Role of IFITs in RBC Alloimmunization - Approximately 1 out of every 70 people (~5,000,000 patients) are transfused each year in the USA alone. Over 340 red blood cell (RBC) alloantigens have been identified in addition to the well-known ABO and RhD antigens. Over time, some patients generate antibody responses against so many RBC alloantigens that compatible RBC units become increasingly difficult to identify, and in some cases are unavailable. This leads to considerable morbidity and in extreme cases mortality. Despite the large number of alloantigens, only a minority of patients ever become alloimmunized (~2-5%). This remains very significant given the number of people transfused each year and leads to the question of why certain people become alloimmunized and others do not. Moreover, patients tend to be either “responders” with a relatively high frequency of alloimmunization or “nonresponders” with no detectable alloantibodies even after chronic transfusion. Responder vs. nonresponder status has been shown to have both environmental and genetic components. Viral infection with RNA viruses at the time of transfusion is associated with RBC alloimmunization in both mice and humans and has been shown to cause alloimmunization in mice by inducing interferon-alpha and signaling through the interferon-alpha/beta receptor (IFNAR). Preliminary data presented in this grant demonstrate that murine cytomegalovirus (mCMV) infection induces RBC alloimmunization through IFNAR independent pathways. Less progress has been made in defining genetic factors that regulate responder vs. nonresponder status. A number of genome wide association studies in humans have identified potential candidates, but running trials with sufficient statistical power is challenging. In such cases, the use of animal models is a reasonable alternative. We have reported that, much like in humans, there is substantial variability in genetics of responder vs nonresponder biology across different strains of inbred mice. Using RBC alloimmunization as a trait, we performed a GWAS study on 584 mice from a diversity outbred population and identified a region of chromosome 19 associated with alloimmunization (LOD score of 8.2, genome-wide p-value (0.036)). This region contains a cluster of IFIT (IFIT1-IFIT3) genes (interferon-induced protein with tetratricopeptide repeats) that are interferon- alpha response genes downstream of IFNAR signaling. Moreover, IFIT induction also occurs downstream of mCMV infection. Thus, the two known pathways of RBC alloimmunization have IFIT induction as a common distal event. Although best known for antiviral function, IFIT genes are increasingly being appreciated as being immunoregulatory. Thus, multiple lines of evidence support the Central Hypothesis of this application: IFIT1 and IFIT2 suppress while IFIT3 enhances RBC alloimmunization. The current application proposes to use mice with deletions of IFIT1, IFIT2, IFIT3 or the whole IFIT locus to test their causal roles in RBC alloimmunization. The proposal is structured as a single Specific Aim, testing the role of each IFIT gene member individually, or as a group, in RBC alloimmunization.
