Symptoms of paroxysmal nocturnal hemoglobinuria (PNH)

Clinical Features paroxysmal nocturnal hemoglobinuria (PNH)

The primary clinical manifestations of PNH are hemolysis, thrombosis, and marrow failure.27 Constitutional symptoms (fatigue, lethargy, malaise, asthenia) dominate the history, but nocturnal hemoglobinuria is a presenting symptom in only approximately 25 percent of patients.28 Directed 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.

Laboratory Features

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

Table 40–1. Recommendation for Screening Patients for PNH*
History of episodic hemoglobinuria
Evidence of nonspherocytic, Coombs negative intravascular hemolysis (must have abnormally high serum lactate dehydrogenase)
Patients with aplastic anemia (screen at diagnosis and once yearly even in the absence of intravascular hemolysis)
Patients with refractory anemia (RA) or refractory cytopenias with multilineage dysplasia (RCMD) variants of myelodysplastic syndrome (MDS)
Patients with venous thrombosis involving unusual sites (usually have evidence of intravascular hemolysis)
Budd-Chiari syndrome
Other intraabdominal sites
Cerebral veins
Dermal veins
*Screening by flow cytometric analysis of GPI-APs on RBCs and polymorphonuclear cells.There is no indication for screening patients with other MDS classifications.

While the clinical manifestations of PNH depend in large part on the size of the PIGA mutant clone, the extent of the associated 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).27

Table 40–2. Classification of PNH*
Category Rate of Intravascular Hemolysis Marrow Flow Cytometry Benefit from Eculizumab
Classic Florid (macroscopic hemoglobinuria is frequent or persistent) Cellular marrow with erythroid hyperplasia and normal or near-normal morphology‡ Large population (>50%) of GPI-AP deficient PMNs Yes
PNH in the setting of another marrow failure syndrome§ Mild to moderate (macroscopic hemoglobinuria is intermittent or absent) Evidence of a concomitant marrow failure syndrome§ Although variable, the percentage of GPI-AP deficient PMNs is usually relatively small (<30%) Dependent on the size of the PNH clone
Subclinical No clinical or biochemical evidence of intravascular hemolysis Evidence of a concomitant marrow failure syndrome§ Small (<1%) population of GPI-AP deficient PMNs detected by high-resolution flow cytometry No
*Based on recommendations of the International PNH Interest Group (Blood 106:3699, 2005).Based on macroscopic hemoglobinuria, serum lactate dehydrogenase concentration, and reticulocyte count.

‡Karyotypic abnormalities are uncommon.

§Aplastic anemia & refractory anemia/MDS are the most commonly associated marrow failure syndromes

Analysis of PMNs is more informative than analysis of RBCs because of selective destruction GPI-AP–deficient RBCs.

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 determining 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 (see Chaps. 34 and 88 and see “PNH and Marrow Failure” below). By using high-sensitivity flow cytometry, approximately 60 percent of patients with aplastic anemia and 20 percent of patients with low-risk myelodysplastic syndrome (MDS) have been found to have a detectable population of GPI-AP–deficient erythrocytes and granulocytes.29–31 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.

Once suspected, diagnosing PNH is straightforward as deficiency of GPI-APs on blood cells is readily demonstrated by flow cytometry32 (Fig. 40–5). Although they have much biologic and historic importance, the acidified serum lysis test (Ham test) and the sucrose lysis test (sugar water test) have been largely 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

Differential Diagnosis

PNH and Marrow Failure

Although the marrow of patients with classic PNH appears fairly normal morphologically (see Table 40–2), numerous in vitro studies have shown that the growth characteristics of marrow-derived stem cells are aberrant.22,33,34 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.22,33 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 intellectually appealing, rigorous experimental support for this hypothesis is lacking.

Using high-resolution flow cytometry with the capacity to detect <0.003 percent GPI-AP–deficient erythrocytes and granulocytes, 50 to 60 percent of patients with aplastic anemia can be shown to have a population of PNH cells at diagnosis,30,35 however, only 10 to 15 percent of patients with aplastic anemia treated with immunosuppressive therapy subsequently develop clinically apparent PNH,36 and development of clinical disease, when it occurs, typically follows diagnosis of aplastic anemia by several years. In the remainder, GPI-AP– cells persist subclinically or disappear, suggesting that mutant PIGA (and the consequent deficiency of GPI-APs) is necessary for clonal selection but is insufficient to account for the clonal expansion required for clinical manifestations of PNH to become apparent. One interpretation of these observations is that factors in addition to mutant PIGA determine the clinical phenotype of the disease by affecting the extent to which the PIGA mutant stem cells expand. Conceivably, a second genetic event that works additively or synergistically with mutant PIGA is required for clonal expansion.25 That the extent of clonal expansion varies markedly among patients, however, suggests that the second event may have diverse etiologies and could involve somatic mutations, epigenetic phenomenon or stochastic processes.

The basis of the relationship between PNH and aplastic anemia is speculative. Most patients with PNH have some evidence of marrow failure (e.g., thrombocytopenia, leukopenia, or both) during the course of their disease.37–40 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.30,31

The presence of PNH cells has also been observed in patients with MDS.29,31,41,42 Of note, the association between PNH and MDS appears to be confined to low-risk categories of MDS, particularly the refractory anemia (RA) variant.29,31,42 Using high-sensitivity flow cytometry in which 0.003 percent GPI-AP–deficient RBCs or PMNs was classified as abnormal, Wang and colleagues reported that 21 of 119 (18%) patients with RA MDS had a population of PNH cells, whereas GPI-AP–deficient cells were not detected in patients with refractory anemia with ringed sideroblast (RARS), refractory anemia with excess of blasts (RAEB), or refractory anemia 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; (6) higher probability of response to cyclosporine therapy.

That a population of PNH cells is associated only with low-risk MDS variants in Japanese patients was confirmed in a North American study of 137 patients42 classified by World Health Organization (WHO) criteria.43 The study found a population of PNH cells in 1 of 5 (20%) patients with 5q– syndrome, in 6 of 17 (35%) patients with RA, and in 2 of 37 (5%) 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.

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.29 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.44,45 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.