Endogenous erythroid and megakaryocytic circulating progenitors, HUMARA clonality assay, and PRV-1 expression are useful tools for diagnosis of polycythemia vera and essential thrombocythemia

Recently, Liu et al¹  and Kralovics et al²  reported in this journal the possible diagnostic value of some biologic markers in chronic myeloproliferative disorders such as polycythemia vera (PV) and essential thrombocythemia (ET). Briefly, they described a strong correlation between endogenous erythroid colony growth in the absence of added erythropoietin (Epo) and polycythemia rubra vera 1 (PRV-1) expression in PV patients.

Our experience demonstrates very similar results using comparable methodologies. We studied the correlations among endogenous growth of circulating erythroid burst-forming unit (eBFU-E) and megakaryocytic colony-forming unit (eCFU-MK) progenitors, clonality by human androgen receptor–polymerase chain reaction (HUMARA-PCR) assay, and expression of PRV-1 in females with PV (n = 11) and ET (n = 17). All patients fulfilled the diagnostic criteria of the Polycythemia Vera Study Group (PVSG). At the time HUMARA and PRV-1 assays were performed, 5 of 17 ET and 2 of 11 PV patients were receiving myelosupressive/platelet-lowering agents. BFU-E and CFU-MK were determined at diagnosis, except for 2 ET patients who were receiving anagrelide.

In vitro cultures were performed as previously reported.³  HUMARA assay was analyzed on granulocytes and CD3⁺ lymphocytes. PRV-1 expression was quantified by real-time reverse-transcriptase (RT)–PCR in RNA from granulocytes.

In PV patients, 11 (100%) of 11 showed eBFU-E and 4 (36%) of 11, eCFU-MK. Clonality was demonstrated in 5 (50%) of 10 patients, and 1 patient was homozygous. PRV-1 overexpression was detected in 10 (91%) of 11 patients. The correlation among the 3 assays gave a concurrent positive result in 5 patients, but considering the 3 assays separately, we found a positive result in all PV patients (Table 1).

In ET patients, 5 (29%) of 17 had eBFU-E, 7 (41%) of 17 had eCFU-MK, and 9 (53%) of 17 had eBFU-E and/or eCFU-MK. Clonality was demonstrated in 6 (40%) of 15 patients, and the other 2 were homozygous. PRV-1 overexpression was observed in 10 (59%) of 17 patients (Table 1). Correlation between eBFU-E and/or eCFU-MK and overexpression of PRV-1 were observed in 7 (41%) of 17 patients. When we compared the 3 assays, a coinciding positive result was observed in 2 patients, but if they were considered separately, a positive result was found in 15 (88%) of 17 ET patients.

In our preliminary results, the best and most reliable correlation was observed between eBFU-E and PRV-1 expression in PV (91% of patients). In ET, a lower correlation (41%) between the 2 assays was found; but if we consider all assays separately, patients with a positive result increase to 88%. Cultures and PRV-1 results are in agreement with Liu et al¹  and Kralovics et al²  except for eCFU-MK, which was not studied. Clonality by HUMARA gene is lower than that obtained by Liu et al¹  using G6PD, IDS, MPP1, BTK, and FHL1 genes.

We conclude that endogenous BFU-E remains the single most useful technique for diagnosis of PV and, together with PRV-1 expression, the most reliable biologic markers for PV. In ET, eBFU-E and eCFU-MK with PRV-1 expression are useful diagnostic tests in more than 70% of ET patients. We suggest, in line with others,^1,2  that the concomitant implementation of in vitro cultures of BFU-E and/or CFU-MK and PRV-1 expression should be applied in PV and ET for their high diagnostic yield and that, whenever possible, HUMARA assay should be added.