Chapter 40: Paroxysmal Nocturnal Hemoglobinuria

Although commonly regarded as a type of hemolytic anemia, paroxysmal nocturnal hemoglobinuria (PNH) is properly categorized as a hematopoietic stem cell disorder. PNH arises as a result of clonal expansion of one or several hematopoietic stem cells that have acquired a somatic mutation of the X-chromosome gene PIGA (phosphatidylinositol glycan class A). As a consequence of mutant PIGA, any progeny of affected stem cells (erythrocytes, granulocytes, monocytes, platelets, and lymphocytes) are deficient in all glycosylphosphatidylinositol-anchored proteins (GPI-APs) that are normally expressed on hematopoietic cells. The clinical manifestations of PNH are hemolytic anemia, thrombophilia, and marrow failure, but only the hemolytic anemia is unequivocally a consequence of somatic mutation of PIGA. It is not a malignant neoplasm in the classical sense of uncontrolled proliferation of cells, spread to tissues other than marrow, or spatial replacement of hematopoiesis. Its effects can be lethal and it can uncommonly undergo clonal evolution to acute myelogenous leukemia.

Comprehensive, scholarly reviews of the history of PNH have been published.^1,2,3,4 The first published clinical description of PNH is attributed to William Gull in 1866, but he failed to distinguish definitively PNH from paroxysmal cold hemoglobinuria. Paul Strübing, in 1882, clearly recognized PNH as a distinct entity and undertook prescient experiments designed to test his hypothesis that the nocturnal hemoglobinuria was a consequence of acidification of plasma that occurred when carbon dioxide and lactic acid accumulated because of slowing of respiration during sleep. In 1911, A.A. Hijmans van den Berg demonstrated that the hemolysis of PNH is caused by a defect in the red cell rather than by the presence of an abnormal plasma factor (as is the case with paroxysmal cold hemoglobinuria; Chap. 54). Thomas Hale Ham is credited with discovering, in the late 1930s, that complement mediates the hemolysis of PNH erythrocytes, although it was not until the alternative pathway of complement was identified and characterized in the mid-1950s by Louis Pillemer that the basis of Ham’s original observations became apparent. Ham developed the acidified serum lysis test (Ham test) that, along with the sucrose lysis test (sugar water test) of Robert Hartmann and David Jenkins, was used as the standard diagnostic test for PNH until being supplanted in the early 1990s by flow cytometry. Both Hartmann and William Crosby brought attention to the important role that thrombosis (particularly the Budd-Chiari syndrome) plays in the natural history of PNH, and John Dacie and his student and colleague S.M. Lewis were the first to systematically characterize the relationship between PNH and marrow failure.

The prevalence of PNH is not known with certainty. Prevalence estimates are influenced by bias in study design and results differ considerably, in large part, because of the heterogeneous nature of the disease. The blood of patients with PNH is a mosaic of normal and abnormal cells, and the extent of the mosaicism varies widely among patients (see “Phenotypic Mosaicism is Characteristic of Paroxysmal Nocturnal Hemoglobinuria” below). Patients with small PNH clones have few or no symptoms related to hemolysis. Thus an argument can be made that asymptomatic patients with small clones do not have clinically significant PNH and should be excluded from prevalence estimates. Others, however, may argue that any patient with flow cytometric evidence of a population of GPI-AP–deficient cells, regardless of clone size, has PNH and should be included in prevalence estimates. Well-designed, rigorous studies of prevalence that address the issue of disease heterogeneity are needed, but, by any definition, PNH is a rare disease. The prevalence of clinically significant PNH (i.e., classic PNH) plus patients with relatively large clones that arise in the setting of another marrow failure syndrome, (see “Clinical Features” and Table 40–2 below) is likely in the order of less than 1 case per 200,000 population, easily fulfilling criteria (<1 case per 50,000 population) for classification as an ultraorphan disease.⁵ There is a close association between PNH and aplastic anemia, and environmental factors, drugs, and toxins, that cause aplastic anemia, concordantly increase the risk of developing PNH. Although PNH is reported in all age groups, the peak incidence is in the third and fourth decades of life, similar to that of aplastic anemia (Chap. 35). PNH is an acquired disorder, and there is no known inherited risk for developing the disease. A number of cases have been reported in which only one of a pair of identical twins was affected.

COMPLEMENT AND PAROXYSMAL NOCTURNAL HEMOGLOBINURIA

The chronic intravascular hemolysis that is the hallmark clinical manifestation of PNH is mediated by the alternative pathway of complement (APC) (Fig. 40–1).⁶ The APC is a component of innate immunity (Chap. 20).⁷ This ancient system evolved to protect the host against invasion by pathogenic microorganisms. Unlike the classical pathway of complement that is part of the system of acquired immunity and requires antibody for initiation of activation, the APC is in a state of continuous activation, armed at all times to protect the host (Chap. 19 provides a detailed review of the complement system). The APC cascade can be divided into two functional components: the amplification C3 and C5 convertases and the membrane attack complex (MAC). The C3 and C5 convertases (Fig. 40–1, top panel) are enzymatic complexes that initiate and amplify the activity of the APC. Generation of C5b by enzymatic cleavage of C5 by the APC C5 convertase activates the terminal pathway of complement that results ultimately in assembly of the cytolytic MAC.

Because the APC is primed for attack at all times, elaborate mechanisms for self-recognition and for protection of the host against APC-mediated injury have evolved. Both fluid-phase and membrane-bound proteins are involved in these processes. Normal human erythrocytes are protected against APC-mediated cytolysis primarily by decay-accelerating factor (DAF, CD55)^8,9,10 and membrane inhibitor of reactive lysis (MIRL, CD59).¹¹ These proteins act at different steps in the complement cascade (see Fig. 40–1, top panel). CD55 regulates the formation and stability of the C3 and C5 convertases, whereas CD59 blocks the formation of the MAC. Deficiency of CD55 and CD59 on the erythrocytes of PNH is the pathophysiologic basis of the Coombs-negative, intravascular hemolysis that is the clinical hallmark of the disease (see Fig. 40–1, bottom panel). But why are PNH erythrocytes deficient in the two complement regulatory proteins?

THE MOLECULAR PATHOGENESIS AND GENETIC BASIS OF PAROXYSMAL NOCTURNAL HEMOGLOBINURIA

PNH is a consequence of clonal expansion of one or more hematopoietic stem cells with mutant PIGA(located on Xp22.1).¹² The protein product of PIGA is a glycosyl transferase^{12,13,14,15,16} that is an obligate constituent of a complex biochemical pathway required for synthesis of the glycosylphosphatidylinositol (GPI) moiety that anchors individual proteins belonging to diverse functional groups to the cell surface (Fig. 40–2). As a result of mutant PIGA, progeny of the affected stem cells are deficient in all GPI-APs. Although more than 20 GPI-APs are expressed by hematopoietic cells, it is deficiency on red blood cells (RBCs) of the two GPI-anchored complement regulatory proteins, CD55 and CD59, that underlies the hemolytic anemia of PNH.¹⁷ RBCs lacking CD55 and CD59 undergo spontaneous intravascular hemolysis as a consequence of unregulated activation of the APC (see Fig. 40–1, bottom panel). Thus, the hallmark clinical manifestation of PNH (intravascular hemolysis and the resultant hemoglobinuria) occurs because the two proteins that regulate complement on erythrocytes happen to be GPI-anchored.

Hypothetically, the PNH phenotype would result from inactivation of any of the more than 25 genes involved in synthesis of the GPI-anchor (see Fig. 40–2), but, with one exception,¹⁸ somatic mutation of no gene involved in GPI-AP synthesis other than PIGA has been reported in patients with PNH. This phenomenon is accounted for by the fact that, of the genes involved in the GPI-anchor synthesis pathway, only PIGA is located on the X-chromosome. Therefore, somatic mutation of only one allele is required for expression of the phenotype as males have one X-chromosome and, as a consequence of X-inactivation during embryogenesis, females have only one functional X-chromosome in somatic tissues (Chap. 10). On the other hand, mutation of two alleles would be required for inactivation of any of the autosomal genes involved in the GPI-anchor synthesis pathway.¹⁸

Cells with PIGA mutations do not appear to have a proliferative advantage in vitro or in hybrid animal models made with PIGA knockouts.¹⁹ They have been found to be relatively resistant to apoptosis in some studies,^20,21,22,23 but not in others.^24,25 Thus, the basis of clonal selection and clonal expansion of PIGA mutant stem cells in patients with PNH remains largely enigmatic²⁶ although a number of hypotheses have been proposed (reviewed in Ref. 17).

PHENOTYPIC MOSAICISM IS CHARACTERISTIC OF PAROXYSMAL NOCTURNAL HEMOGLOBINURIA

The blood of patients with PNH is a mosaic of normal and abnormal cells (Fig. 40–3). Although PNH is a clonal disease, the extent to which the PIGA-mutant clone expands varies widely among patients.¹⁷ As an example, in some cases, greater than 90 percent of the blood cells may be derived from the PIGA-mutant clone, whereas in others, less than 10 percent of the blood cells may be GPI-AP deficient. This unique feature (variability in extent of mosaicism) is clinically relevant because patients with relatively small PNH clones have minimal or no symptoms and require no PNH-specific treatment, whereas those with large clones are often debilitated by the consequences of chronic complement-mediated intravascular hemolysis and respond dramatically to complement inhibitory therapy.

Another remarkable feature of PNH is phenotypic mosaicism (see Fig. 40–3A) based on PIGA genotype²⁷ (see Fig. 40–3B) that determines the degree of GPI-AP deficiency.¹⁷ PNH III cells are completely deficient in GPI-APs, PNH II cells are partially (approximately 90 percent) deficient and PNH I cells express GPI-APs at normal density (putatively, these cells are progeny of residual normal stem cells; see Fig. 40–3A). Phenotype varies among patients (Fig. 40–4). Some patients have only type I and type III cells (the most common phenotype), some have type I, type II, and type III (the second most common phenotype), and some patients have only type I and type II cells (the least-common phenotype). Furthermore, the contribution of each phenotype to the composition of the blood varies. Phenotypic mosaicism is clinically important because PNH II cells are relatively resistant to spontaneous hemolysis, and patients with a high percentage of type II cells have a relatively benign clinical course with respect to hemolysis (Fig. 40–4).

The anemia of PNH is multifactorial as an element of marrow failure is present in all patients, although the degree of marrow dysfunction is variable.²⁸ In some patients, PNH arises in the setting of aplastic anemia. In this case, marrow failure is the dominant cause of anemia. In other patients with PNH, evidence of marrow dysfunction may be subtle (e.g., an inappropriately low reticulocyte count) with the degree of anemia being determined primarily by the rate of hemolysis that is, in turn, determined by PNH clone size.

The primary clinical manifestations of PNH are hemolysis, thrombosis, and marrow failure.²⁸ Constitutional symptoms (fatigue, lethargy, malaise, asthenia) dominate the history, with nocturnal hemoglobinuria being a presenting symptom in only approximately 25 percent of patients.²⁹ Direct questioning frequently elicits a history of episodic dysphagia and odynophagia, abdominal pain, and male impotence. Venous thrombosis, often occurring at unusual sites (Budd-Chiari syndrome, mesenteric, dermal, or cerebral veins), may complicate PNH. Arterial thrombosis is less common.

PNH should be suspected in all patients with nonspherocytic, Coombs-negative intravascular hemolysis (Table 40–1).

Although the clinical manifestations of PNH depend in large part on the size of the PIGA-mutant clone, the extent of the associate marrow failure also contributes significantly to disease manifestations. Thus, PNH is not a binary process and based on clinical features, marrow characteristics, and the size of the mutant clone as determined by the percentage of GPI-AP–deficient polymorphonuclear cells (PMNs), the International PNH Interest Group recognizes three disease subcategories (Table 40–2).²⁸

Reticulocytosis reflects the response to hemolysis, although the reticulocyte count may be lower than expected for the degree of anemia because of underlying marrow failure (see Table 40–1). Serum lactate dehydrogenase (LDH) concentration is always abnormally high in patients with clinically significant hemolysis and serves as an important surrogate marker for estimating and following the rate of intravascular hemolysis. A close association exists between PNH and aplastic anemia, and to a lesser extent between PNH and low-risk myelodysplastic syndromes (MDSs) (Chaps. 35 and 87 and see “Paroxysmal Nocturnal Hemoglobinuria and Marrow Failure” below). By using high-sensitivity flow cytometry, approximately 50 percent of patients with aplastic anemia and 15 percent of patients with low-risk MDS have been found to have a detectable population of GPI-AP–deficient erythrocytes and granulocytes.^30,31,32,33 In approximately 80 percent of these cases, the proportion of GPI-AP deficient cells is <1.0 percent of the total. These patients with very small populations of GPI-AP–deficient erythrocytes have no clinical or biochemical evidence of hemolysis and are designated as subclinical PNH (PNH-sc; see Table 40–2). Varying degrees of leukopenia, thrombocytopenia, and relative reticulocytopenia reflect the extent of marrow insufficiency (see “Paroxysmal Nocturnal Hemoglobinuria and Marrow Failure” below).

Once suspected, diagnosing PNH is straightforward as deficiency of GPI-APs on blood cells is readily demonstrated by flow cytometry (Fig. 40–5).^34,35 Although they have much biologic and historic importance, the acidified serum lysis test (Ham test) and the sucrose lysis test (sugar water test) have largely been abandoned as diagnostic assays because they are both less sensitive and less quantitative than flow cytometry. Flow cytometric analysis of both RBCs and PMNs is warranted, as clone size will be underestimated if only RBCs are examined because GPI-AP–deficient red cells are selectively destroyed by complement. Recent transfusion will also affect the estimate of clone size, if only RBCs are analyzed, but delineation of PNH phenotypes (i.e., the percentage of types I, II, and III cells) requires flow cytometric analysis of the erythrocyte population.

In addition to flow cytometric analysis, the basic initial evaluation of a patient with PNH should include complete blood count to assess the effects of the disease on production of leukocytes and platelets, as well as on erythrocytes (Table 40–3). In patients with classic PNH, the leukocyte and platelet counts are usually normal or nearly normal, whereas leukopenia, thrombocytopenia, or both invariably accompany PNH/aplastic anemia and PNH/MDS. The reticulocyte count is needed to assess the ongoing capacity of the marrow to respond to the anemia. Although the reticulocyte count is elevated in patients with classic PNH, as noted above, it may be inappropriately low for the degree of anemia, reflecting underlying relative insufficiency of hematopoiesis that is characteristic of the disease. The reticulocyte count is decreased in patients with PNH with concomitant aplastic anemia or low-risk MDS. Serum LDH is always markedly elevated in classic PNH. The degree of serum LDH elevation is variable in patients with PNH/aplastic anemia and PNH/MDS, depending on the size of the PNH clone (see Table 40–2). By definition, patients with PNH-sc have neither clinical nor biochemical evidence of hemolysis (see Table 40–2). Patients with classic PNH are often iron deficient as a result of chronic iron loss in the form of hemoglobinuria and hemosiderinuria (Chap. 43). Marrow aspirate and biopsy are needed to distinguish classic PNH from PNH in the setting of another marrow abnormality. Nonrandom cytogenetic abnormalities are rare in PNH.²⁶

PAROXYSMAL NOCTURNAL HEMOGLOBINURIA AND MARROW FAILURE

Although the marrow of patients with classic PNH appears relatively normal morphologically (see Table 40–2), numerous in vitro studies have shown that the growth characteristics of marrow-derived stem cells are aberrant.^21,36,37 Moreover, when stem cells are sorted into GPI-AP− and GPI-AP+ populations, compared to the GPI-AP+ population, the growth characteristics of the GPI-AP− population more closely approach those of normal control cells.^21,36 One plausible explanation for this observation is that the GPI-AP− cells are relatively protected from the pathophysiologic process that mediates the marrow injury, thereby providing a basis for natural selection of the PIGA mutant clone. In this view of PNH, outgrowth of the PIGA mutant clone is seen as an example of Darwinian evolution occurring within the microenvironment of the marrow. Although appealing, definitive experimental support for this hypothesis is lacking.

A close association exists between PNH and aplastic anemia and to a lesser extent between PNH and low-risk MDSs. By using high-resolution flow cytometry,³⁴ approximately 50 to 60 percent of patients with aplastic anemia and 15 to 20 percent of patients with low-risk MDS have been found to have a detectable population of GPI-AP–deficient erythrocytes and granulocytes.^{30,31,33,38,39} In approximately 90 percent of these cases, the proportion of GPI-AP–deficient blood neutrophils is less than 25 percent of the total.³⁸ Those patients with very small populations of GPI-AP–deficient erythrocytes (designated PNH-sc) have no clinical or biochemical evidence of hemolysis and require no specific treatment for PNH (see Table 40–2).

Studies have investigated the natural history of PNH clones in the setting of marrow failure.^38,40,41 The threshold that separates PNH-sc from clinical PNH is reached when the neutrophil clone size is in the range of 20 to 25 percent with a corresponding GPI-AP–deficient erythrocyte population of 3 to 5 percent.⁴⁰ Longitudinal studies indicate that clonal expansion occurs in 15 to 50 percent of cases.^38,40,41 In 10 to 25 percent of cases the clone disappears, and in 25 to 60 percent of cases the clone size persists unchanged.^38,40,41 Available evidence indicates that patients who present with PNH-sc do not progress to clinical PNH.^38,40,41 Among patients who present with clinical PNH in the setting of marrow failure, treatment for complications of PNH (eculizumab for hemolysis or anticoagulation for thrombosis) is required in approximately 50 percent of cases.⁴¹ There is no evidence that treatment with immunosuppressive therapy influences clonal expansion either positively or negatively.

The basis of the relationship between PNH and aplastic anemia is speculative. The vast majority of patients with PNH have some evidence of marrow failure (e.g., thrombocytopenia, leukopenia, or both) during the course of their disease.^26,42,43 Therefore, marrow injury may play a central role in the development of PNH by providing the conditions that favor the growth/survival of PIGA-mutant, GPI-AP–deficient stem cells. Finding a population of GPI-AP–deficient erythrocytes in patients with aplastic anemia is clinically relevant, as these patients have a particularly high probability of responding to immunosuppressive therapy, and the onset of the response appears to be more rapid compared to patients with aplastic anemia without a population of GPI-AP–deficient erythrocytes.^31,33,44

The presence of PNH cells has also been observed in patients with MDS.^30,33,39,45 Notably, the association between PNH and MDS appears to be confined to low-risk categories of MDS, particularly the refractory anemia (RA) variant.^30,33,39 Using high-sensitivity flow cytometry in which equal to or greater than 0.003 percent GPI-AP–deficient RBCs or PMNs was classified as abnormal, Wang and colleagues reported that 21 of 119 (18 percent) patients with RA MDS had a population of PNH cells, whereas GPI-AP–deficient cells were not detected in patients with RA with ringed sideroblast (RARS), RA with excess of blasts (RAEB), or RA with excess of blasts in transformation (RAEB-t). Compared to patients with RA without a population of PNH cells (RA-PNH−), patients with RA with a population of PNH cells (RA-PNH+) had a distinct clinical profile characterized by the following features: (1) less-pronounced morphologic abnormalities of blood cells; (2) more severe thrombocytopenia; (3) lower rates of karyotypic abnormalities; (4) higher incidence of HLA-DR15; (5) lower rate of progression to acute leukemia; and (6) higher probability of response to cyclosporine therapy.

That a population of PNH cells is associated only with low-risk MDS was confirmed in a North American study of 137 patients classified by World Health Organization (WHO) criteria.^39,46 The study found a population of PNH cells in 1 of 5 (20 percent) patients with 5q− syndrome, in 6 of 17 (35 percent) patients with RA, and in 2 of 37 (5 percent) patients with refractory cytopenias with multilineage dysplasia (RCMD), whereas no patient with RARS (0 of 9), RCMD-ringed sideroblasts (0 of 6), RAEB (0 of 26), MDS unspecified (0 of 10), myelodysplastic/myeloproliferative disease (0 of 10), primary myelofibrosis (0 of 5), chronic myelomonocytic leukemia (0 of 5), or acute myeloid leukemia (0 of 6) had a detectable population of GPI-AP–deficient blood cells.³⁹

When combined with evidence of polyclonal hematopoiesis (based on the pattern of X-chromosome inactivation in female patients), the presence of a population of PNH cells in patients with MDS predicts a relatively benign clinical course and a high probability of response to immunosuppressive therapy in patients.³⁰ A relatively good response to immunosuppressive therapy for patients with MDS and aplastic anemia was also predicted by expression of HLA-DR15 in studies of both North American and Japanese patients.^47,48 Together, these observations provide compelling indirect evidence that aplastic anemia and a subgroup of low-risk MDS are immune-mediated diseases and that the immune pathophysiologic process provides the selection pressure that favors the outgrowth of PIGA mutant, GPI-AP–deficient stem cells.

ECULIZUMAB

The complement-mediated intravascular hemolysis of PNH can be inhibited by blocking formation of the terminal complement pathway generated MAC, the cytolytic component of the complement system (see Fig. 40–1). The MAC consists of complement components C5b, C6, C7, C8, and multiple molecules of C9. Eculizumab (Soliris) is a humanized monoclonal antibody that binds to complement C5, preventing its activation to C5b and thereby inhibiting MAC formation (see Fig. 40–1).⁴⁹ In 2007, eculizumab was approved by both the FDA and the European Union Commission (now the European Medicines Agency) for treatment of the hemolysis of PNH. Treatment with eculizumab reduces transfusion requirements, ameliorates the anemia of PNH, and markedly improves quality of life by resolving the debilitating constitutional symptoms (fatigue, lethargy, asthenia) associated with chronic complement-mediated intravascular hemolysis.⁵⁰ Following treatment, serum LDH concentration returns to normal, but mild to moderate anemia and reticulocytosis usually persist, likely the result of ongoing extravascular hemolysis mediated by opsonization of PNH erythrocytes by activated complement C3, as eculizumab does not block the activity of the APC C3 convertase (see Fig. 40–1).^51,52 In some cases, extravascular hemolysis is sufficiently severe so as to require therapy.⁵³

Thromboembolic events are the major cause of morbidity and mortality in PNH,²⁸ and eculizumab appears to ameliorate the thrombophilia of PNH, although the studies that support that conclusion had suboptimal design.⁵⁴

Eculizumab is given by intravenous infusion on a biweekly schedule following an initial loading period consisting of five weekly treatments. In general, the drug is well tolerated; however, patients with congenital deficiency of complement C5 have an increased risk of infection with Neisseria species. For this reason, patients treated with eculizumab (which blocks the function of C5; see Fig. 40–1) are at risk for meningococcal septicemia. All patients must be inoculated with a meningococcal vaccine 2 weeks before starting therapy, but the vaccine is not 100 percent protective. Whether prophylactic antibiotic therapy aimed at preventing meningococcal infection is justified for a patient receiving eculizumab remains to be determined. Despite the fact that the percentage of GPI-AP–deficient erythrocytes increases during treatment with eculizumab,⁵⁵ there have been no reports of catastrophic hemolytic crises in the relatively few PNH patients who have discontinued treatment with eculizumab.^54,56

Eculizumab is expensive (approximately $400,000/year in the United States), and it has no effect on either the underlying stem cell abnormality or the associated marrow failure. Consequently, treatment must continue indefinitely and leukopenia, thrombocytopenia, and reticulocytopenia, if present, persist.

OTHER TREATMENT FOR PAROXYSMAL NOCTURNAL HEMOGLOBINURIA

Other than eculizumab, there is no specific treatment for PNH, and for patients who are not being treated with eculizumab, management is largely supportive (reviewed in Ref. 28). Although hemolysis is ameliorated in some patients by treatment with glucocorticoids or androgens, the use of steroids in the management of patients with PNH is controversial.²⁸ The main value of glucocorticoids may be in attenuating acute hemolytic exacerbations. Under these circumstances, brief pulses of prednisone may reduce the severity and duration of the crisis. The value of glucocorticoids in treating chronic hemolysis is limited by toxicity, and the harm that can accrue from long-term use cannot be overemphasized. An every-other-day schedule may attenuate some of the adverse effects of chronic glucocorticoid use,⁵⁷ but patients may note worsening of symptoms on the off day.

Androgen therapy, either alone or in combination with glucocorticoids, has been used successfully to treat the anemia of PNH.^57,58 As with glucocorticoids, the mechanism by which androgenic steroids ameliorate the anemia of PNH is not fully understood, although the rapid onset of action is consistent with complement inhibition.⁵⁸ Potential complications of androgen therapy include liver toxicity, prostatic hypertrophy, and virilizing effects. The toxicity profile is more favorable for attenuated synthetic androgens such as danazol, making long-term use of this drug a reasonable management option in responding patients. A starting dose of 400 mg twice a day is recommended, but a lower dose (100 to 400 mg/day) may be adequate to control chronic hemolysis.²⁸

Patients with PNH frequently become iron deficient as a result of both hemoglobinuria and hemosiderinuria.^57,58 Clinically important iron loss from hemosiderinuria can occur (Chap. 43), even in the absence of gross hemoglobinuria. Replacement is often associated with exacerbation of hemolysis, regardless of the route of administration.^57,58 Compared with parenteral replacement, oral administration of iron may be accompanied by less-severe hemolytic exacerbations, but urinary iron loss may be so great that repletion may not be achieved.⁵⁷ Concern for inducing a hemolytic exacerbation should not deter iron repletion.⁵⁷ If a hemolytic exacerbation occurs in the setting of iron repletion, the episode can be controlled by treatment with glucocorticoids or androgens or by suppression of erythropoiesis by transfusion. There is no concern about iron-replacement therapy inducing a hemolytic exacerbation in patients being treated with eculizumab as hemolysis is inhibited by the drug.

Because the hemolysis is a consequence of a defect intrinsic to patient’s erythrocytes, the anemia of PNH responds to red cell transfusion. Concerns about inducing a hemolytic exacerbation as a consequence of infusion of small amounts of donor plasma that may be included in red cell preparations appear unwarranted.⁵⁹ However, hemofiltration is recommended to prevent transfusion reaction arising from the interaction between donor leukocytes and recipient antibodies. Iatrogenic hemochromatosis from chronic transfusion may be delayed in patients with PNH as a result of iron loss from hemoglobinuria/hemosiderinuria,⁵⁷ but iron overload remains a concern in patients who require chronic transfusion when the anemia is primarily a consequence of marrow failure rather than intravascular hemolysis.

Supplemental folate (1 mg/day) is recommended to compensate for increased use (Chap. 41) associated with heightened erythropoiesis that is a consequence of ongoing hemolysis.²⁸

The role of splenectomy in the management of patients with PNH has not been investigated systematically. Reports of amelioration of hemolysis and improvement in cytopenias following splenectomy are anecdotal. Concerns about lack of proven efficacy and the potential for postoperative complications, particularly thrombosis, have led some to argue that splenectomy has no role in the management of PNH.²⁸

ALLOGENEIC HEMATOPOIETIC STEM CELL TRANSPLANTATION

Prior to the availability of eculizumab, the primary indications for transplantation were marrow failure, recurrent, life-threatening thrombosis, and uncontrollable hemolysis (Table 40–4).²⁸ The latter process can be eliminated by treatment with eculizumab and the thrombophilia of PNH may also respond to inhibition of intravascular hemolysis by eculizumab.^54,60 Nonetheless, transplantation is the only curative therapy for PNH, and the availability of molecularly defined, matched, unrelated donors, less-toxic conditioning regimens, reduction in transplantation-related morbidity and mortality, and improvements in posttransplantation supportive care make this option a viable alternative to medical management. Studies (see “Course and Prognosis” below) indicate a normal survival for patients with PNH treated with eculizumab, making the decision of whether to recommend medical management or hematopoietic stem cell transplant particularly complex.⁶⁰ An understanding of the unique pathobiology of PNH and the input of physicians experienced in transplantation and medical management of PNH are essential to develop an appropriate management plan for transplantation-eligible patients.⁶¹

For patients who are receiving transplantation for marrow failure, the focus of management is on the etiology of the marrow failure (see Table 40–4). For patients with aplastic anemia and a small PNH clone who undergo matched sibling donor allotransplantation, the conditioning regimen of antithymocyte globulin and cyclophosphamide coupled with graft-versus-host effects appear sufficient to eradicate the PNH clone.²⁸ However, in the unusual situation in which the patient has a syngeneic twin, a more intense conditioning regimen is required, as graft-versus-PNH effect does not contribute to clonal eradication in this circumstance.⁶² In the event that a patient with low-risk MDS with a PNH clone requires allotransplantation, the conditioning regimen (marrow ablative or reduced intensity) in combination with graft-versus-tumor effects usually is sufficient to eradicate the PNH clone.

Transplantation for classic PNH is aimed at eradicating the PNH clone, and both marrow ablative^63,64,65 and reduced-intensity^66,67 conditioning regimens are effective, although experience with the latter is more limited. Successful outcomes have been reported using matched, unrelated donors, as well as matched, sibling donors.^66,68 There are no PNH-specific adverse events associated with transplantation; severe, acute graft-versus-host disease (GVHD) occurs in more than one-third of the patients and the incidence of chronic GVHD is approximately 35 percent. Overall survival for unselected PNH patients who undergo transplantation using an human leukocyte antigen (HLA)-matched sibling donor is in the range of 50 to 60 percent.²⁸

MANAGEMENT OF THE THROMBOPHILIA OF PAROXYSMAL NOCTURNAL HEMOGLOBINURIA

Thromboembolic complications are the leading cause of morbidity and mortality in PNH.⁶⁹ Prophylaxis against thromboembolic events in patients with PNH is an issue of active debate.⁶⁹ Current estimates of risk are based on retrospective analysis,^{54,70,71,72,73} but risk appears to correlate with size of the PNH clone (based on flow cytometric determination of the percentage of GPI-AP–deficient PMNs), leading to the recommendation that patients with greater than 50 to 60 percent GPI-AP–deficient PMNs be offered prophylactic anticoagulation.^70,71 Treatment with warfarin with a goal international normalized ratio (INR) of between 2.0 and 3.0 is recommended for patients with PNH who require chronic anticoagulation either for treatment of a thromboembolic event or for prophylaxis. There are no empiric data to guide the use of low-molecular-weight heparin or novel oral anticoagulants in these settings, but their use can be considered in patients with adequate renal function who fail warfarin or in patients have difficulty maintaining a consistent therapeutic INR.

Although arterial thrombosis may be observed,⁵⁴ thromboembolic events in patients with PNH usually involve the venous system. Acute thrombotic events require anticoagulation with heparin. Systemic thrombolytic therapy,^74,75 or thrombolytic therapy delivered via canalization directly to the affected site,⁷⁶ should be strongly considered in patients with acute onset of Budd-Chiari syndrome.

Thrombocytopenia often complicates PNH, and this issue should be addressed when formulating an anticoagulation management plan. Thrombocytopenia is a relative contraindication to anticoagulation, and transfusions should be given to maintain the platelet count in a safe range rather than withholding therapy.⁷⁷ Patients with PNH who experience a thromboembolic event should be anticoagulated indefinitely. Recurrent, life-threatening thrombosis merits consideration of marrow transplantation, but such patients are at high-risk for transplantation-related adverse events (see Table 40–4).⁶¹

Eculizumab reduces the risk of thromboembolic complications.⁵⁴ For patients being treated with eculizumab who have no prior history of thromboembolic complications, prophylactic anticoagulation may not be necessary.

PREGNANCY AND PAROXYSMAL NOCTURNAL HEMOGLOBINURIA

Women with PNH can have serious morbidity and increased mortality during pregnancy.^77,78 Because of concerns about fetal/maternal risks from exposure to potentially toxic therapy, anticoagulation and transfusion have been the mainstay of management. Eculizumab has been assigned to pregnancy category C (risk cannot be ruled out) by the FDA. There are no controlled studies of the use of eculizumab in human pregnancy; however, anecdotal reports and small series have identified no significant adverse effects when eculizumab is used during pregnancy, including early in gestation and, in one case, from the time of conception.^51,79,80 Nonetheless, until more is known about the safety of eculizumab in pregnancy, it seems prudent to restricted use of the drug to the third trimester and then only for patients who are at high-risk for thrombosis and who have no acceptable therapeutic alternatives.

Moderate to severe thrombocytopenia may complicate the pregnancy, and clinically significant bleeding in this setting necessitates platelet transfusion. The incidence of clinically apparent venous thromboembolism during pregnancy in women with PNH is approximately 10 percent,⁷⁷ and these events are associated with a high risk of mortality.^77,78 Similar to nonpregnant patients with PNH, cerebral and hepatic veins are commonly involved sites of thrombosis. Thrombolytic therapy should be considered for those with Budd-Chiari syndrome.

The role of prophylactic anticoagulation for pregnant women with PNH has not been studied prospectively; however, because of the significant morbidity and mortality associated with thromboembolism in this setting, prophylaxis is recommended. Coumadin is contraindicated because of teratogenic potential in the first trimester and hemorrhagic risks later in gestation (Chap. 8). Anticoagulation with heparin should begin immediately once the pregnancy is documented. Low-molecular-weight heparin has a hypothetical advantage over unfractionated heparin because of a lower incidence of drug-induced thrombocytopenia and less osteopenia (Chap. 118). Careful monitoring of the platelet count is required because thrombocytopenia may worsen during the period of anticoagulation. Anticoagulation can be discontinued briefly around the time of delivery. However, it should be restarted as soon as is feasible and continued for at least 6 weeks into the postpartum period, as thrombosis during the puerperium is a major concern.^77,78 Most deliveries can be accomplished vaginally, although premature delivery may be necessary. Despite the many concerns surrounding PNH and pregnancy, successful outcomes appear to be the rule rather than the exception^72,77; however, management is complicated and should involve the combined efforts of a knowledgeable hematologist and an obstetrician experienced in dealing with high-risk pregnancies.²⁷

PEDIATRIC PAROXYSMAL NOCTURNAL HEMOGLOBINURIA

PNH can occur in the young (approximately 10 percent of patients are younger than age 21 years at the time of diagnosis).²⁸ A retrospective analysis of 26 cases, underscored the many similarities between childhood and adult PNH.⁸¹ Signs and symptoms of hemolysis, marrow failure, and thrombosis dominate the clinical picture, although gross hemoglobinuria as a presenting symptom may be less common in young patients. A generally good response to immunosuppressive therapy was observed,⁸¹ but based on poor long-term survival, hematopoietic cell transplantation is the recommended treatment for childhood PNH. One study⁸² confirmed the common presentation of marrow failure in 11 children with PNH, and reported that five patients eventually underwent hematopoietic cell transplant (three matched unrelated donors and two matched family donors), of whom four were long-term survivors. In another study of 12 young patients over an 18-year period, 10 presented with evidence of marrow failure and only one with hemoglobinuria.⁴⁰ There were six children with thrombosis and five with myelodysplastic features, indicating that the clinical presentation may be more similar to adult PNH than previously recognized. The safety and effectiveness of eculizumab in pediatric patients below the age of 18 years has not been established, however, there are anecdotal reports of its use in pediatric patients with PNH.⁸³ Although eculizumab is not approved for PNH patients younger than age 18 years, approval will likely be sought once pharmacodynamic and pharmacokinetic characteristics of the drug are defined for the pediatric/adolescent population.⁸⁴ The availability of eculizumab for pediatric PNH may be particularly advantageous as a bridge prior to implementation of more definitive therapy.

The clinical course of PNH is enormously variable. In rare instances, the patient may succumb to this disease within a few months of the first onset of symptoms. Most patients experience a chronic course in which the severity of the disease waxes and wanes as the normal cells and the PNH clone alternately appear to gain ascendancy. Rarely, the abnormal clone disappears altogether, and the patient appears to be cured. Transformation to acute leukemia is uncommon (in the range of 1 percent). In some instances, but not in others, leukemic blasts are GPI-AP–deficient.⁴⁶

As with so many other diseases, initial reports on PNH tended to emphasize the more severely affected patients, so the prognosis was generally deemed to be very grave. As physicians developed a higher index of suspicion concerning this disorder, and as simplified methods for diagnosis became available, milder cases were diagnosed with a better long-term outlook. Nonetheless, even today, the disease must be considered a very serious one. The most common lethal event is a thrombotic episode such as the Budd-Chiari syndrome,^69,73,85 but the various complications of pancytopenia also may lead to death,^29,43 and in a few patients, the terminal process is development of acute leukemia.⁴⁶ In a study of 220 patients with PNH followed for up to 46 years in the preeculizumab era, the Kaplan-Meier survival estimate was 65 percent at 10 years and 48 percent at 15 years after diagnosis.⁸⁵ In another preeculizumab era study of 80 consecutive patients the outlook was similar: the median survival after diagnosis was 10 years, with 28 percent of patients surviving for 25 years.⁴³ Eight-year cumulative incidence rates of the main complications of pancytopenia, thrombosis, and MDS were 15 percent, 28 percent, and 5 percent, respectively. Poor survival was associated with age older than 55 years at the time of diagnosis, the occurrence of thrombosis as a complication, evolution to pancytopenia, MDS or acute leukemia, and thrombocytopenia at diagnosis. The prognosis of patients in whom aplastic anemia antedated PNH was better than in those in whom it did not.⁸⁵

In addition to symptomatic benefit, treatment with eculizumab appears to influence the natural history of PNH. In a retrospective study of the clinical history of 79 patients with classic PNH or PNH/marrow failure treated with eculizumab, the median age at diagnosis was 37 years (range: 12 to 79 years) and the median age at the time of initiation of treatment with eculizumab was 46 years (range: 14 to 84 years).⁶⁰ The mean duration of treatment with eculizumab was 39 months (range: 1 to 98 months). Based on flow cytometric analysis of blood neutrophils, the average clone size among the treated patients was 96.4 percent (range: 41.8 to 100 percent). Twenty-four patients (30 percent) had a history of a marrow failure syndrome at the time of diagnosis (23 with aplastic anemia and one with MDS). Thrombotic episodes were reported in 27 percent of patients prior to starting eculizumab (including 12 cases of Budd-Chiari syndrome, four cases of mesenteric vein thrombosis, three cases of cerebral vein thrombosis). The investigators found that treatment with eculizumab reduced the mean yearly transfusion requirement from 19.3 units to 5.0 units. Of 61 patients who had been on eculizumab for more than 1 year, 40 (66 percent) became transfusion-independent. Thrombosis was observed in two patients while on eculizumab. No thrombotic events were reported in 21 patients who discontinue prophylactic anticoagulation after starting treatment with eculizumab.

Survival of the 79 patients treated with eculizumab was the same as that of a group of age- and sex-matched controls from the general population. Two eculizumab patients developed documented meningococcal infection with Neisseria meningitidis serogroup B, and thereafter, a program of antibiotic prophylaxis was instituted by the investigators.

A multicenter study involving 195 patients followed for 66 months largely confirmed these findings.⁸⁶

Together, these results demonstrate that treatment with eculizumab alters the natural history of PNH both by reducing or eliminating transfusion requirements through inhibition of intravascular hemolysis and by virtually eradicating thromboembolic complications. Eculizumab treatment may also reduce disease related mortality although the extent to which the drug enhances survival cannot be accurately determined from these studies as the experimental design did not include a randomized control group of patients. Eculizumab does not appear to affect either the marrow failure component of the disease or the clonal hematopoiesis that underlies disease pathophysiology. Although PNH is a clonal disease, it is not a malignant disease, as previously discussed, and it is this characteristic of PNH that allows for successful long-term symptomatic management in the absence of a treatment strategy aimed at eradicating the PIGA mutant hematopoietic stem cells.