(P-IG-12) Are Compound Heterozygous Partial D Individuals at risk for allo-anti-D?

Background/Case Studies: There are hundreds of RHD variant alleles, with many expressing partial RhD antigens lacking one or more epitopes. When an individual expresses RhD antigen lacking an epitope and is exposed to that epitope via blood transfusion or pregnancy, they are at risk of alloimmunization. The impact of compound heterozygosity among various RhD variants is not well studied. We used a panel of anti-D monoclonal antibodies to examine agglutination of RBCs expressing only one partial RhD antigen or two distinct partial RhD antigens. This work can be used to predict the risk of RhD alloimmunization in specific compound heterozygote pairs.

Study

Design/Methods: RHD genotyping was performed using RHD BeadChip (Immucor) and PCR-RFLP for RHD c.1136C>T. Specimens identified by RHD genotyping to be hemizygous or homozygous for a specific partial D antigen and those compound heterozygous for two different partial D antigens were evaluated for agglutination with the unlicensed Quotient RhD Variant Investigation Kit (Albaclone) of human/murine monoclonal IgG/IgM reagents.

Results/Findings: Two samples compound heterozygous for partial D alleles were identified: RHD*DIV/RHD*DVI (Sample 3) and RHD*DAR/RHD*DAU5 (Sample 4). Agglutination of sample 3 was compared to RBCs carrying RHD*DVI (Sample 1) or RHD*DIV (Sample 2), while agglutination of Sample 4 was compared to RBCs carrying RHD*DAR (Sample 5). Agglutination results are shown in Figure 1. While RBCs carrying RHD*DVI (Sample 1) or RHD*DIV (Sample 2) lacked several epitopes, the RHD*DVI/RHD*DIV RBCs reacted with all antisera in the panel. While RBCs carrying RHD*DAR (Sample 5) did not agglutinate with 4 antisera, the RHD*DAR/RHD*DAU5 RBCs reacted 3 of those but showed no agglutination with LHM174/102.

Conclusions: There are hundreds of variant RHD alleles with many expressing partial D phenotype. When a patient carries two different partial D alleles, the risk of RhD alloimmunization is not known. By examination of the reactivity with a panel of anti-D monoclonal antisera, epitope presence or loss can be determined. One of the two compound heterozygotes studied here (RHD*DIV/RHD*DVI) showed 3+ to 4+ reactivity with all anti-D reagents suggesting all epitopes tested by the panel are present. The other compound heterozygote (RHD*DAR/RHD*DAU5) failed to agglutinate with LHM174/102 reagent. This supports that these RBCs lack at least one epitope and therefore suggests that the patient is at risk of anti-D alloimmunization. This approach of evaluation of epitope loss and tracking of outcomes in patients with this combination can inform about anti-D risk and aid in more personalized management, especially in women of child-bearing age.

Importance of research: There are hundreds of variant RHD alleles, many with partial D phenotypes. It is unclear if individuals who carry two different partial D alleles are at risk of allo-anti-D. This study evaluates the epitopes of compound heterozygous RBCs versus those expressing only one partial D antigen using a monoclonal RhD typing kit. This information is useful in helping to understand the risk of alloimmunization in compound heterozygotes.