Non-invasive foetal RHD genotyping in transfusion medicine.

Abstract

Non-invasive genetic tests using foetal DNA in maternal plasma were developed largely to reduce pregnancy loss related to invasive diagnostic tests for the inheritance of foetal genetic and chromosomal abnormalities1. The immediate application in transfusion medicine was to determine the foetal inheritance of RHD among pregnant women alloimmunised to the D antigen. The article by Laura Salvaneschi et al3 in this issue of Blood Transfusion underscores the value of an international approach to validating any diagnostic test used to predict inheritance of foetal RHD. In the past, the foetal risk of anti-D haemolytic disease of the foetus or newborn (HDFN) was determined from amniotic fluid or chorionic villi-derived DNA and PCR-based assays which accurately detected RHD when it was present but carried a specific level of risk of pregnancy loss and transplacental haemorrhage leading to increased antibody titres. Assays were modified to detect variant and nonfunctional RHD alleles as they were characterised4,5. Foetal RHD typing from maternal plasma has benefited from refinements of specific RHD targets that ensure accuracy, which has been adopted by the International Society for Blood Transfusion (ISBT) Working Party on Red Cell Immunogenetics and Blood Group Terminology. The immediate goal of testing foetal RHD in maternal plasma is to avoid the use of invasive procedures to assess the risk of HDFN, with the option of returning alloimmunised pregnant women to their primary care physician when the foetus is predicted to be RhD-negative. The immediate outcome addressed is a reduction of foetal morbidity/mortality in the management of HDFN. It is particularly noteworthy that Salvaneschi et al chose a validation plan that integrated international standards. Their validation plan incorporated evaluations of the specificity, sensitivity, variability and robustness of an internationally established protocol, with specific exon targets for the detection of foetal RHD in maternal plasma, and international reference reagents6,7. They included samples from the 2010 ISBT workshop, blinded patients’ samples, and a reference reagent from the National Institute for Biological Standards and Control. The traceability of their work to the international protocol and standards was obtained because the reference reagent was used in the First Collaborative Study to establish the sensitivity standard for non-invasive testing of foetal RHD in maternal plasma. The authors were astute to outline the next steps required to evaluate the feasibility of foetal testing in the prenatal care of all RhD-negative pregnancies. Their comment that “international collaborations are essential for monitoring the performance of the laboratory” resonates well in both perinatal care and transfusion medicine. Italy is poised now to evaluate the clinical, ethical, and economic impact of early gestational testing of foetal RHD in maternal plasma using rapid and high-throughput analyses to limit the use of Rh immune globulin for RhD-negative women whose foetus is RhD-negative.