The Genomics of Chronic Allograft Rejection (GoCAR) is a multicenter study investigating the ability of differential gene expression to predict the development of Chronic allograft nephropathy (CAN). 589 patients have been enrolled. Allograft biopsies were obtained were obtained at 0, 3, 12, and 24 months post-transplant with Chronic allograft dysfunction index score (CADI) reported from a core lab. Gene expression microarray analysis was performed on 3-month biopsies (Affymetrix: human exon-1chip) and correlation to 12-month CADI and eGFR analysed (n=160). Overexpression and lentiviral suppression studies were performed on human primary tubular cells (RPTE). SHROOM3-gene expression was found to correlate linearly with fibrosis and negatively with eGFR at 1 year (n=101; p<0.05). This was confirmed by RT-PCR independently (n=36) No correlation was seen between SHROOM3 expression and 3 month CADI (n=135). A SNP in SHROOM3 has been linked to CKD in genome-wide association studies. We found that presence of 1 or 2 copies of the risk allele (ie A/G or A/A) in donors was associated with higher intragraft SHROOM3 expression at 3 months(n=136;p=0.02). The risk allele was more prevalent in white vs. non-white donors(p<0.0001). While in recipients of white-donor kidneys SHROOM3 expression was predictive of CADI at 12 mths, this was not true for non-whites. Type-1 Collagen production in RPTE was increased by SHROOM3 overexpression and markedly diminished by lentiviral knockdown (p<0.01). In RPTE, SHROOM3 overexpression facilitated, whilst knockdown inhibited, canonical TGF-Β signaling as evidenced by Smad3-phosphorylation and downstream profibrotic marker production. Further, in murine models of CKD (HIV-nephropathy & Unilateral Ureteric Obstruction), Shroom3 expression was increased compared to controls. In summary, SHROOM3 is a novel candidate gene whose expression in the renal allograft precedes and predicts the development of renal dysfunction and progression of CADI in CAN through facilitation of TGF-Β signaling. The previously described CKD-associated SHROOM3 locus likely mediates its effect through increased SHROOM3 expression. SHROOM3 may be a common therapeutic target for CKD and CAN.

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