Background: Development of chronic lung allograft dysfunction (CLAD) limits long-term success from lung transplantation (LTx). Elicitation of de novo antibody (Ab) to mismatched donor HLA (DSA) and lung-associated self-antigens (SAgs) are risk factors for CLAD following human LTx. Aim of this study was to analyze repertoire of non-HLA Abs and identify dominance as well as diversity of SAgs.

Methods: Plasma from LTx recipients (LTxRs) was analyzed by high-density protein microarrays. Longitudinal samples from LTxRs with CLAD were stratified based on DSA status (pre-DSA, peak-DSA and DSA-resolved) and plasma from CLAD-free stable LTxR was analyzed at one, three and four post-LTx years. The Ab specificity was evaluated by ProtoArray microchips and results were analyzed using ProtoArray Prospector v5.2 for immune response biomarker profiling. Significant SAgs identified in CLAD, but not in CLAD-free LTxRs, were analyzed by Ingenuity Pathway Analysis (IPA).

Results: Ab mapping by ProtoArray revealed a unique repertoire in LTxRs that developed CLAD compared with CLAD-free LTxRs. Fold change for Ab targets was calculated as increase in signal intensity at peak-DSA or DSA-resolved compared to pre-DSA. Overall, peak-DSA LTxRs demonstrated a significant increase in Ab signals, determined by a Z-Factor >0.4, for 1221 SAgs of which signal for 75 SAgs was significant at Z-factor >0.4, Z-score >3 and CI p<0.0001. IPA identified a number of enriched pathways including B-cell receptor signaling, S-phase entry, netrin signaling and role of TOB in T cell signaling. Furthermore, peak-DSA plasma exhibited an expanded Ab repertoire compared to that of pre-DSA, which later contracted when DSA was resolved following Ab directed therapy.

Conclusion: We document, for the first time, the repertoire of SAgs targeted by B cells following human LTx. The spread of Ab targets encompassed proteins that are expressed on cell surface, secreted and cytosolic indicating a complex immunologic priming event. Furthermore, de novo DSA was associated with development of immune responses to a spectrum of SAgs. We conclude that de novo DSA is a trigger for the development of anti-SAg humoral responses including an expanded Ab repertoire that may be involved in the pathogenesis of CLAD.

CITATION INFORMATION: Nayak D, Garcia K, Pirrotte P, Hachem R, Bremner R, Smith M, Mohanakumar T. Antibody Mapping by Protein Microarray Revealed a Unique Repertoire in Lung Transplant Recipients That Developed Chronic Lung Allograft Dysfunction. Am J Transplant. 2017;17 (suppl 3).

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