A Combined HLA Molecular Mismatch and Expression Analysis for Evaluating HLA-DPB1 de novo Donor-Specific Antibody Risk in Pediatric Solid Organ Transplantation: Implications for Solid Organ & Hematopoietic Stem Cell Transplantation

Background HLA molecular mismatch (MM) has been shown to be a risk factor for de novo donor-specific antibody ( dn DSA) development in solid organ transplantation (SOT). HLA expression differences have also been associated with adverse outcomes in hematopoietic cell transplantation. We sought to study both MM and expression in assessing dn DSA risk. Methods One-hundred-and-three HLA-DP-mismatched SOT pairs were retrospectively analysed. MM was computed using amino acids (aa), eplets and, supplementarily, Grantham/Epstein scores. DPB1 alleles were classified as rs9277534-A (low-expression) or -G (high-expression)–linked. To determine the associations between risk factors and dn DSA, logistic regression, linkage disequilibrium (LD) and population-based analyses were performed. Results A high-risk AA:GX (recipient:donor) expression combination (X=A or G) demonstrated strong association with DP- dn DSA (p=0.001). MM was also associated with DP-dnDSA when evaluated by itself (eplet\_p=0.007, aa\_p=0.003, Grantham\_p=0.005, Epstein\_p=0.004). When attempting to determine the relative individual effects of the risk factors in multivariable analysis, only AA:GX expression status retained a strong association (RR=18.6, p=0.007 with eplet; RR=15.8, p=0.02 with aa), while MM was no longer significant (eplet\_p=0.56, aa\_p=0.51). Importantly, these risk factors are correlated, due to LD between the expression-tagging SNP and polymorphisms along HLA-DPB1. Conclusions The MM and expression risk factors each appear to be strong predictors of DP- dn DSA and to possess clinical utility; however, the two risk factors are closely correlated. These metrics may represent distinct ways of characterizing a common overlapping dn DSA risk profile, but they are not independent. Further, we demonstrate the importance and detailed implications of LD effects in risk assessment of dn DSA and possibly transplantation overall. ### Competing Interest Statement J.L.D., D.F., and D.S.M. receive royalties from Omixon Inc. ### Funding Statement This work was supported by institutional funds from The Children’s Hospital of Philadelphia to D.S.M. Neither the authors nor their institutions received payment or services at any time from a third party for any aspect of the submitted work. ### Author Declarations All relevant ethical guidelines have been followed; any necessary IRB and/or ethics committee approvals have been obtained and details of the IRB/oversight body are included in the manuscript. Yes All necessary patient/participant consent has been obtained and the appropriate institutional forms have been archived. Yes I understand that all clinical trials and any other prospective interventional studies must be registered with an ICMJE-approved registry, such as ClinicalTrials.gov. I confirm that any such study reported in the manuscript has been registered and the trial registration ID is provided (note: if posting a prospective study registered retrospectively, please provide a statement in the trial ID field explaining why the study was not registered in advance). Yes I have followed all appropriate research reporting guidelines and uploaded the relevant EQUATOR Network research reporting checklist(s) and other pertinent material as supplementary files, if applicable. Yes The data contained in this study can be made available upon reasonable request. * aa : amino acid CHOP : Children’s Hospital of Philadelphia dn DSA : de novo donor-specific antibody FWP : frequency-weighted permutation GVHD : graft versus host disease HLA : human leukocyte antigen HSCT : hematopoietic stem cell transplantation IRB : Institutional Review Board LD : linkage disequilibrium MFI : mean fluorescence intensity MM : molecular mismatch MW-U : Mann-Whitney U test NGS : next-generation sequencing SAB : single-antigen bead SDC : supplemental digital content SNP : single-nucleotide polymorphism SOT : solid-organ transplantation TCE : T-cell epitope