High Frequency of HPA-1B in Leukemia Patients.

Background: The HPA-1 alloantigen is a biallelic co-dominant system on human platelets carried on glycoprotein IIIa. Alloimmunization against the HPA-1 has clinical relevance in multiple transfused patients, in neonates with alloimmune thrombocytopenia and in platelet refractoriness. Accurate HPA genotyping is important to facilitate the provision of HPA-matched blood components for these patients. The purpose of this study was to determine the HPA-1 gene frequencies in apheresis blood donors and in leukemia patients.

Methods: Genomic DNA was extracted from 149 peripheral blood samples of apheresis regular donors and from 84 leukemia patients with chronic myeloid leukemia (CML=56), acute myeloid leukemia (AML=13), chronic lymphoid leukemia (CLL=6) and acute lymphoid leukemia (ALL=9) using a commercial DNA extraction kit (QIAamp DNA, QIAGEN, Germany). Sequence specific PCR (SSP-PCR) was performed amplifying the HPA-1a/HPA-1b gene using a pair of primer for each allele under the following conditions: the PCR was carried out in a final volume of 20μL, containing 0.2μg of genomic DNA, 0.2mM of each dNTP, 5% of glycerol, 1.5 units of platinum Taq polymerase (Invitrogen, U.S.A) in the buffer supplied, 5.0mM MgCl2 and 10.0 pmoles of each primer. Fragments of 196 bp derived from HPA-1a/1a mutation were separated for 90 minutes at 102V using 0.5μg.mL-1 ethidium bromide-stained 2.0% agarose gels and visualized in a UV light apparatus (Eagle Eye II, EUA).

Results: Table 1 shows the results of HPA1a/b genotyping and gene frequencies in apheresis regular donors and in leukemia patients. HPA-1b/HPA-1b genotype frequency in patients with AML was higher when compared to apheresis blood donors.

Conclusion: In our study, high prevalence of HPA-1b in homozygous in AML patients was detected. This data can be useful to evaluate the alloimmunization against platelet specific antigens in acute myeloid leukemia patients.