paroxysmal nocturnal hemoglobinuria (PNH)


 

Summary of paroxysmal nocturnal hemoglobinuria (PNH)

In contrast to all other intrinsic abnormalities of the erythrocyte, paroxysmal nocturnal hemoglobinuria (PNH) is an acquired disorder. PNH arises as a consequence of somatic mutation, in one or more hematopoietic stem cells, of PIGA, a gene located on the X chromosome that is required for synthesis of the glycosyl phosphatidylinositol (GPI) moiety that anchors some proteins to cell surface. Consequently, all GPI-anchored proteins (GPI-APs) that are normally expressed are deficient on the mutant hematopoietic stem cells and their progeny. The complement-mediated intravascular hemolytic anemia and the resulting hemoglobinuria that are the clinical hallmarks of PNH are a consequence of deficiency of the GPI-anchored complement regulatory proteins, CD55 and CD59. Although PNH is a clonal disease, it is not a malignant disease, and the extent to which the mutant clones expand varies greatly among patients. Thus, the blood of patients with PNH is a mosaic of abnormal and phenotypically normal cells. The size of the mutant clone is an important determinant of the clinical manifestations of the disease that include thrombophilia and marrow failure in addition to hemolysis. The diagnosis of PNH is straightforward using flow cytometry to detect and quantify the percentage of blood erythrocytes and neutrophils that lack GPI-APs. The intravascular hemolysis of PNH can be controlled with eculizumab, a humanized monoclonal antibody that blocks formation of the cytolytic membrane attack complex of complement. Eculizumab, however, has no effect on the underlying disease process. The mutant clone can be eradicated and normal hematopoiesis restored by allogeneic hematopoietic stem cell transplant.

Acronyms and Abbreviations

Acronyms and abbreviations that appear in this chapter include: APC, alternative pathway of complement; DAF, decay accelerating factor; EtN, ethanolamine; GLcN, glucosamine; GPI, glycosyl phosphatidylinositol; GPI-APs, glycosyl phosphatidylinositol-anchored proteins; LDH, lactate dehydrogenase; MAC, membrane attack complex of complement; MDS, myelodysplastic syndrome; MIRL, membrane inhibitor of reactive lysis; PIGA, phosphatidylinositol glycan class A; PMN, polymorphonuclear cell; PNH, paroxysmal nocturnal hemoglobinuria; PNH-sc, subclinical PNH; RA, refractory anemia; RAEB, refractory anemia with excess of blasts; RAEB-t, refractory anemia with excess of blasts in transformation; RA-PNH+, RA with a population of PNH cells; RA-PNH–, RA without a population of PNH cells; RARS, refractory anemia with ringed sideroblast; RBCs, red blood cells; RCMD, refractory cytopenias with multilineage dysplasia; RCMD-RS, RCMD with ringed sideroblasts; WHO, World Health Organization.

Definition and Early History

Although commonly regarded as a type of hemolytic anemia, paroxysmal nocturnal hemoglobinuria (PNH) is actually a hematopoietic stem cell disorder. PNH arises as a result of nonmalignant 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, progeny of affected stem cells (erythrocytes, granulocytes, monocytes, platelets, and lymphocytes) are deficient in all glycosyl phosphatidylinositol-anchored proteins (GPI-APs) that are normally expressed on hematopoietic cells (and all GPI-APs that are normally expressed on hematopoietic cells are deficient on progeny of PIGA mutant stem 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.

Comprehensive, scholarly reviews of the history of PNH have been published.1–4 The first 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; see Chap. 53). 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’s test) that along with the sucrose lysis test (sugar water test) of Robert Hartmann and David Jenkins were used as the standard diagnostic tests for PNH until being supplanted in the mid-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 pupil and colleague S.M. Lewis first systematically characterized the relationship between PNH and marrow failure.

Epidemiology

The prevalence of PNH is not known with certainty. Prevalence estimates are primarily anecdotal and 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 are Characteristic of PNH” 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. 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” below) is likely in the order of <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 has been reported in all age groups, the peak incidence is in the third and fourth decades of life, similar to that of aplastic anemia. 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.

Cause of paroxysmal nocturnal hemoglobinuria (PNH)

Complement and PNH

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.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 (see Chap. 17 for 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 cytolytic 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 and ultimately generate the MAC (the MAC is the common cytolytic subunit of the classical and lectin pathways of complement as well as the APC [see Chap. 17]).

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)8–10 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 (Fig. 40–1, bottom panel). But why are PNH erythrocytes deficient in the two complement regulatory proteins?

The Molecular Pathogenesis and Genetic Basis of PNH

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 transferase12–16 that is an obligate constituent of a complex biochemical pathway required for synthesis of the glycosyl phosphatidylinositol (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) is an epiphenomenon, occurring because the two proteins that regulate complement on erythrocytes happen to be GPI-anchored.

Another remarkable feature of PNH is phenotypic mosaicism (see Fig. 40–3A) based on PIGA genotype26 (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 (~90%) 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 relevant 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 (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.27 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.