A Single Point Mutation Gln₇₁₆His on the α₂ Integrin: The New Platelet Alloantigen Cas^a.

We report here the identification and characterization of a new low-frequency platelet alloantigen Cas^a involved in a case of neonatal alloimmune thrombocytopenia (NAIT). A 29-year-old mother gave birth to a full-term male infant who exhibited petechiae at birth. Nine hours post-delivery the platelet counts revealed a severe thrombocytopenia (16.10⁹platelets/L) leading to platelet transfusion therapy associated with IVIG. The outcome was uneventful. Blood samples from the parents and infant were referred to our laboratory for investigation because of suspected NAIT. Maternal serum showed a specific positive reaction with the antigen-capture assay (MAIPA) only when it was tested with the paternal platelets and the monoclonal antibodies Gi9 (Immunotech, Marseille, France), P16 (NIBSC, Bristol, UK), and AK7 (Abcys, Paris, France) directed against the GPIa-IIa (a₂b₁ integrin). Nucleotide sequence analysis of GPIa cDNA of the father and newborn showed a nucleotide substitution at position 2235 (2235G>T according to the International Nomenclature). This substitution induces a Q716H amino acid change in the GPIa mature protein, located outside the I domain involved in cell-adhesion for collagen. In vitro analysis of recombinant CHO cells expressing wild-type or mutant (Q716H) human GPIa allowed us to demonstrate that this single amino-acid substitution is responsible and sufficient for inducing Cas^a antigen expression. Adhesion of CHO cells to collagen coated on microtiter plates was not modified by the Cas polymorphism, nor by the maternal anti Cas^a alloantibodies, indicating that the mutation does not affect the function of the integrin a₂b₁. PCR-SSP was developed for Cas^a genotyping. In a Caucasian population study none of the 100 unrelated blood donors was found to be Cas^a carrier. In conclusion, the Cas^a antigen described here, implicated in a case of severe neonatal thrombocytopenia is a low-frequency platelet antigen in the Caucasian population. This study highlights the high polymorphism of the GPIa gene.