Over the last two decades, over 300,000 solid organ transplantations have been performed in the USA alone. While there has been considerable progress with immunosuppression therapies there is still significant risk of graft organ rejection, even when appropriate HLA matching has been performed. Transplant rejections from male donor to male-recipient and females to females are typically much lower than male to female. Cross-ethnic rejection rates are also higher, suggesting that additional genetic factors are at play in the rejection process. Possible sources of genetic variation underpinning rejection are homozygous deletion copy number variants (CNVs) spanning whole gene or exon regions and Loss of Function (LoF) variants ablating two copies of a given gene in a recipient but not in the doors genome, resulting in potential incompatibility across the proteomes of donor and recipient.

It has been shown through large-scale whole genome sequencing studies that the average human has 20 genes disrupted (i.e. with LoF) in two copies. We have analyzed CNV data from over 68,000 individuals in ubjected to genome-wide association studies (GWAS), mostly using 610,000 SNPs arrays, as well as published GWAS and second generation sequencing (SGS) datasets. We designed 6.5MegaBase (Mb) of targeted capture DNA for over 300 hdCNVs, and non-exonic PGx content, and spiked it into >64,000,000 bases (EZ v3.0 whole exome capture product from Roche). In total, DNA from 562 tissue/blood samples from transplant donors and recipients (lung, liver, heart and kidney) within the Clinical Trials in Organ Transplantation 3 study (CTOT3) were subjected to the 70.5Mb capture tool followed by SGS on HiSeq2000 (Illumina). We compare hdCNV and LoF carriages across the entire donor pool compare with the entire recipient pool in order to assess inherent differences between these two populations and demonstrate over 3 dozen putative candidate allogenic targets. We followed up these putative LoF targets using peptide probes to over 175,000 12-mer peptide probes across > 800 protein targets in > 100 longitudinal sera samples of CTOT3 individuals with biopsy proven rejection, or no-rejection status to investigate allo-antibody status in non-HLA as well as HLA targets.

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