Examining the Mechanisms of RBC Alloimmunization Hyperresponders

Summary: Red blood cell (RBC) alloimmunization can make it difficult to procure compatible RBCs for future
transfusion, which can directly increase morbidity and mortality in transfusion-dependent individuals. While
patients who develop multiple alloantibodies against distinct alloantigens are particularly challenging to manage,
the immune events during initial alloimmunization that may increase the likelihood of generating additional
alloantibodies following subsequent transfusion remain unknown. Our long-term goal is to identify immune
factors that enhance subsequent alloimmunization events in previously alloimmunized individuals in order to
prevent the accumulation of multiple alloantibodies in transfusion dependent individuals. Our central hypothesis
is that initial alloimmunization events directly enhance subsequent RBC alloimmunization by inducing CD4 T
cells that possess the ability to directly activate B cells against a completely unrelated RBC alloantigen following
subsequent transfusion. Our hypothesis is formulated on the basis of our recent discovery that B cells specific
for one antigen (the HOD (HEL, OVA and Duffy) antigen) not only internalize HOD following RBC engagement,
but likewise remove and internalize additional RBC components, suggesting that B cells may possess the ability
to remove multiple antigens following engagement of the target antigen. Consistent with this, adoptive transfer
of CD4 T cells primed by KEL RBC transfusion in the presence of poly I:C, which induces viral-like inflammation,
directly enhances alloantibody formation against the completely distinct HOD antigen following subsequent
transfusion of RBCs expressing HOD and KEL. Depletion of marginal zone (MZ) B cells, a unique B cell
population previously shown to be critical in the initiation of alloantibodies, inhibits KEL RBC priming and the
HOD RBC boost following HOD x KEL RBC transfusion, suggesting that MZ B cells work in concert with
previously recognized bridging channel 33D1+ dendritic cells (33D1+ DCs) shown to be critical in the initial
activation of CD4 T cells following HOD RBC transfusion. In contrast, while KEL RBC-induced alloimmunization
requires type I interferons (IFNab) and HOD RBC-induced alloimmunization requires toll-like receptor (TLR)
signaling, KEL-induced alloimmunization in the presence of PIC requires both IFNab and TLRs, suggesting that
while innate immune pathways may differ for KEL and HOD RBC-induced alloimmunization, PIC allows KEL
RBCs to engage TLRs and prime a subsequent HOD boost. We will use a series of pre-clinical models to define
the key priming and subsequent boosting pathways by testing the following specific aims: Aim 1: Define the role
of MZ B cells, 33D1+ DCs, IFNab and TLRs in PIC/KEL RBC-induced priming. Aim 2: Define the role of MZ B
cells, 33D1+ DCs, and TLRs in subsequent KEL-mediated HOD RBC boost. We think that successful completion
of these aims will define key immunological priming and boosting events that facilitate alloimmunization and
therefore will provide an important framework to develop rational approaches to prevent the development of RBC
alloantibodies against multiple alloantigens in chronically transfused individuals.

Grant Number: 5R01HL154034-05
NIH Institute/Center: NIH
Principal Investigator: Connie Arthur