in selected elderly patients with CML without adverse impact on overall and progression-free survival.⁶⁴⁷

HIGH-DOSE CHEMOTHERAPY WITH AUTOLOGOUS STEM CELL INFUSION

Since the availability of imatinib, autografting in CML is rarely used.⁶⁴⁸ Ph chromosome–negative stem cells are present in most patients with CML at the time of diagnosis. Techniques that use these cells to reconstitute hematopoiesis after high-dose therapy have been developed.⁶⁴⁹ Ph chromosome–negative progenitors can be mobilized with G-CSF and collected from the blood of patients who have responded to prior treatment with a TKI.⁶⁵⁰ Such cells also can be collected after recovery from chemotherapy regimens, such as after idarubicin and cytarabine, followed by G-CSF stimulation.⁶⁴⁹ G-CSF was used for at least 4 days while imatinib treatment was continued for stem cell mobilization in 58 patients with a CCyR. The cells were collected in two cytapheresis procedures in 74 percent of patients, and the cells of 84 percent of those cytapheresis products were negative for the Ph chromosome.⁶⁵¹

In another series, stem cells were mobilized in 32 patients in complete cytogenetic remission after imatinib, with uninterrupted imatinib therapy in 50 percent of patients and with imatinib temporarily withheld in approximately 50 percent. Blood levels of BCR-ABL1 transcripts were not changed by the use of G-CSF.⁶⁵² In yet another series, 13 of 15 patients were successfully mobilized with G-CSF while receiving imatinib, and 28 percent of stem cell harvests were negative for BCR-ABL mRNA. No change in blood BCR-ABL1 transcript level was noted after stem cell mobilization as assessed by RT-PCR.⁶⁵³ No series of patients autografted with cells mobilized while they were receiving imatinib have been reported.⁶⁵⁴ Autografting might find a role in cases of imatinib resistance,⁶⁵⁵ or to reduce the level of residual disease in cases without a molecular response.⁶⁵⁶ Imatinib can be effective and safe in chronic phase CML patients who have previously undergone autografting,⁶⁵⁷ although there is an increased frequency of hematologic toxicity.

ALLOGENEIC HEMATOPOIETIC STEM CELL TRANSPLANTATION

Until imatinib became available in 2001, allogeneic transplantation was used in most new patients with CML who were younger than 65 years of age and who had a suitable donor. The advent of imatinib treatment and the projected survival of patients with a complete cytogenetic remission has changed the indications for transplantation in CML.^658,659 There has been a marked reduction in number of transplants performed for CML worldwide and a decline in the proportion performed in first chronic phase.^660,661 Although no randomized trials of imatinib versus transplantation have been or are likely to be conducted, there is circumstantial evidence that survival is superior for populations of patients treated with drugs who have a CCyR as compared to transplantation.⁶⁶² Allografting continues to play a role in the treatment of patients, who are refractory or intolerant to serial TKIs, and remains the optimal therapy in those who progress to accelerated or blast crisis in the face of treatment with a series of TKIs.

Patients in the chronic phase of CML who are younger than approximately 70 years and who have an identical twin,⁶⁶³ or a histocompatible sibling,^664,665 or access to a histocompatible unrelated donor,⁶⁶⁶ can be transplanted after intensive therapy, usually with cyclophosphamide and fractionated total-body irradiation (TBI) or a combination of busulfan and cyclophosphamide. Busulfan can be administered as an intravenous preparation and as a single daily dose.⁶⁶⁷ When targeted steady-state busulfan levels are used, a 3-year survival rate of 86 percent and a disease-free survival rate of 78 percent with no age effect is achieved.⁶⁶⁸ With nonmyeloablative or “reduced-intensity” conditioning regimens, older patients and those with comorbidities can undergo successful allografting.⁶⁶⁹ With the success of TKI therapy, allogeneic stem cell transplantation is no longer recommended as a treatment option in chronic phase CML responding to any sequence of TKIs.⁶⁷⁰ Allogeneic transplant should be considered for patients with disease progression to accelerated or blast phase on TKI therapy. In these cases, TKIs to which the patient has not been previously exposed and for which they do not possess a resistant mutation can be used to bridge or prepare for the transplant.⁶⁷¹ Allogeneic transplant can be used in those rare patients who present with blast crisis and in those with T315I mutations who do not respond to ponatinib.^672,673 In those who do not respond to their first TKI exposure and are switched to a second TKI, transplantation in chronic phase would be indicated for those patients with BCR-ABL1 transcripts greater than 10 percent or less than a PCyR at 3 and 6 months, minor or no cytogenetic response at 12 months, and only a PCyR at 18 months or cytogenetic relapse at 12 or 18 months.⁴¹⁷

Myeloablative Allogeneic Transplants

Stem cell transplantation from HLA-compatible siblings results in engraftment and an actual or projected long-term survival in 45 to 85 percent of recipients.^674,675,676 In patients older than age 50 years, survival rates are slightly less at 5 years. The risk of CML relapse is approximately 20 percent, with a plateau of relapse at 5 to 7 years. Transplanted T lymphocytes, especially if activated by a (mild) GVHD, may be an important factor in preventing leukemic relapse. This phenomenon, referred to as graft-versus-leukemia reaction, is thought to suppress the leukemic process through T-cell–mediated cytotoxicity.⁶³⁴ The relative benefit of marrow compared to mobilized blood stem cells as the source of the allograft has not been established.^677,678 Mobilized blood stem cells engraft more rapidly but may be associated with more chronic GVHD. The majority of survivors have no evidence of residual leukemia.⁶⁷⁹

For younger patients who do not have a histocompatible sibling, an unrelated donor or a mismatched family member as a source of stem cells is feasible. When class I HLA genes are typed with molecular methods, an improvement in matching and better outcomes using unrelated donors have been demonstrated and are comparable to those transplanted with a matched-sibling donor. When matched-unrelated donor and sibling donor transplants were compared, unrelated donor transplants had increased risk of graft failure and acute GVHD, but only a slightly poorer survival and disease-free survival. For patients who survived to 1 year, only a slightly inferior disease-free survival was observed.⁶⁸⁰ The rate of extensive chronic GVHD is up to 60 percent with unrelated donor transplants, but 63 percent disease-free survival in younger CML chronic phase patients has been reported. Cord blood stem cell transplantation from an unrelated donor has also been used in adults with CML.⁶⁸¹

Pretransplantation imatinib is not associated with increased transplant-related morbidity or decreased survival, but those who are transplanted with suboptimal response to imatinib or loss of response to imatinib fare worse, probably related to a higher disease burden at the time of transplantation and more aggressive disease.^682,683 Second-generation TKIs do not increase transplant-related toxicity.⁶⁸⁴ Disease status after allografting can be monitored with cytogenetic studies, PCR, or FISH analysis. A positive PCR assay 3 months after allogeneic transplantation has not been found to correlate with an increased risk of relapse compared with PCR-negative patients. A positive assay at 6 months and beyond is associated with subsequent relapse. In one series, 42 percent of patients with a positive PCR assay at 6 to 12 months relapsed versus 3 percent with a negative assay.⁶⁸⁵ Paradoxically, patients who remain BCR-ABL1 positive more than 36 months after transplantation have little propensity for relapse.⁶⁸⁵ Serial quantitative RT-PCR analysis of blood specimens has been proposed to distinguish patients destined to relapse.⁶⁸⁶ Patients who remain in remission have undetectable, low, or falling BCR-ABL1 levels on sequential analysis. After 6 to 9 months, these levels are undetectable in most cases. Recognition of relapse at the molecular level may allow for early therapeutic intervention.

Killer immunoglobulin-like receptors (KIRs) are expressed by NK cells and subpopulations of T cells. NK clones from a single individual can vary substantially in the type of KIR molecules they express. The ligands for several of the inhibitory KIR have been shown to be subsets of HLA class I molecules. Missing KIR ligands in recipients lead to less relapse and increased GVHD based on NK alloreactivity.⁶⁸⁷ KIR ligand mismatch has also been found to be an important prognostic factor in achieving molecular responses after transplantation for CML.⁶⁸⁸ Increased frequency of regulatory T cells characterized as CD4+, CD25-high are associated with higher rates of relapse after allografting in CML.⁶⁸⁹

Nonmyeloablative Allogeneic Transplants

Nonablative regimens have been developed in an attempt to expand the indication for allogeneic transplantation to older patients. These regimens rely on immunosuppressive therapy to allow engraftment of cells that potentially will generate a graft-versus-leukemia effect. These procedures in general are associated with acceptable degrees of engraftment, less mortality, similar rates of GVHD, and possible durable effects on persistent or recurrent disease.⁶⁹⁰ Approximately 60 percent of patients, mostly in initial chronic phase (median age: 50 years) and transplanted with reduced-intensity conditioning regimens, had a 3-year survival and about one-third had a 3-year progression-free survival.⁶⁹¹ Patients no longer in first chronic phase do not fare as well.⁷¹² Conditioning regimens include fludarabine and busulfan,^693,694 low-dose TBI and fludarabine, and low-dose TBI and cyclophosphamide, but no prospective randomized trials comparing regimens or comparing ablative and nonmyeloablative transplant approaches have been conducted.^693,695,696 In one case, imatinib given concurrently with nonmyeloablative stem cell transplantation did not compromise engraftment and resulted in a cytogenetic remission in a patient with CML in blast crisis.⁶⁹⁷ Whereas nonmyeloablative or reduced intensity regimens are still considered investigational, they can achieve molecular remissions.

USE OF TYROSINE KINASE INHIBITORS AFTER STEM CELL TRANSPLANT

In patients treated with stem cell transplantation before the wide availability of imatinib, complete cytogenetic remissions with imatinib treatment can occur if treated at relapse after allografting and after donor lymphocyte infusion (DLI) fails to give a response. Such remissions may include a molecular response.⁶⁹⁸ Complete responses after imatinib therapy have been noted in accelerated or blast phase CML persisting after stem cell transplantation.⁶⁹⁹ In another series, 45 percent of relapsed patients had a CCyR of up to 28 months and without significant GVHD.⁷⁰⁰ In a series of 28 adults with relapse after allogeneic stem cell transplantation who then received imatinib, the response rate was 74 percent, and the complete cytogenetic remission rate was 35 percent. Five patients had recurrence of GVHD, and 13 had received previous DLI infusions.⁶⁹⁸ In one series, imatinib was able to generate complete molecular remissions in 26 percent of chronic phase patients after allografting, with full donor chimerism usually observed.⁷⁰¹ One study found that more relapsed patients had a molecular remission with DLI at 5 years than with imatinib alone. Most of these patients had cytogenetic relapses only.⁷⁰² In a retrospective comparison of 37 patients with hematologic or molecular relapse of CML who received imatinib, DLI, or a combination of both in concurrent or sequential regimens, the overall survival was 100, 89, and 54 percent for both modalities, imatinib, alone, and DLI alone, respectively.⁷⁰³ There are few studies examining use of nilotinib or dasatinib for posttransplantation relapses.⁷⁰⁴

Prophylactic administration of imatinib after transplantation has been used for patients at high risk of relapse.⁷⁰⁵ Posttransplantation imatinib may also postpone the requirement for DLI with its attendant risks of GVHD and marrow aplasia.⁷⁰⁶ Some have found that DLI are superior to imatinib therapy in preventing relapse and increasing leukemia-free survival. Posttransplantation imatinib is usually well-tolerated; pancytopenia is the principal toxicity. The cytopenias resolve with doses adjustments or temporary drug discontinuation. Many questions remain regarding the use of posttransplantation TKIs. When should they be started? Which drug is superior? For how long should the drugs be used? Should they only be started in cases of molecular relapse? How can they be best combined with DLI in cases of relapse?

IMMUNOTHERAPY: ADOPTIVE CELL THERAPY FOR POSTTRANSPLANTATION RELAPSE

Substantial evidence indicates that the effectiveness of allografting in CML does not result solely from the eradication of the leukemic clone with high-dose chemoradiotherapy conditioning regimens, but also from adoptive immunotherapy provided by lymphocytes in the allograft, the graft-versus-leukemia effect (see “Myeloablative Allogeneic Transplants” above).⁷⁰⁷ This phenomenon has been recreated to produce a therapeutic response by infusing the lymphocytes from the stem cell donor after a relapse following allogeneic stem cell transplantation.^708,709 The overall response rate to DLI is approximately 75 percent. The response rate is higher when this approach is used early after detecting a relapse by PCR,⁷¹⁰ compared to use after a hematologic or cytogenetic relapse. Patients with a short interval between transplantation and DLI have a higher probability of response than patients with longer intervals. Responses are the same with related versus unrelated donors.⁷¹¹ Some patients show a very rapid decline of BCR-ABL1 transcript levels (<6 months after DLI), whereas other patients demonstrate PCR negativity only over a longer period.⁷¹² The responses to DLI can be durable.⁷¹³ Molecular responses can occur in up to two-thirds of patients.⁷¹⁴

The main toxicities of DLI have been the induction of GVHD and myelosuppression. Chronic GVHD can occur in up to 60 percent of cases.⁷¹⁵ Attempts to diminish these toxicities have included use of CD8-depleted DLIs and infusion of smaller numbers of T cells.^716,717 Lower initial cell dose is associated with less myelosuppression, the same response rate, better survival, and less DLI-related mortality, leading to suggestions that the initial dose should not exceed 0.2 × 10⁸ mononuclear cells/kg.⁷¹⁸ The initial doses should be lower when matched unrelated donors are used. Immune suppression should first be tapered before DLIs are administered. As noted above, imatinib may synergize with DLI to foster rapid molecular responses after relapse.⁷¹⁹

Imatinib was first used experimentally for CML treatment in June 1998. Although it has completely altered the treatment approach to CML, its use has raised several questions. Studies require long-term followup of survival, but the degree of cytogenetic response, and degree of molecular response can be used as surrogate end points. The durability of cytogenetic and molecular responses in the face of persistent minimal residual disease during imatinib therapy require longer followup of larger numbers of patients, as more than 95 percent of cases have molecular evidence of disease at 2 years.

With the advent of TKI treatment, median survival in chronic phase CML is estimated to be 25 to 30 years. The prevalence of CML in the TKI era is estimated to reach a plateau 35 times the annual incidence with plateau estimated to occur in 2050. Therefore, the prevalence of CML is predicted to increase by a factor of 10.⁷²⁰ Using the Swedish Cancer Registry over a 36-year period, relative survival rates compared with the total population for CML improved from 0.21 in 1973 to 1979 to 0.80 for 2001 to 2008; imatinib was introduced in Sweden in 2001.⁷²¹ Another study from the Swedish CML registry of 779 patients showed that the mean survival ratio at 5 years was close to 1.0 for those younger than 60 years old and 0.9 for those 60 to 80 years old. Only 3 percent had progressed to accelerated or blast phase at 12 months.⁷²² Resistance rates decline with each passing year, and adverse effects have not emerged over time.⁷²³ Those who require 1 year or more of imatinib therapy to attain a complete cytogenetic remission have comparable rates of molecular response, progression-free, and overall survival to those who achieve a cytogenetic remission sooner.⁷²⁴ In patients with failure to respond or intolerance to imatinib the estimated 3-year survival rate was approximately 70 percent for patients in chronic phase. Survival in chronic phase was better when subsequent therapy was nilotinib or dasatinib as compared to allogeneic hematopoietic stem cell transplantation or to others agents. This result was at a median followup of 2 years.⁷²⁵ Older patients have lower response rates when treated in late chronic phase, but if they attain a CCyR, no difference was found in the level of molecular response.⁷²⁴ Table 89–7 shows the most recent 5-year relative survival data by age at diagnosis in the United States.

The introduction of imatinib has minimized the impact of prognostic factors at diagnosis of chronic phase CML.⁷²⁶ Several prognostic scales have been proposed in CML, including the Sokal and Hasford systems for patients at the time of diagnosis (Table 89–8). The European Bone Marrow Transplantation Consortium Risk Score was introduced for patients undergoing allogeneic hematopoietic stem cell transplantation,⁷²⁷ in which performance status is added to five variables (age, spleen size, blood blast cell count, basophil and eosinophil count, and platelet count [Hasford score]), and has been validated with good discrimination for survival.⁷²⁸ The Sokal score based on age, spleen size, platelet count, and percent blasts was developed much earlier during the busulfan era of treatment and was less accurate in patients treated with IFN. The European Treatment and Outcome Study (EUTOS) score uses basophils and spleen size but has not been well-validated. A simple prognostic scale that includes donor type, stage of disease at time of transplantation, age of recipient, sex of donor and recipient, and interval between diagnosis and transplantation has been proposed to predict outcome of allogeneic hematopoietic stem cell transplantation.⁷²⁹ These prognostic scales require revalidation given the dramatic impact of conversion to TKI therapy. There is evidence that a patient with chronic phase CML and a favorable Sokal Score at the time of diagnosis has a higher proportion of CHR and CCyR than other patients.⁷³⁰ Cytogenetic response and molecular response as a surrogate marker for survival is useful in patients undergoing TKI therapy.⁷³¹ The score of a patient may be calculated, easily, using an online website (Table 89–8). Studies suggest that in certain healthcare systems, healthcare setting may influence survival time, especially for those in advanced phases of the disease.⁷³²

Outcomes after allogenic hematopoietic stem cell transplantation have also improved in the imatinib era, but those in chronic phase have better 3-year survival rates than those in advanced phases (91 percent vs. 59 percent, respectively).⁷³³ There is favorable long-term survival after 5 years free of relapse after allogeneic hematopoietic stem cell transplantation.⁷³⁴ Prognostic factors in the TKI era for allogeneic transplantation are also being defined, and in addition to disease phase, donor-match, age, sex, and calculated comorbidity indices are of importance.⁷³⁵

DETECTION OF MINIMAL RESIDUAL DISEASE

Detection of minimal residual disease by molecular probes makes possible the identification of approximately one cell in 1,000,000 that is derived from the CML clone.⁷³⁶ Techniques used to monitor residual disease have been reviewed.^737,738 PCR permits observation of regression or persistence of subclinical disease following therapy and of progression of subclinical disease prior to the disease becoming overt, and it is therefore critical for monitoring responses to CML treatment.^739,740 The stable persistence of subclinical disease does not invariably predict early relapse.^741,742

Efforts are underway to standardize the technique for measuring and reporting real-time RT-PCR,⁷⁴³ and serial measurements are required for treatment decisions based on increases in transcript numbers.⁷⁴⁴ Some studies show that marrow values tend to be higher than blood in real-time RT-PCR assays, but both follow a similar trend during treatment.⁷⁴⁵ Interchanging these may lead to misinterpretation of disease status.⁷⁴⁵ In patients on imatinib, who have MMR, confirmed by real-time quantitative PCR, no marrow cell cytogenetic abnormalities were found, indicating that patients with MMR do not require regular marrow examinations for cytogenetics. The International Standard uses baseline diagnosis levels in the IRIS study as 100 percent and fixes a 3-log reduction from a standardized baseline (MMR) at 0.1 percent. Widespread use of the International Standard will require laboratories to send in specimens for analysis,^743,746 but is a priority in order to ascertain response to therapy accurately and to be able to analyze responses among treatment centers.

Patients who achieve a MMR (expressed as a 3-log reduction from median baseline value) at the time of achieving a CCyR have been found to have longer cytogenetic remissions than those without this magnitude of molecular response.⁷⁴⁹ The achievement of a 2-log molecular response at the time of a CCyR or a 3-log response anytime thereafter is an independent prognostic marker of progression-free survival.⁷⁴⁸ Other studies, however, show that patients who achieve CCyR do not derive additional benefit from a CMR.⁷⁴⁹ The treatment response, to imatinib, nilotinib, or dasatinib showed the 3-year event-free survival was 95 percent for those with BCR-ABL1 transcripts of 1 percent or less at 3 months, 98 percent for greater than 1 percent to 10 percent, and 61 percent for greater than 10 percent. These 3-month responses translated into overall survival of 98, 96, and 92 percent, respectively.⁷⁵⁰

Interphase FISH is not standardized, but a large number of cells can be rapidly analyzed (100 to 500). Fixation, specimen preparation, and hybridization conditions may account for differing false-positive ranges and scoring criteria.⁷⁵¹ In CML patients treated with imatinib, FISH for BCR-ABL1 on interphase blood neutrophils, but not unselected white cells, correlates with marrow cytogenetics.⁷⁵² FISH and RT-PCR can be useful complementary techniques,⁷⁵³ but FISH is generally not suitable for monitoring minimal residual disease.⁷⁵⁴

For patients who are undergoing allogeneic hematopoietic stem cell transplantation, the kinetics of minimal residual disease in either standard or nonmyeloablative transplants differ. BCR-ABL1/ABL1 ratios were 0.2 percent with reduced-intensity transplants versus 0.01 percent in transplantation patients with traditional conditioning regimens in the first 3 months. By 12 months, however, 20 percent of patients who received standard transplants and 50 percent of patients who received reduced-intensity transplants had reached a level less than 0.01 percent, supporting the concept of different kinetics of disease eradication between the two transplantation modalities.⁷⁵⁵ Patients who relapse after allografting have reappearance and/or rising levels of BCR-ABL1 transcripts.⁷⁵⁶ Use of quantitative RT-PCR early (3 to 5 months) after stem cell transplantation can project long-term outcomes.⁷⁵⁷ When RT-PCR was negative, the 3-year risk of relapse was 16.7 percent; when RT-PCR was positive at a ratio of less than 0.02 percent, the relapse rate was 42.9 percent; and when RT-PCR was positive at a level greater than 0.02 percent, the relapse rate was 86.5 percent. Another group found that detection of blood BCR-ABL1 at 18 or more months after transplantation was associated with a highly significant risk of relapse and that patients who had a positive test result but failed to relapse generally had only one positive test result at a low copy number.⁷⁵⁸ Performance of quantitative PCR (qPCR) at regular intervals after allogeneic transplant (every 2 to 4 months in the first year and every 6 months thereafter) is appropriate. If the PCR results are persistently positive or become positive, qPCR should be performed at monthly or shorter intervals. Molecular relapse is defined as a 10-fold increase of PCR positivity without any signs of cytogenetic relapse.⁷⁵⁹ Detection of increasing recipient chimerism by FISH for the male chromosome in sex-mismatched donor–recipient pairs or variable number of tandem repeats after allogeneic transplantation or after DLI infusion also is usually associated with a relapse.⁷⁶⁰

In an imatinib-treated patient, the absence of BCR-ABL1 transcripts should not be interpreted as an absence of the leukemic clone.⁷⁶¹ In terms of response to second-generation TKIs, there was no difference in event-free survival and CCyR duration between patients with CCyR with and without MMR up to 18 months with followup at 3-month intervals.⁷⁶² One study examined 116 patients with durable cytogenetic responses on imatinib who had increased PCR levels on at least two occasions. Only 9 percent of those had CML progression. Ten patients had lost MMR or had never had it, and all of these had more than 1-log increase in qPCR.⁷⁶³ With second-generation TKI, the BCR-ABL1 transcript levels at 3 months are also correlated with CCyR and MMR by 24 months, with levels less than 10 percent at 3 months being optimal.⁷⁶⁴ The European Leukemia Net criteria for failure or suboptimal response may also identify CML in early chronic phase treated with imatinib where the eventual outcome will be poor. These landmarks at 3, 6, 12, and 18 months are associated with poorer overall survival and cytogenetic responses.⁵⁰⁵ Only 60 percent of patients on the IRIS trial were in CCyR on imatinib after 6 years of therapy, indicating the need for close monitoring and for alternative therapies.⁷⁶⁵ There is also evidence that the BCR-ABL1 transcript doubling-time more reliably assesses CML relapse as compared to the fold rise in BCR-ABL1 transcripts. A short doubling-time for a patient in chronic phase should raise suspicion of nonadherence.⁷⁶⁶

DEFINITION

In all patients with chronic phase CML, the disease has the potential to evolve into a more aggressive, more symptomatic, and troublesome phase, which is poorly responsive to the therapy that formerly controlled the chronic phase. The failure of therapy to restore or maintain near-normal red cell and white cell counts, increased spleen size, increased numbers of marrow blasts and blood basophils, loss of the sense of well-being, and appearance of extramedullary tumors are the most consistent clinical hallmarks of the metamorphosis of the chronic to the accelerated phase of CML. The most objective findings are a blood blast percentage greater than 10, a platelet count less than 100 × 10⁹/L, blood basophils greater than 20 percent, and new clonal cytogenetic abnormalities accompanying the Ph chromosome.⁷⁶⁷

Several criteria have been published to define accelerated phase and blast crisis.^768,769,770 The terminology used has included accelerated phase, acute phase, acute transformation, or, in its most dramatic expression, blast crisis, but the metamorphosis, which can be acute, often is more gradual, hence the preference for transformation or accelerated phase to describe this transition from a controllable to a poorly controlled malignancy. Blast phase is the most severe manifestation of the accelerated phase and can occur abruptly or after a period of worsening disease. Blast crisis is in effect the evolution to overt acute leukemia, either myeloid or lymphoid.

PATHOGENESIS

Effect of Tyrosine Kinase Inhibitors in Rate of Progression

The advent of TKI therapy for CML has resulted in a marked increase in the duration of a subclinical chronic phase, with normal blood counts and spleen size, often with the loss of identifiable Ph chromosome-bearing cells in blood and marrow, and sometimes with the loss of laboratory evidence of the BCR-ABL1 oncogene as judged by PCR. This therapeutic advance has greatly delayed the evolution to accelerated phase and blast crisis, but the risk for such a conversion exists since under experimental conditions CML stem cells do not undergo apoptosis when exposed to BCR-ABL1 TKIs and BCR-ABL1–positive cells return in virtually all patients if tyrosine kinase therapy is interrupted. There is evidence that genomic instability may derive from an imatinib-refractory CML stem cell.⁷⁷¹

Blast Crisis Stem Cells

The onset of accelerated phase is thought to occur in a BCR-ABL1–bearing granulocyte-monocyte progenitor. Experimental^772,773 and theoretical⁷⁷⁴ evidence supports this concept. This progenitor for clonal evolution also could explain the reversion to chronic phase in some patients in whom the suppression of the advanced phase of the disease is achieved.

Molecular and Genetic Alterations

The transformation of chronic phase CML to accelerated and then blast crisis or directly to blast crisis is thought to be the result of seven molecular processes: (1) maturation arrest, (2) failure of genome surveillance, (3) failure of adequate DNA repair, (4) development of a mutator phenotype, (5) telomere shortening, (6) loss of tumor-suppressor function, and (7) unknown factors.^775,776

Progression of chronic phase to accelerated phase is marked by an increase in BCR-ABL1 expression.^777,778 Superimposed on the increased transcription of mRNA^BCR-ABL1 are additional cytogenetic abnormalities that are added to the persistent Ph chromosome in approximately 50 to 65 percent of patients.^779,780,781 In lymphoid blast crisis, in which the blast cells have a lymphocytic phenotype, acquisition of mutations in tumor suppressor genes, such as p16/ARF, occurs in approximately 50 percent of cases, and RB gene mutations occurs in approximately 20 percent of cases. In myeloid blast crisis, in which blast cells have a myeloid phenotype, approximately 25 percent of cases have cells containing a p53 mutation.⁷⁸² The possible role of loss of p53 function in fostering transformation of a human chronic phase CML clone has been demonstrated in transgenic mice in which p53 function was abrogated.^783,784 Progression of the clone to a more malignant clone is reflected in a more disordered growth and maturation pattern of progenitor cells in culture, ultimately mimicking the growth failure of acute leukemia,⁷⁷⁹ and in increased morphologic and functional abnormalities of blood cells,^785,786 eventuating in a block in maturation and replacement of blood and marrow by blast cells.

Approximately 65 percent of patients have cytogenetic abnormalities in addition to the Ph chromosome. A double Ph chromosome, trisomy 8, and isochromosome 17p are the secondary changes most commonly seen.^782,787 Because the frequency of trisomy 8 was greater after treatment with busulfan compared to hydroxyurea, the frequency of secondary chromosomal changes may be quite different after imatinib therapy.⁷⁸² Clonal instability has also been found in cases of lymphoid blast crisis. Clones distinct from those identified later may be detected before overt lymphoid transformation. Identification of these abortive clones suggests clonal instability before the onset of transformation, which might have prognostic value.⁷⁸⁸ FISH has been used to determine which cells have secondary cytogenetic abnormalities, and these cells often are not the blast cells. This finding suggests that some chromosomal abnormalities merely denote genomic instability.⁷⁸⁹ The abnormal mRNA and protein product p210^BCR-ABL1 are present in the marrow and blood cells of patients who have transformed to acute leukemia.^790,791,792

Although the breakpoint site on M-bcr was thought to be correlated with the time of the onset of the accelerated phase,⁷⁹³ subsequent studies have not indicated a correlation between length of chronic phase and the specific site of the BCR-ABL1 fusion.⁷⁹⁴ Rare cases have displayed deletion of the BCR-ABL1 fusion gene, loss of transcription of the message, and loss of expression of the p210 tyrosine kinase after transformation, the latter finding indicating the abnormal protein kinase may not always play a unique role in sustaining the acute state.⁷⁹⁵ In contrast, the frequent response, albeit temporary, to imatinib suggests that the mutant BCR-ABL1 product usually plays a role at this stage of the disease.

Numerous molecular changes identified in the cells of patients with acute transformation that might contribute to the increased malignant behavior of the CML clone, include activation of the N-RAS gene,^796,797 rearrangement of the p53 gene,^{797,798,799,800} hypermethylation of the calcitonin gene,⁸⁰¹ and methylation of the ABL1 gene.⁸⁰² One report described p53 mutations in 17 percent of blast crisis patients. An association between the failure of CML cells to express the RB1 gene product and acute blast crisis with a megakaryoblastic phenotype has been reported.⁸⁰² Homozygous deletions of the p16 gene are associated with lymphoid transformation of CML,⁸⁰⁴ but such deletions are not seen in the chronic phase and in myeloid blast crisis. p16 is also known as the cyclin-dependent kinase 4 inhibitor gene and is located on chromosome 9p21.^805,806 This gene inhibits the kinase CDK-4, which regulates a cell-cycle checkpoint prior to commitment to DNA synthesis. The Wilms tumor (WT) gene on chromosome 11p13 encodes a zinc finger motif-containing transcription factor found in CML patients only after progression to blast crisis.⁸⁰⁷ Overexpression of the EVI-1 gene has also been found in CML blast crisis.^808,809 Microsatellite instability has not been found to be involved with progression to blast crisis.⁸¹⁰ BCL-2, c-MYC, RUNX1, IKZF1, ASXL1, WT1, TET2, IDH1, NRAS, KRAS, CBL, and various other genes have also been implicated in the evolution of CML.^{811,812,813,814,815}

Approximately 50 genes have been identified that could play a role in the progression to accelerated phase or blast crisis,⁷⁷⁵ including genes identified by expression profiling that are dysregulated in accelerated phase compared to chronic phase.^816,817 These include the WNT-β-catenin and JunB pathways.

CLINICAL FEATURES

Signs and Symptoms

The features that might signal the conversion of the chronic to the accelerated phase include unexplained fever, bone pain, weakness, night sweats, weight loss, loss of sense of well-being, arthralgia, and left upper quadrant pain related to splenic enlargement or infarcts. These features may occur weeks in advance of laboratory evidence of the accelerated phase. Localized or diffuse lymphadenopathy or enlarging masses in extralymphatic and extramedullary sites containing BCR-ABL1–positive myeloblasts or lymphoblasts may develop. A poor response of blood cell counts and splenic enlargement despite previously effective therapy may be evident.^{767,782,818,819,820} Symptoms caused by histamine excess in basophilic crisis can be present.⁸²¹

Several of these changes may occur in series or in parallel. The time of onset of transformation and the appearance of a blast crisis and its clinical expression are unpredictable.

LABORATORY FEATURES

Blood Findings

Anemia may worsen and be associated with increasing poikilocytosis, anisocytosis, and anisochromia.^{782,818,819,820} The number of nucleated red cells in the blood may increase. These red cell changes may be accentuated further if advancing marrow fibrosis is a feature of the disease.

The total leukocyte count may fall without treatment. The proportion of blasts increases to greater than 10 percent in blood and marrow in the accelerated phase and when blast crisis ensues represents 20 to 90 percent of the cells. The morphology of the blast cells may be lymphoid or myeloid. Myelocytes decrease in number. Hyposegmented neutrophils (Pelger-Huët cells) and other dysmorphic changes may become evident. Basophils increase and often represent 20 to 80 percent of the total blood leukocytes. A decrease of the platelet count to less than 100 × 10⁹/L develops. Giant platelets, micromegakaryocytes, and megakaryocyte fragments may enter the blood. Decreased progenitor cell growth in culture is present, akin to that in acute leukemia.

Marrow Findings

The marrow findings are widely variable.^{782,818,819,820} Marked dysmorphic changes in one, two, or three of the major cell lineages; an increase in blast count to greater than 10 percent; marrow morphology simulating subacute myelomonocytic leukemia; or, in the extreme, florid blastic transformation with blast counts greater than 30 percent can occur. Reticulin fibers may increase in prominence, and occasionally severe reticulin and collagen fibrosis develop. Additional clonal cytogenetic abnormalities develop in as many as half the patients in accelerated phase (see “Cytogenetic Studies” below).

EXTRAMEDULLARY BLAST CRISIS

A variety of symptoms or signs may occur as a result of the specific effects of new extramedullary blastic tumors, referred to as extramedullary blast crisis.^{821,822,823,824} Extramedullary blast crisis is the first manifestation of accelerated phase in approximately 10 percent of patients with CML. Lymph nodes,^822,823,824 serosal surfaces,^825,826 skin and soft tissue,^{821,822,823,824} breast,^824,827 gastrointestinal or genitourinary tract,^822,824 bone,^{822,824,824,825,826,827,828,829,830,831} and central nervous system^{822,832,833,834,835,836} are among the principal areas involved. Isolated or diffuse lymphadenopathy may occur. Bone involvement may lead to severe pain, tenderness, and pathologic fracture, and may be evident on imaging of the involved area. Central nervous system involvement usually is meningeal and may be preceded by headache, vomiting, stupor, cranial nerve palsies, and papilledema and is associated with an increase in cells, protein, and the presence of blasts in the spinal fluid.^{824,832,833,834}

Appropriate histochemical and immunologic tests are required to determine if the extramedullary disease is composed of phenotypic myeloblasts or lymphoblasts. Because the tumor cells may have features of lymphoma cells, the terms myeloid or granulocytic sarcoma, chloroma, and myeloblastoma can be misnomers, and the term extramedullary blast crisis is used for this circumstance in CML.^{833,835,836,837} The lymphoblasts, like the myeloblasts, are Ph chromosome–positive. A combination of morphology, histochemistry (e.g., peroxidase, lysozyme), terminal deoxynucleotidyl transferase assay, and monoclonal antibodies specific for lymphoid or myeloid cells can be used to classify the extramedullary blast cells.

MARROW BLAST CRISIS

Approximately half of patients with CML enter the accelerated phase by developing acute leukemia. The onset of blast crisis can develop from days^838,839,840 to decades after diagnosis of CML. The signs and symptoms may include fever, hemorrhage, bone pain, and lymphadenopathy.^{776,838,839,840} The morphology of the acute leukemia usually is myeloblastic or myelomonocytic.^776,841 A substantial proportion of myeloid leukemia in this setting may not have myeloperoxidase demonstrable by cytochemistry.⁸⁴² The proportion of cases classified as erythroblastic leukemia is approximately 10 percent, based on morphologic features,⁸⁴³ but may be as high as 20 percent if expression of glycophorin-A is used as the determinant.⁸⁴⁴ Occasional cases have megakaryoblastic transformation.^803,845 These cases may be difficult to identify by light microscopy because the megakaryoblasts may be mistaken for lymphoid cells or undifferentiated blasts. Myelofibrosis is a feature of this variant. Antiplatelet glycoprotein antibodies and other monoclonal antiplatelet antibodies now are available as reagents to identify megakaryoblasts without the need for ultrastructural studies.⁸⁴⁵ Promyelocytic^846,847,848 and eosinophilic⁸⁴⁹ blast crises also can occur. Basophilic leukemia is a known variant of CML.⁸⁵⁰ Patients with promyelocytic crisis often have t(15;17) in addition to the Ph chromosome, and some have presented with disseminated intravascular coagulation.⁸⁵¹

CML may transform into acute lymphoblastic leukemia in approximately 30 percent of blastic crisis cases.^{767,852,853,854,855,856} The lymphoid cells generally express terminal deoxynucleotidyl transferase (TdT)^852,853 and are of the B-cell lineage,^856-878 as judged by antiimmunoglobulin staining. TdT is a DNA polymerase that adds deoxynucleoside monophosphates from triphosphate substrates to single-stranded DNA by end addition, differing in the latter respect from replicative polymerases.⁸⁵⁹ The enzyme is present in normal immature thymocytes and in the blast cells of nearly all patients with acute lymphoblastic leukemia. Rare patients have blasts with a T-lymphocyte phenotype.^{835,836,860,861,862} Some cases are biphenotypic; the blasts have both lymphoid and myeloid markers.^{841,863,864,865} Some cases may have myeloperoxidase activity in blast cells and express CD33 or CD13. Myeloid to lymphoid clonal succession following autologous transplantation in the second chronic phase has been described.⁸⁶⁶ Patients with lymphoid blast crisis seldom have an intermediate accelerated phase, have less splenomegaly and basophilia, and usually have a higher degree of marrow blast infiltration. With non-TKI therapy, remission rate and survival were somewhat longer in cases of lymphoid than in myeloid blast crisis.⁸⁶⁷

CYTOGENETIC STUDIES

Most large studies have shown seven recurrent changes in patients’ cells prior to, or during, the accelerated phase: trisomy 8 (33 percent of cases), additional 22q− (30 percent of cases), isochromosome 17 (20 percent of cases), trisomy 19 (12 percent of cases), loss of Y chromosome (8 percent of males), trisomy 21 (7 percent of cases), and monosomy 7 (5 percent of cases).^{867,868,869,870} In addition, a large number of other chromosome abnormalities have been described.^{871,872,873,874,875} In one study, 46 (63 percent) of 73 blast crisis patients had secondary cytogenetic abnormalities. These abnormalities were more common in myeloid blast crisis and were associated with shorter remission.⁷⁸⁸ The changes may be features of myeloid blast crisis compared to lymphoid crisis.^869,874 Some abnormalities, such as inv16, are associated with early transformation to AML.^{870,874,875,876,877} A significant proportion (50 percent) of patients in the accelerated phase or blast crisis have no additional cytogenetic abnormalities beyond t(9;22)(q34;q11) after banding and multicolor FISH analysis.⁸⁷⁶ In cases where the blastic transformation is in extramedullary sites, such as lymph nodes or spleen, the additional cytogenetic abnormalities may be in the cells at those sites but not in cells in the blood or marrow.⁸⁷⁸

TREATMENT

Optimal treatment is allogeneic stem cell transplantation if the patient is eligible based upon patient’s age and donor availability. The role of stem cell transplantation is also evolving as TKI use in these phases of diseases becomes better defined. Thus far, treatment with TKIs has improved survival only modestly in blast crisis, and most long-term survivors have been transplanted. At present, it is recommended that patients in blast crisis be treated with TKIs with or without chemotherapy to a second chronic phase and proceed to stem cell transplant as soon as possible once a donor is identified. One of the major goals of treatment of chronic phase CML is the prevention of evolution to accelerated or blast phases of the disease.

Tyrosine Kinase Inhibitors in Accelerated and Blast Crisis

The initial dose of imatinib in accelerated phase is 600 mg/day.⁸⁷⁹ Imatinib, dasatinib 140 mg/day, and nilotinib 400 mg BID, bosutinib 500 mg daily, and ponatinib 45 mg/day have been used as bridging therapies to permit allogeneic stem cell transplantation in accelerated phase.⁸⁸⁰ Dasatinib and nilotinib can achieve a better molecular response, and thus the role for and timing of transplantation in accelerated phase CML is being redefined, Imatinib can be combined with an anthracycline plus cytarabine for patients in myeloid blast crisis.⁸⁸¹ Imatinib has produced complete hematologic remissions in approximately 20 percent of patients.^882,883 However, CCyRs are uncommon. Central nervous system and other extramedullary blast crisis can occur during imatinib therapy for accelerated phase disease,^884,885 and all types of blast crisis, including promyelocytic blast crisis,⁸⁸⁶ can occur during imatinib therapy. Compared to historical controls in which various combinations of chemotherapy were utilized, imatinib used alone results in comparable outcomes (6-month median survival of patients in blast crisis).⁸⁸⁷ Although dasatinib therapy can result in CCyRs in 29 percent of blast crisis CML, and nilotinib can result in CCyRs in 27 percent of myeloid blast crisis and in 43 percent of lymphoid blast crisis, these responses are rarely durable, so in a patient of appropriate age with an acceptable donor, transplantation options should be considered with the second-line TKIs used as a bridge to transplantation therapy.^888,889 The choice of TKI in accelerated phase is based on prior therapy and/or mutational status. If response to TKI therapy is inadequate in accelerated phase, allogeneic hematopoietic stem cell transplantation should be considered. In lymphoid or myeloid blast phase, allogeneic hematopoietic stem cell transplantation is recommended if there is a suitable donor.⁴¹⁷ Omacetaxine has shown activity in patients with disease progression to accelerated phase CML after prior TKI use.⁸⁹⁰

Chemotherapy

The treatment approach is predicated on the phenotype of the blast cells in CML patients with blast crisis and is rarely used in accelerated phase. In patients with myeloid phenotypes, the approach has been similar to that used for AML: combinations of an anthracycline antibiotic, such as idarubicin or daunorubicin, with cytosine arabinoside and sometimes etoposide.⁸⁹¹ Because this approach produces few remissions that are of short duration (median survival approximately 6 months), a variety of other drug combinations have been used, but with no significant improvement in outcome.⁸⁸⁷ Complete hematologic response is observed in only a quarter to a third of patients with this approach.

In patients with lymphoid phenotypes, vincristine sulfate 1.4 mg/m² (not to exceed 2 mg/dose) given intravenously once per week and prednisone 60 mg/m² per day given orally are the mainstay of treatment. A minimum of two cycles of treatment (2 weeks) should be given to judge responsivity. Other more intense ALL-type regimens, such as HyperCVAD (cyclophosphamide, vincristine, doxorubicin, and dexamethasone),⁸⁹² have also been used. Approximately one-third of patients with lymphoid blast transformation reenter the chronic phase after such treatment. However, because only about one-third of patients have lymphoid blasts, this number represents a remission rate of only approximately 10 percent of patients who enter blast crisis using this approach. The benefit of intensive chemotherapy has been small because remission durations have been modest. TdT-positive, CD10 (CALLA)-positive lymphoblasts may be the lymphoblast phenotype most responsive to vincristine and prednisone.⁸⁵⁵ mTOR inhibitors may be useful in lymphoid blast crisis.⁸⁹⁷

Use of Chemotherapy with Tyrosine Kinase Inhibitors in Accelerated Phase or Blast Crisis

Imatinib has been successfully combined with decitabine, low-dose cytarabine, and standard anthracycline plus cytarabine (“7 + 3”) where complete remission can be as high as 75 percent. Median time to relapse and overall survival are brief, however.⁸¹⁵ Imatinib and dasatinib have been combined with HyperCVAD in lymphoid blast crisis.^892,893

Allogeneic Hematopoietic Stem Cell Transplantation

Stem cell transplantation from an appropriately HLA-matched donor has been used in some patients after entry into the blastic crisis. Occasional patients have had long-term survival. The 3-year survival rate is approximately 15 to 20 percent,^894,895,896 unlike transplantation in the chronic phase, in which the 3-year survival rate is 50 to 60 percent. However, for patients who present in blast crisis, who develop blast crisis in the first year of the chronic phase, or who delay transplantation for other reasons, transplantation remains the best hope for long-term survival if a histocompatible donor is available.^832,833 Relapse of accelerated phase after allogeneic stem cell transplantation has responded to infusion of donor cytotoxic T lymphocytes.⁸⁹⁷ The use of TKIs before allogeneic stem cell transplantation for patients in advanced phases of disease may favorably improve transplantation outcomes, especially when MCyR occurs before transplantation.⁸⁹⁸

Autologous Hematopoietic Stem Cell Transplantation

Autografting in the accelerated phase or blast crisis, either with stem cells collected during chronic phase or with mobilized Ph chromosome–negative progenitor cells collected upon cell rebound after intensive chemotherapy, has resulted in apparent prolonged remission in some patients, but this procedure is rarely used in advanced disease because of the high rate of relapse.⁸⁹⁹ Whether Ph chromosome–negative cells collected during imatinib therapy have the same potential is unknown.

Splenectomy

Splenectomy may be performed for palliation of painful splenic infarctions or hemorrhage. However, the complication rates are high, and the procedure performed in this setting should be avoided if possible.⁹⁰⁰

COURSE AND PROGNOSIS

The accelerated phase of CML generally is very poorly responsive or refractory to treatment and is a morbid state that can be fatal in weeks to months in all but a few patients who undergo a successful stem cell transplant from a histocompatible donor. Patients with myeloid blast crisis have a median survival of approximately 6 months, whereas patients with lymphoid blast crisis have a median survival of approximately 12 months.^887-901 In the earliest study of imatinib in blast crisis, patients with myeloblastic crisis had a slightly longer median survival (approximately 5 months) than patients with lymphoid blast crisis plus Ph chromosome–positive ALL (3 months). The results are poor in either case. A worse survival was seen with abnormalities of chromosome 17, other superimposed translocations, or a high percentage of abnormal metaphases.⁹⁰² Severe cytopenias from repeated courses of cytotoxic therapy contribute to infections, hemorrhage, and organ dysfunction, especially liver and kidney dysfunction. Opportunistic infections with herpes viruses, cytomegalovirus, or fungi often supervene. The addition of imatinib or other TKI therapies has made only a small difference in long-term outcome, although formal studies of combinations of chemotherapy and imatinib at a higher dose (600 mg/day) have not been completed, and the results of trials with second-generation TKIs are anticipated.

In addition to classic CML with the BCR-ABL1 fusion gene, there are several other chronic myeloid leukemias which are listed in Table 89–9.

CHRONIC MYELOMONOCYTIC LEUKEMIA

This leukemia is part of the spectrum of clonal myeloid diseases that may have findings that simulate CML. In the past, when rigorous criteria for the diagnosis of CML were not applied, CMML was among a heterogenous group of related diseases that sometimes were referred to as Ph chromosome–negative CML. These diseases share the feature of originating in the clonal expansion of a primitive multipotential hematopoietic cell.⁹⁰³

Epidemiology

Most patients with CMML are older than age 50 years, the median age is approximately 72 years, and approximately 90 percent of patients are older than age 60 years at the time of diagnosis.⁹⁰⁴ Occasional cases have been reported in older children and younger adults. Men are affected more frequently than women (approximately 2:1).^904,905,906 An evaluation of exogenous factors that might increase the incidence of CMML, did not find an association with benzene or other occupational or non-occupational risk factors.⁹⁰⁷

The disease may occur following therapy for an unrelated malignancy, most commonly lymphoma, breast, or prostate cancer. The prior therapy was radiation, combined radiation and chemotherapy, or chemotherapy, alone. The median time of onset of CMML was 6 years.⁹⁰⁸

Clinical Findings

Signs and Symptoms The onset usually is insidious, and weakness, infection, or exaggerated bleeding may bring patients to medical attention.^905,906 Hepatomegaly and splenomegaly occur in approximately 50 percent of patients. Leukemia cutis occurs in a small proportion of patients and the skin cellular infiltrate is usually has a monocytic phenotype: CD45, CD68, and lysozyme positive by immunostaining. Immune manifestations, such as vasculitis, pyoderma gangrenosum, immune cytopenias, and connective tissue diseases, may occur in coincidence with CMML.⁹⁰⁹

Blood and Marrow Findings The disease is characterized by anemia and blood monocytosis greater than 1 × 10⁹/L.⁹¹⁰ The white cell count may be decreased, normal, or moderately elevated. In one study of 275 patients, the range in 247 informative patients was 0.9 to 160 × 10⁹/L.⁹⁰⁴ Occasional patients, however, may have hyperleukocytosis with total white cell counts of 250 to 300 × 10⁹/L associated with respiratory insufficiency resulting from pulmonary leukostasis.⁹¹¹ Promonocytes and monocytes are present in blood and may have dysmorphic features. Immature granulocytes (promyelocytes and myelocytes) may be present in the blood. Blood myeloblasts are absent in approximately 75 percent of patients or, when present, usually do not exceed 10 percent of total white cells. Most patients have thrombocytopenia, but normal or elevated platelet counts may occur (range: 3.0 to 1385 × 10⁹/L among 227 patients in one series).⁹⁰⁴ Eosinophilia may be so prominent in occasional cases that the designation chronic eosinophilic leukemia may be appropriate.^905,906,912

The marrow is hypercellular as a result of granulomonocytic hyperplasia; the dominant cells are early myelocytes. Blasts cells are 1 to 4 percent in about two-thirds of patients and are from 5 to 19 percent in one-third of cases.⁹⁰⁴ The proportion of promyelocytes is increased. Promonocytes and monocytes also are increased in number. Distinction between poorly granulated myelocytes and promonocytes with primary granules can be difficult. Macronormoblasts and hypersegmented or hyposegmented, often bilobed (acquired Pelger-Huët anomaly), neutrophils may be evident but are more frequent in cases with lower white cell counts. Despite thrombocytopenia, megakaryocytes usually are present in the marrow. Microvessel density is increased in the marrow, and myelomonocytic cells contain cytoplasmic mRNA for VEGF and membrane VEGF receptors.^912,913 In vitro colony studies suggest that autocrine stimulation of cell growth by VEGF may occur. “Spontaneous” cluster/colony growth of granulocyte-monocyte colony-forming cells occurs in vitro. The spontaneous growth may result from autocrine or paracrine production of GM-CSF, based on anti–GM-CSF inhibition of colony growth.⁹¹⁴

Cytogenetic and Genetic Findings Patients with CMML have an approximately 35 percent frequency of chromosomal abnormalities. Trisomy 8 and, to a lesser extent, monosomy 7 and −Y are the most prevalent findings. A low–risk karyotype is either a normal pattern or isolated −Y, an intermediate-risk is any other abnormality, and high-risk is trisomy 8 or complex karyotypes (more than three abnormalities). Approximately 35 percent of patients have point mutations of the K-RAS or N-RAS gene.⁹⁰⁴ The RAS gene also may be involved in the transforming events. Abnormal methylation of p15^INK4B is a common finding in CMML.⁹¹⁵ Translocation between the gene PDGFR-β on chromosome 5(q33) and four partner genes—TEL at 12(p13), HIP-1 at 7(q11.2), H4 at 10(q22), and Rabaptin-5 at 17(p13)—occur in a very small proportion of patients (approximately 3 to 4 percent).^{905,916,917,918,919} This mutation juxtaposes the gene encoding the PDGFR-β with a partner gene, which results in the encoding of a mutant tyrosine kinase that is constitutively activated and sensitive to inhibition by imatinib or a congener⁹²⁰ (see “Treatment” below). The cases with PDGFR-β translocations are more likely to be accompanied by eosinophilia than are cases with other cytogenetic abnormalities. Somatic mutations in CMML cells include SRSF2, TET2, ASXL1, RUNX1, SETBP1, KRAS, EZH2, CBL and NRAS, as well as the novel CMML genes FAT4, ARIH1, DNAH2 and CSMD1, each mutated in ≥ 10 percent of patients. Other gene mutations identified in patients' CMML cells include IDH1/2, JAK2, DNMT3A, UTX, and several others. The gene SRSF2 (serine/arginine-rich splicing factor 2), was found to be the most frequently mutated gene in patients with CMML. Many of the genes commonly mutated in CMML have been associated with age-related clonal hematopoiesis (ARCH). Most CMML patients (71%) had mutations in ≥2 ARCH genes and 52 percent had ≥7 mutations overall. Higher mutation burden was associated with shorter survival. ARCH is characterized by a myelomonocytic differentiation bias. These findings are consistent with a model in which clinical CMML ensues when a sufficient number of stochastically acquired age-related mutations has accumulated. CMML appears to represent the leukemic conversion of the myelomonocytic-lineage-biased aged hematopoietic system.^920a In an effort to determine the frequency of the mutation in a large sample of patients and to look for coincident mutations with SRSF2 in CMML patient’s cells, eight other genes known to be mutated in some patients with CMML were studied among 275 patients with the disease.⁹⁰⁴ SRSF2 was mutated in 47 percent of patients in this large series, and had a significantly increased coincidence with EZH2 and TET2. Ninety-three percent of patients had at least one mutated gene.

Serum and Urine Findings Plasma and urine lysozyme concentrations nearly always are elevated. Plasma levels of VEGF, hepatocyte growth factor, and tumor necrosis factor alpha are elevated. Serum vitamin B₁₂, β₂-microglobulin, and LDH levels often are elevated.^905,906

Treatment

Treatment of most patients with CMML has been unsatisfactory, and remissions of any duration are uncommon. The age and performance status of the patient are considered in determining the intensity of treatment. Cytarabine, either standard or low-dose, etoposide, hydroxyurea, and other approaches used for the oligoblastic myelogenous leukemias have been attempted, but with little success (Chap. 88). Decitabine and 5-azacytidine have been useful in a small proportion of patients.^905,906 One study of azacytidine treatment reported a complete response in 11 percent, a partial response in 3 percent, and hematologic improvement in 25 percent. The median survival of responders was 15 months compared to 12 months among nonresponders, a modest result.⁹²¹ Unfortunately, although a particular approach may confer significant benefit in a small proportion of cases, determining which patients will respond is not possible, except by trial and error. An exception is the patient with a translocation involving PDGFR-β, which itself occurs in only a small percentage of patients. In the case of PDGFR-β fusion genes with several of the partner genes, imatinib 400 mg/day has resulted in normal blood counts, cytogenetic remissions, and, occasionally, molecular remission.^{915,916,917,922,923} These fortunate patients probably will benefit from this treatment, as evidenced by prolonged remissions and survival, compared to other drug options, but the number of patients and duration of followup do not permit quantitative estimates at this point. Allogeneic stem cell transplantation is an option for the small proportion of younger patients with an appropriate matched-related or unrelated donor.⁹²⁴

Course and Prognosis

Median survival in CMML is approximately 12 months, with a range from approximately 1 to more than 60 months. Approximately 20 percent of patients progress to frank AML. Arbitrary stratification of CMML into types 1 (<5 percent blood myeloblasts) and 2 (5 to 20 percent blood myeloblasts) based on the height of the blast count has been proposed, but distinguishing patients by whether, for example, they have 4 (type 1) or 8 percent blasts (type 2) on a marrow examination is of little use for care of an individual patient. Blast percentage may be a significant correlate with outcome in any large group of patients with a clonal myeloid disease, and this factor among several others should guide therapy. Clusters of prognostic variables have been used to stratify patients into risk groups for survival duration. In general, the severity of the anemia, the cytogenetic risk category, and the height of the blast percentage are the most important. Other variables that may confer a shorter life expectancy are height of the absolute lymphocyte count, high spontaneous rates of myelomonocytic colony growth, higher total leukocyte counts, higher LDH level, and larger spleen size.^925,926,927 These risk factors for progression are validated in large groups of patients but the confidence intervals are not shown. In an individual patient the variability in outcome is great based on such variables; careful clinical observation for progression is most important. Unfortunately, at this time, unless the patient is a candidate for imatinib therapy or allogeneic stem cell transplantation, long-term salutary therapeutic effects are uncommon.

CHRONIC EOSINOPHILIC LEUKEMIA

History and Definition

The recognition of eosinophilic lineage prominence in myelogenous leukemia dates to a case published in 1912.⁹²⁸ In 1968, the term hypereosinophilic syndrome was introduced to encompass a group of disorders with (1) prolonged exaggerated eosinophilia without an apparent cause, (2) frequent cardiac and neurologic tissue damage, (3) a poor or transient response to therapy, and (4) a progressive course and a high fatality rate. Shortly thereafter, Benvenisti and Ultmann⁹²⁹ presented five cases of eosinophilic leukemia and reviewed the literature regarding that phenotypic designation. In 1975, Chusid and colleagues⁹³⁰ described 14 cases of hypereosinophilic syndrome, highlighted the frequency of secondary cardiac and neurologic disorders, and suggested the existence of a continuum of manifestations. Because some cases had clonal cytogenetic abnormalities and hematologic findings compatible with a clonal myeloid disease, the presence of eosinophilic leukemia was suspected in this apparently heterogeneous group of patients.

The relationship of blood eosinophilia to clonal myeloid diseases is complex because the former can be reactive or represent acute eosinophilic leukemia, chronic eosinophilic leukemia, or eosinophilia associated with a different category of disease, such as BCR-ABL1–positive CML, primary myelofibrosis, oligoblastic leukemia (MDS), or mastocytosis.⁹³¹ Chronic eosinophilic leukemia is a BCR-ABL1–negative, clonal myeloid disease with a striking eosinophilia in the blood and marrow, often with clonal cytogenetic abnormalities that have features including, when present, cytogenetic findings that usually distinguish chronic eosinophilic leukemia from other clonal myeloid diseases that may have an associated eosinophilia, such as CMML. The phenotype of the eosinophilic variant of CMML overlaps somewhat with that of chronic eosinophilic leukemia. However, the fusion gene associated with CMML involves the PDGFR-β (see “Chronic Myelomonocytic Leukemia” above), whereas in chronic eosinophilic leukemia the cytogenetic findings are different and in some cases involve PDGFR-α. This definition of chronic eosinophilic leukemia recognizes that, at the margins, classification may be arbitrary. Studies in cases with the FIP1L1–PDGFR-α translocation were consistent with multilineage involvement, consistent with an origin in a pluripotential lymphohematopoietic cell.⁹³² Another report found multilineage involvement but with a presumptive lesion in a multipotential, not pluripotential, hematopoietic cell.⁹³³

Signs and Symptoms

Fever, cough, weakness, easy fatigability, dyspnea, abdominal pain, maculopapular rash, cardiac symptoms and signs of heart failure, and a variety of neurologic manifestations ranging from peripheral neuropathy to cerebral encephalomalacia may occur, ranging from mild to severe in expression. Splenomegaly often is evident.

Laboratory Findings

Eosinophilia is a constant finding (see Chap. 62, Fig. 62–3). Anemia usually but not always is present at the time of presentation. The leukocyte count may be high-normal or more often elevated. Platelet counts often are normal or mildly decreased. The marrow shows myelocytic and eosinophilic hyperplasia and, occasionally, Charcot-Leyden crystals. Mast cells, often spindle-shaped, may be increased. In the FIP1L1–PDGFR-α type of chronic eosinophilic leukemia, which may make up approximately 14 percent of cases of primary eosinophilia not found to be reactive to another disease, marrow aggregates of spindle-shaped mast cells are invariably found (see “Cytogenetic Findings” below).^933,934 Megakaryocytes usually are present but may appear dysmorphic. Reticulin fibrosis is common. Immunophenotyping and PCR do not show evidence of either a clonal T-cell population or T-cell–receptor rearrangement. Pulmonary function studies may provide evidence of fibrotic (restrictive) lung disease. Echocardiography may detect mural thrombi, thickening (fibrosis) of the ventricular wall, valvular dysfunction from papillary muscle, and chordae fibrosis. Magnetic resonance imaging can detect subendocardial fibrosis, thickening of ventricles, and markedly reduced ventricular lumen volume. Serum immunoglobulin (Ig) E, vitamin B₁₂, and tryptase levels usually are elevated. Skin biopsy of lesions uncovers intense eosinophilic infiltrates. Neural or brain biopsy may disclose eosinophilic infiltrates, often perivascular, with microthrombi, axonal degeneration, and gliosis.

Cytogenetic Findings A wide array of cytogenetic findings have been reported in cases of chronic eosinophilic leukemia.⁹³⁵ Notable translocations include a high frequency of translocations involving chromosome 5, t(1;5), t(2;5), t(5;12), t(6;11), 8p11, trisomy 8, and numerous others, infrequently. Chromosome 5 often is translocated at the site of the PDGFR-β gene, and the phenotype usually is more compatible with CMML with eosinophilia. Chromosome 5 from band q31–35 contains several genes relevant to eosinophilopoiesis, including those encoding IL-5, IL-3, GM-CSF, and PDGFR-β. A cryptic interstitial CHIC2 deletion on chromosome 4 (q12;q12) results in the fusion gene FIL1L1–PDGFR-α, normally separated by the CHIC2 gene, and in a phenotype that can be considered a form of chronic eosinophilic leukemia, virtually always associated with marrow mastocytosis, which is of particular note because, like the PDGFR-β mutations in CMML with eosinophilia, of a near-universal response to treatment with imatinib.^935,936,937

Serum Tryptase Level Elevation Versus Normal Levels

The elevation of serum tryptase level (>11.5 ng/mL) has been used to distinguish a subset of patients who (1) are male, (2) have marrows that are intensely hypercellular with a higher proportion of immature eosinophils and with dysmorphic mast cells with a CD117−CD25+CD2− genotype and phenotype (distinguishing these cells from classic mastocytosis, which are CD117+CD25+CD2+), (3) have dramatically higher serum vitamin B₁₂ and IgE levels, (4) are more prone to restrictive pulmonary disease and endomyocardial fibrosis, (5) have the FIP1L1–PDGFR-α fusion gene, and (6) are responsive to imatinib.⁸⁶⁹ Patients with normal serum tryptase levels are more prone to obstructive pulmonary restrictive disease, eosinophilic dermatitis, and gastrointestinal complaints.

Differential Diagnosis

Eosinophilia can occur for many reasons (Chap. 62). The first step is to identify signs that may point to a clonal myeloid disease. These signs include anemia, thrombocytopenia, splenomegaly, immature eosinophils in the marrow examination, evidence of dysmorphic cells in blood or marrow, for example, atypical megakaryocytes or dysmorphic mast cells, cardiac or pulmonary manifestations, which may occur secondary to chronic eosinophilic leukemia, and markedly elevated serum tryptase or vitamin B₁₂ level. The former signs, especially in the aggregate, are highly suggestive, but the presence of a cytogenetic abnormality in myeloid cells is diagnostic of a clonal myeloid disease (leukemia). If the latter is not evident, PCR and/or flow cytometry to search for a clonal T lymphocyte abnormality should be performed. Whether the eosinophilic leukemia is typical or represents an eosinophilia with idiopathic myelofibrosis, CMML, or MDS is less important than if it has a mutation that is imatinib sensitive (e.g., PDGFR mutation).

Therapy

Patients (nearly always men) whose cells display a FIP1L1–PDGFR-α have a very high probability of responding to imatinib at a dose of 100 to 400 mg/day.^{936,937,938,939,940} The tyrosine kinase activity of this fusion protein is two orders of magnitude more sensitive to imatinib than that of BCR-ABL1. However, because not all patients taking 400 mg/day achieve a molecular remission, and that goal may be more likely to result in long-term remission, initial therapy remains at 400 mg/day or an equivalent dose of another TKI, such as dasatinib or nilotinib, and PCR monitoring is appropriate. Dose adjustment upward if molecular remission is not achieved can be considered. Unlike the case in CML, patients with chronic eosinophilic leukemia with significant side effects when taking imatinib, 400 mg/day, have a reasonable probability of having a good response at lower doses.⁹⁵⁰ Dasatinib and nilotinib are also active.⁹⁴¹

In patients with eosinophilic leukemia without an imatinib-sensitive mutation or in patients who become resistant to imatinib and unresponsive to a second-generation TKI (e.g., dasatinib or nilotinib) and who are progressing, ablative or nonablative allogeneic stem cell transplantation can be considered if they are in an acceptable age range and have access to a matched-related or matched-unrelated donor.^942,943

In TKI-insensitive patients without the option of transplantation, empirical treatment with glucocorticoids, hydroxyurea, or anti–IL-5^944,945 to decrease eosinophil counts and mute the progress of eosinophil-mediated cutaneous, cardiac, pulmonary, and neurologic tissue damage should be considered. These approaches may relieve symptoms for a time, but are temporizing if therapy with drugs that might be effective in inhibiting clonal expansion or evolution is not effective (e.g., cytarabine, anthracycline antibiotic, etoposide).

Course and Prognosis

If chronic eosinophilic leukemia is not TKI-sensitive, the long-term outlook is one of probable progressive cardiac and neurologic disability. Transformation to acute eosinophilic or myelogenous leukemia can occur. Allogeneic stem cell transplantation is potentially curative. In tyrosine kinase sensitive cases, hematologic normalization, reversal of marrow fibrosis and mastocytosis, resolution of skin lesions, normalization of spleen size, and restoration of well-being occurs in the great preponderance of cases. Cardiac, neurologic, and pulmonary changes usually cannot be reversed but should be stabilized. The long-term outlook with tyrosine kinase therapy is uncertain. However, it likely will decisively and dramatically improve survival compared to other prior therapy and should greatly improve the prognosis of patients with a molecular target for the drug.

CHRONIC BASOPHILIC LEUKEMIA

This type of clinical disorder, in which the patient has marrow and blood basophilia and other findings compatible with a clonal myeloid disease without evidence of the BCR-ABL1 translocation, is rare. Two reports of such a syndrome occurring in five patients have been published.^946,947 The marrow was intensely hypercellular in the three major lineages. Dysmorphic megakaryocytes were evident. Basophilia in marrow and blood was striking, although eosinophilia also was evident in two patients and increased mast cells in three patients. The clinical effects of basophilic mediator release were evident in two patients. One patient evolved to AML; another recovered after allogeneic transplantation. The cases had similar findings, leading to the suggestion they represented Ph chromosome–negative chronic basophilic leukemia. In one case, a PRKG2–PDGFR-β fusion gene was evident and the patient responded to imatinib.

JUVENILE MYELOMONOCYTIC LEUKEMIA

Epidemiology

Ph chromosome–positive, adult-type CML occurring in children younger than age 15 years makes up approximately 3 percent of childhood leukemias and approximately 10 percent of all cases of CML.⁹⁴⁸ Although CML occurs in children of all ages, it is rare in children younger than age 5 years. With the exception of a propensity to present with higher total leukocyte counts and with leukostatic signs or symptoms, CML in children has the typical manifestations and course of the disorder seen in adults.

A disorder different from adult-type CML, designated juvenile myelomonocytic leukemia, represents approximately 1.5 percent of childhood leukemias. It occurs most often in infants and children younger than age 4 years and is similar in some respects to adult subacute or CMML because the two diseases share a prominent monocytic component in the leukemic cell population.^{949,950,951,952}

Pathogenesis

This disorder is a clonal myeloid disease that originates in an early hematopoietic multipotential cell. Evidence indicates this cell may be pluripotential (myeloid-lymphoid) in some cases and myeloid in others.^{953,954,955,956} Patient-derived induced pluripotential stem cells recapitulated the growth patterns in vitro of the human disease and drug inhibition of MEK kinase reduced their GM-CSF growth potential.⁹⁵⁷ RAS mutations in hematopoietic cells are present in approximately 20 percent of patients.⁹⁵⁸ Approximately one in 10 patients with juvenile myelomonocytic leukemia have mutations of NF1 and manifest type 1 neurofibromatosis. This frequency is approximately 400 times the expected occurrence in a comparable pediatric population.^959,960,961 The linkage between neurofibromin, the protein encoded by the NF1 gene, GTPase activity proteins, and the activation state of RAS-encoded proteins has led to a postulated sequence of events that may be triggered by the extraordinarily heightened sensitivity of the colony-forming cells in the marrow and blood of infants with the disease to the proliferative effects of GM-CSF. The latter initiates signal transduction from the cell membrane to the nucleus via RAS protein activation.^961,962 Mutations in the PTPN11 gene have been found in approximately one-third of children with juvenile myelomonocytic leukemia, and the mutations in NF1, RAS, and PTPN11 usually do not coincide.^1025,1026 However, they each may act through a common pathway. PTPN11 encodes SHP-2, a phosphatase, which is an upstream regulator of RAS; thus, all three mutations can contribute to deregulation of RAS signaling. As an aside, children with Noonan syndrome, which is characterized by short stature, dysmorphic facies, skeletal abnormalities, and cardiac defects, have a germ-cell mutation of PTPN1. These children may have a transient disorder that closely mimics juvenile myelomonocytic leukemia.⁹⁶³

Clinical Findings

Symptoms and Signs Infants present with failure to thrive, and children present with malaise, fever, persistent infections, and exaggerated skin, oral, or nasal bleeding. Hepatomegaly can occur. Splenomegaly, sometimes massive, is present in almost all cases. Lymphadenopathy is frequent.^{949,950,951,952} More than half of the patients have eczematoid or maculopapular skin lesions⁹⁶⁴ and xanthomatous lesions, and multiple café-au-lait spots (neurofibromatosis type 1) may occur.⁹⁵⁰ The xanthomas may be the earliest signs of neurofibromatosis.^950,951 Noonan syndrome (dysmorphic facies, short stature, heart disease, mental retardation, cryptorchidism, webbed neck, chest deformities, and bleeding diathesis) may coexist.⁹⁵¹

Laboratory Findings Anemia, thrombocytopenia, and mild to moderate leukocytosis are common. The leukocyte count usually is greater than 10 × 10⁹/L with a median leukocyte count at diagnosis of approximately 35 × 10⁹/L. The blood has an increased monocyte concentration of 1 to 100 × 10⁹/L, immature granulocytes including a small percentage of blast cells, and nucleated red cells. Fetal hemoglobin concentration is increased in approximately two-thirds of the patients. The marrow aspirate is hypercellular as a result of granulocytic hyperplasia; the number of erythroblasts and megakaryocytes usually are decreased. Monocytic cells are increased but may not be as striking as in the blood. Leukemic blast cells are present in modest proportions (<20 percent).

Cell culture of blood and marrow shows a striking preponderance of monocytic progenitors, even in the absence of overt monocytosis in the marrow.^965,966 Granulocyte-monocyte colony-forming cells show a marked tendency to spontaneous growth if adherent (monocytic) cells are not depleted from culture.⁹⁶⁶ The effect is mediated by a release of large quantities of GM-CSF by monocytes in culture.⁹⁶⁷

Although clonal chromosome abnormalities have been found in some cases,⁹⁶⁸ the cytogenetic abnormalities have no consistent pattern, and more than half of the patients have normal karyotypes. The BCR-ABL1 fusion gene is not present.^968,969,970 The phenotype of monosomy 7 syndrome overlaps with juvenile myelomonocytic leukemia, and an abnormality of chromosome 7 (del 7, del 7q, others) is present in approximately one-fifth of patients.⁹⁴⁹

Course, Prognosis, and Treatment

The median survival of patients with juvenile myelomonocytic leukemia has been less than 2 years.^949,950 Children younger than 2 years are more likely to have a protracted course.⁸⁸² The disease has been refractory to most chemotherapy. In a study of nine patients, four of whom were treated with a five- or six-drug intensive regimen, remissions were 11 months to more than 27 months, compared with untreated or lightly treated patients, four of whom died within 7 months.⁹⁷¹ Even in the treated patients, complete suppression of the disease did not occur, and treatment protocols to induce and sustain remissions were lacking.⁹⁶⁶ A program of cytosine arabinoside, etoposide, vincristine, and isotretinoin resulted in a highly favorable response in five children treated. Three patients relapsed and were treated with cytarabine by infusion and subcutaneously and with etoposide. All patients were alive, and the range of survival at the time of publication was 8 to 89 months, with a median survival of 27 months. The resistance of these cells to currently available therapy is distressingly highlighted by the sense of success in prolonging the life of infants and young children by a few years. Intensive therapy can control disease, but curative chemotherapy has been elusive.⁹⁷² The inclusion of isotretinoin was based on a prior report of responsiveness to the drug used alone; however, this observation has not been confirmed.⁹⁷³ The GM-CSF antagonist E21R, inhibitors of RAF-1 gene expression, blockers of RAS protein farnesylation, and angiogenesis inhibitors are among other drug approaches to the disease being studied.^952,974

Allogeneic stem cell transplantation is an important approach to therapy and may provide the best chance of long-term survival in selected children.^975,976,977 Hence, a rapid search for a matched-unrelated donor, including cord blood sources, is important in patients without matched sibling donors. Transplantation from a histocompatible sibling or matched-unrelated donor resulted in an event-free survival at 5 years of approximately 50 percent, and from matched-cord blood stem cells of approximately 45 percent, unless monosomy 7 was present, which lowers 5-year survival to approximately 25 percent.⁹⁷⁷ Children transplanted before age 1 year had better results (approximately 50 percent) than did older children (approximately 30 percent).⁹⁷⁷ DLI has placed a posttransplantation patient in relapse into remission,⁹⁷⁸ but, in general, this procedure is ineffective in patients who relapse after transplantation.

A minority of patients have a smoldering course for 2 to 4 years. Thereafter, the disease usually rapidly progresses, and patients die of infection or hemorrhage. Occasional patients have a very long survival (>10 years) despite persistence of abnormal blood counts and splenomegaly, independent of the type or intensity of therapy. Some children convert to a full-blown AML with a rapidly fatal outcome. Cases of juvenile myelomonocytic leukemia may be associated with transformation to acute lymphoblastic leukemia.⁹⁷⁹

CHRONIC NEUTROPHILIC LEUKEMIA

History, Pathogenesis, and Epidemiology

In 1920, Tuohey⁹⁸⁰ described the first recorded case of an unusual sustained neutrophilia with splenomegaly without fever, inflammation, cancer, or other cause of a leukemoid reaction. Use of X-chromosome-linked polymorphic genes in blood cells and FISH of chromosome abnormalities have been indicative of a clonal myeloid disorder.^981,982,983 Some cases may arise in the hematopoietic multipotential cell, others in a neutrophil progenitor cell (Chap. 85).^{981,982,983,984,985} Evidence points to defective apoptotic signals accounting, in part, for the striking accumulation of segmented neutrophils in the blood.⁹⁸⁶ The median age at onset is approximately age 65 years. Younger patients may be affected.⁹⁸⁷ As in most clonal myeloid diseases, men are affected more frequently than are women.

Clinical Features

Symptoms and Signs Patients may complain of weakness, anorexia, weight loss, abdominal pain, and easy bruising. Symptoms and signs of gouty arthritis occur in approximately one-third of cases. The spleen is enlarged in almost all cases, and the liver frequently is enlarged. Lymphadenopathy is very infrequent.⁹⁸⁵ A hemorrhagic tendency is present in some patients.

Laboratory Findings Although some patients have a normal hemoglobin concentration at the time of presentation, most have mild to moderate anemia on presentation. The reticulocyte count usually is between 0.5 and 3.0 percent. The platelet count rarely is less than 125 × 10⁹/L and usually is normal. Coagulation times are normal. The total leukocyte count usually is between 25 and 100 × 10⁹/L in most cases, and only rarely is less than 20 × 10⁹/L or exceeds 100 × 10⁹/L. Neutrophils compose 85 to 95 percent of the white cells. Although segmented cells usually dominate, occasional cases have a high proportion of band forms. Very infrequently, metamyelocytes, myelocytes, and nucleated red cells may be present in patients. Basophil and eosinophil counts are not increased. Blasts nearly always are absent from the blood. Neutrophil alkaline phosphatase activity is increased in almost all cases.

The marrow invariably shows granulocytic hyperplasia with myeloid-to-erythroid ratios as high as 10:1. Myeloblasts are not overtly increased in number (0.5 to 3.0 percent). Megakaryocytes are either normal or slightly increased in number and have normal distribution and morphology. Erythropoiesis usually is mildly decreased. Unlike CML, reticulin fibrosis is unusual. A few cases with dysmorphic features in the marrow (acquired Pelger-Hüet anomaly, erythroid, dysplasia, micromegakaryocytes) have been reported. Serum vitamin B₁₂-binding protein and vitamin B₁₂ levels both are markedly increased above normal. Serum uric acid concentration is increased, and serum LDH activity may be increased.

Almost every case examined postmortem had liver and splenic enlargement. Portal hepatic and splenic red pulp infiltrates of neutrophils or islands of extramedullary hematopoiesis with immature myeloid cells and megakaryocytes are characteristic.

Cytogenetic and Genetic Findings By definition, the Ph chromosome, BCR gene rearrangements, and BCR-ABL1 transcripts are absent.^{987,988,989,990} Most patients have normal karyotypes, but approximately 25 percent of patients have nonrandom abnormalities of chromosomes.⁹⁹⁰ Deletions of chromosome 20q and trisomy 21 or 9 are the most common abnormalities. The disease has been shown to be associated with a mutation in the colony-stimulating factor 3 receptor gene (CSF3R) alone (approximately 30 percent of cases), or a combination of mutated CSF3R and a SET binding protein gene (SETBP1) mutation (approximately 60 percent of cases) or the JAK2^V617F mutation alone (approximately 10 percent of cases).^991,992 Two principal mutations were observed in CSF3R: membrane proximal CSF3R^S783fs mutation and truncated CSF3R^T618I mutation. The former results in deregulation of the JAK family kinases and may respond to JAK inhibitors, whereas the later deregulates SRC family-TNK2 kinases and confers sensitivity to dasatinib.⁹⁹¹ Four of five prior reports of JAK2 mutations in a single patient with chronic neutrophilic leukemia were cited. In the fifth report, six patients with the disease were studied and one had a JAK2^V617F mutation.

Differential Diagnosis

Most leukemoid reactions are associated with an obvious underlying cause, such as pancreatitis, carcinoma, connective tissue disease, smoker’s neutrophilia, and chronic bacterial or fungal infection. The leukocyte alkaline phosphatase level usually is markedly elevated in chronic neutrophilic leukemia and markedly decreased in CML. More to the point, molecular studies identifying BCR gene rearrangement or the presence of BCR-ABL1 transcripts should distinguish chronic neutrophilic leukemia (BCR-ABL1–negative) from neutrophilic CML (BCR-ABL1–positive; see “Special Clinical Features” above). In the latter case, more than half of the patients have thrombocytosis and megakaryocytic hyperplasia, which are uncharacteristic of chronic neutrophilic leukemia. The presence of a CSF3R or JAK2 mutation with a classical clinical picture would be strong diagnostic evidence for the disease.

Treatment

No systematic studies of treatment have been reported. Although hydroxyurea, IFN-α, or cytarabine may decrease the white count and spleen size, long-term benefit is unusual.^{987,988,989,990} Intensive therapy has led to early posttreatment deaths. With identification of a specific genetic mutation, either a JAK2 inhibitor (e.g., ruxolitinib) (for the CSF3R^T618I mutation) or dasatinib (for the CSF3R^S783fs mutation) should be considered. The disease is rare and clinical trials would be difficult. In vitro studies of cell sensitivity and reports of individual responses may have to be relied upon.^991,992,993 Allogeneic stem cell transplantation in eligible patients may be curative.⁹⁹⁴

Course and Prognosis

The disease is fatal, with a median survival of approximately 2.5 years and a range of 0.5 to 6 years.^{987,988,989,990} A case of spontaneous remission has been reported. The prognosis is considerably worse than the prognosis for CML despite the prevalence of mature neutrophils and the paucity of blasts. Newer approaches with dasatinib or ruxolitinib for the appropriate genetic mutations (see “Treatment” above) may provide better outcomes. Causes of death have included (1) intracranial hemorrhage, sometimes in the presence of adequate platelet counts and coagulation times, suggesting a vascular infiltrative process; (2) severe infection; (3) transformation to AML; and (4) the toxic effects of intensive therapy. The disease usually afflicts older persons, and cardiac, pulmonary, and vascular diseases contribute to a fatal outcome.

A remarkable frequency of concordant essential monoclonal gammopathy or myeloma has been described.^{983,995,996,997,998,999,1000,1001,1002,1003} In two cases, the extreme neutrophilia proved to be a polyclonal response to a plasma cell disorder.^983,1004 Chronic neutrophilic leukemia has evolved from polycythemia vera or oligoblastic leukemia,^{1005,1006,1007,1008,1009} supporting its relationship to the clonal hemopathies.^{983,1010,1011}

BCR REARRANGEMENT-NEGATIVE PHENOTYPICALLY TYPICAL CHRONIC MYELOGENOUS LEUKEMIA

A very small proportion of patients (approximately 4 percent) with clinical manifestations within the limits usually applied to the diagnosis of CML have neither a Ph chromosome (classic, variant, or masked) nor evidence of rearrangement of BCR on chromosome 22. This circumstance represents BCR-negative CML. The literature describing Ph chromosome–negative CML prior to 1987 is difficult to evaluate because many cases were not studied carefully for masked or variant translocations and for the BCR gene rearrangement. Ph chromosome–negative CML is a clonal disease⁹⁰³ that has the propensity for lymphoid and myeloid transformation.^1012,1013 Although most cases of BCR rearrangement–negative CML are closer in manifestations to CMML,^{903,927,1014,1015,1016} some cases are indistinguishable from classic CML but without the BCR-ABL1 after exhaustive molecular diagnostic evaluation.^{1017,1018,1019,1020,1021} In a report of 76 such patients, the median age was 66 years (range: 24 to 88), splenomegaly was present in 50 percent of cases, the median white cell count was 38,000 cells/μL (38 × 10⁹ cells/L; range: 11 to 296), and the median hemoglobin was 11 g/dL (range: 7 to 16), with classical morphologic features in blood and marrow.¹⁰²¹ As the disease progressed, patients developed severe cytopenias.¹⁰¹⁸ Median survival was 24 months and only 7 percent survived for more than 5 years. Myeloid blast phase occurred in one-third of those followed until their death. Occasional patients had extended complete remissions with IFN-γ therapy.¹⁰²⁰ Hydroxyurea can be useful as palliative therapy.

Some patients have transposition of ABL1 to chromosome 22 but not the classic translocation. In such cases, including TEL-ABL1 translocations, transient responses to imatinib have been observed.¹⁰²²

Uncommon cases of coexisting BCR-negative and BCR-positive clones have been described, and the basis for such cases is in dispute.¹⁰²³ One proposed explanation is that this case is an example of “field carcinogenesis” in which multiple clones coexist.¹⁰²³ An alternative explanation is that these cases represent the dual progeny of a single unstable clone.¹⁰²⁴ The long-term survival of patients with CML may permit the emergence of a drug-induced or spontaneous second malignancy, and in the former it may be notably associated with imatinib (see “Secondary Chromosomal Changes with Tyrosine Kinase Inhibitors above).^{1025,1026,1027}

ATYPICAL MYELOPROLIFERATIVE DISEASE

Definition

There is a small but measurable frequency of patients who do not fit the diagnostic criteria of the other chronic myeloproliferative diseases, yet their findings represent the manifestations of a clonal myeloid neoplasm.^{1028,1029,1030,1031} A neoplasm of a hematopoietic stem cell has many variations in expression (Chap. 83), and it is surprising that nearly all can be pigeonholed into a generally agreed upon phenotype. These cases are among those that do not fit a standard diagnostic pattern. These cases have been classified by the WHO as atypical CML (aCML) or alternatively myelodysplastic-myeloproliferative syndrome, which category excludes BCR-rearrangement–positive CML, CMML, chronic neutrophilic leukemia, refractory sideroblastic anemia with thrombocytosis, and other classical syndromes, as judged phenotypically and genotypically. An attempt at teasing apart these syndromes into specific classifications proved how difficult that was, even when very arbitrary criteria were applied.¹⁰³¹ Until a genetic basis is more clearly defined and specific therapy becomes available, we prefer to use the term atypical myeloproliferative disease for this category of hematopoietic multipotential (stem) cell disorder because, when starting with a single patient, the clinical features do not allow clear distinctions among patients and therapy is based on the manifestations in a given case, the consensus between patient and physician about the desirability and likely utility of therapy, and the rate of progression of the disease in that patient.

Clinical Features These patients are principally in the 60 to 90 year-old age range, but atypical expression of a multipotential hematopoietic cell neoplasm can occur at any age. Hepatomegaly and (or) splenomegaly are present in a minority of patients. Anemia and nearly always granulocytosis (granulocytes, notably neutrophils, and granulocytic precursors), sometimes with neutrophilic dysmorphia (e.g., acquired Pelger-Huët neutrophils) are characteristic. Neutrophilic precursors represent less than 15 percent of blood cells. The blast cell count in blood and marrow is low, usually less than 10 percent. Monocytes are not increased and eosinophils or basophils are usually not increased but may be as high as 10 percent of total leukocytes.

Laboratory Features The LDH is often elevated. The marrow is hypercellular with variable evidence of dysmorphic granulopoiesis and dysmorphic megakaryocytopoiesis. Mild marrow reticular fibrosis may be evident. Clonal cytogenetic abnormalities may occur but do not include translocations characteristic of classical chronic myeloid neoplasms, such as a BCR-ABL1 or translocations involving PDGFR-α, PDGFR-β, or FGFR1 seen in chronic eosinophilic leukemia with or without mastocytosis. Common myeloid-related cytogenetic abnormalities may occur, such as trisomy 8, del(20q), and others.

Therapy This type of neoplasm has no specific treatment and is usually treated “symptomatically” with red cell or platelet transfusion and an agent to reduce the white cell count, if that is a problem (e.g., hydroxyurea, 5-azacytidine, low-dose cytarabine, numerous others).¹⁰³¹ Appropriate patients may be considered for allogeneic hematopoietic stem cell transplantation, if a suitable donor is available, but this approach is problematic in an older population because they would be transplanted during active disease and with a higher-risk of GVHD. Nonmyeloablative allogeneic stem cell transplantation has also been used.

Course and Prognosis This patient population has a median survival of approximately 18 months, but the upper range is approximately 5 years. The disease may undergo clonal evolution to AML.