Top-Down Proteomics in the Assessment of Kidney Donor Quality: a proof-of-concept for increased organ utilization.

A persistent barrier to optimizing the utilization of deceased donor kidneys, nearly 30% of which are discarded, is the absence of objective, molecularly resolved metrics of organ quality. To address this gap, we integrated high-resolution proteoform imaging mass spectrometry (PiMS) of donor kidney biopsies with deep LC-MS-based intact proteoform profiling of recipient peripheral blood mononuclear cells (PBMCs) and standard clinical assays. This approach enables direct delineation of proteoform landscapes and reveals immunological and cell stress signatures. Across donor types, living donor (LD) kidneys were enriched for proteoforms associated with oxidative metabolism and cellular homeostasis, whereas kidneys from donation after brain death (DBD), donation after circulatory death (DCD), and discarded organs exhibited elevated abundance of stress-responsive proteoforms, including acetylated CRYAB, PARK7, S100A4, and ACTG1. Despite more pronounced early allograft dysfunction in DCD recipients, DCD tissue proteoform landscapes were globally more similar to LD than to DBD kidneys, indicating distinct injury biology not captured by clinical metrics or peptide-centric proteomics. To extend these findings beyond the graft, we performed the first paired tissue and PBMC proteoform profiling across donor types. Baseline PBMC proteoform differences were minimal; however, by day 7 post-transplant, recipients of deceased donor kidneys exhibited platelet activation, cytoskeletal stress, and inflammatory signaling. Several PBMC-derived proteoforms correlated with serum creatinine, highlighting their potential as early, objective indicators of graft adaptation. Collectively, this proof-of-concept study defines conserved injury-associated proteoforms across tissue and immune compartments and support the idea that proteoform-resolved analysis may provide a mechanistically grounded, objective framework for assessing donor kidney quality and early graft function.