Immunohematology and Genetic Testing (red cells/leukocytes and platelets) - Immunohematology (includes serology)
(P-IG-46) Third Example of Exceedingly Rare Anti-ERSA (Er4) in a Pregnant Native American: Clinical Relevance in HDFN and Serologic Findings of Her ERSA- Phenotype
New York Blood Center Enterprises, Immunohematology and Genomics Laboratory New York, New York, United States
Background/Case Studies: The Er blood group system consists of 5 antigens (Ag): 4 high prevalence Era, Er3, ERSA , ERAMA and low prevalence Erb. Ags reside on Piezo-type mechanosensitive ion channel component 1 (Piezo-1) and are encoded by PIEZO1. Although Piezo-1 is large protein, it is present in only a few hundred copies per RBC but has wide tissue distribution. PIEZO1 pathogenic mutations can be associated with blood disorders or non-immune hydrops fetalis. Data on clinical relevance of antibodies (Ab) to Er Ags is sparce, but 1 example of anti-Er4 has been associated with severe HDFN. We studied a sample from multiparous G4P3 pregnant Native American with an Ab to a high prevalence Ag.
Study
Design/Methods: Serologic testing was by standard hemagglutination. Rare RBCs and Abs were from our in-house collection. RBCs were treated with papain, trypsin, dithiothreitol (DTT) or EDTA glycine acid (EGA, Werfen). Genomic DNA was isolated from WBCs. Sanger sequencing of PIEZO1 exons 24, 37-39, 45-51 and flanking introns was done.
Results/Findings: The patient was group O+. Patient plasma (PP) reacted 1+-2+ by IAT with all RBCs tested; autocontrol was negative (neg); reactivity remained with papain, trypsin or DTT treated RBCs. Patient RBCs (PR) were tested with a battery of Abs to high prevalence Ags; all were positive but anti-Era reacted only weakly and suggested possible Er related Ab. PP reacted mi+ with 3 Er(a-) RBCs and was neg with EGA treated RBCs. PR reacted mi+ with 4 other anti-Era; this showed a likely Er target Ag, but not Era. PIEZO1 sequencing identified homozygous nonsynonymous changes, c.5538C >G, 6570A >G, 6642G >C, 7059T >C, 7500C >T and homozygous for missense variant c.7219G >A (p.2407Lys) predicting ERSA- (ER:-4).
Conclusions: We report the 3rd known case of anti-ERSA in a multiparous woman. She was never transfused or had prenatal care. Records noted 1st positive Ab screen at birth with G2; all deliveries were term and unremarkable with no indication of HDFN. Variants c.7219G >C (Glu2407Gln) or 7219G >A (Glu2407Lys) define a ERSA- phenotype. A p.Gly2394 is required for Era expression. Lack of Gly2394, or a p.Glu2392Lys mutation in cis, results in a Er(a-) phenotype. Crew et al (Blood,141,2: 135) reported weakened Era on some but not all ERSA- RBCs (papain treated). Their structural modeling of p.2392Lys suggested it impacts protein conformation around the Era epitope. Here, the c.7219G >A (p.Glu2407Lys) variant in PIEZO1 gene is associated with markedly weakened expression of Era antigen as it was nearly undetectable on patient RBCs with 4 of 5 anti-Era tested. Substitution of glutamic acid with a positively charged lysine at position 2407 may alter the conformation of Piezo-1 partially masking or disrupting Era. Also the anti-ERSA in this patient reacted exceedingly weaker with Er(a-) RBCs. This study highlights the ongoing and increasing value of combined serology and genotyping to resolve complex cases.
