Targeting erythroid cell-derived asparagine inhibits alloimmunization in sickle cell disease

Red blood cell (RBC) transfusions remain a life-saving therapy for patients with sickle cell disease (SCD), yet their safety and efficacy are limited by a high incidence of alloimmunization. Emerging evidence suggests that metabolites, including amino acids, can influence humoral immune responses. Here, we found that levels of l-asparagine (Asn), a metabolite previously implicated in immunoregulation, are increased in both plasma and erythroid cells in SCD mice. Reciprocal transfusion indicated that circulating erythroid cells contribute to plasma Asn levels, as transfusion of normal RBCs into SCD mice reduced plasma Asn, whereas transfusion of SCD erythroid cells into wild-type mice increased plasma Asn. We further observed that increased mitochondria-positive (Mito+) erythroid cells in SCD were associated with enhanced Asn synthesis. Functionally, depletion of Asn using asparaginase (ASNase) reduced RBC alloimmunization in SCD mice, albeit more strongly than in control mice, whereas exogenous Asn administration increased alloimmunization in wild-type mice. In SCD mice, treatment with ASNase was associated with inhibition of plasma cell differentiation while sparing resting B cells. In vitro studies revealed that ASNase inhibited, whereas Asn supplementation promoted, human B-cell differentiation, with a more pronounced effect in cells from patients with SCD. ASNase treatment was also associated with reducing Src family kinase (SFK) activation in SCD, suggesting a potential link between Asn and SCD B-cell signaling. Together, these findings point to a potential role for Mito+ erythroid cell Asn-SFK axis in regulating RBC alloimmunization in SCD. Modulation of this pathway may provide a basis for future therapeutic exploration.