BK VIRUS INFECTION

A. Liacini¹, L.A. Tibbles^2
1Medicine, University of Calgary, Calgary/CANADA, ²Medicine/nephrology, University of Calgary, Calgary/AB/CANADA

Body: Introduction: Human polyomavirus BK is the causative agent in post-transplant BK Nephropathy, which is now the leading cause of early renal graft loss. Since BK Nephropathy emerged during the era of more intensive immunosuppression, the current therapy is reduction of immunosuppressive drugs. No randomized clinical trials have supported this therapy, or quantified the inherent risk of increased rejection with decreased immunosuppression. In order to understand the pathogenesis of BK virus, we examined the host intracellular signaling pathways triggered by viral infection. We hypothesized that inhibition of the intracellular protein kinase pathways activated by BK virus may be an effective therapeutic strategy. Methods: We have developed two models of human renal tubular cell infection with BK virus, using either commercially available renal tubular epithelial cell lines CCD1103, CCD1105, or Human Primary Tubular Cells (HPTCs) exposed to BK virus. Results: In our acute infection model, we found increased phosphorylation of p70S6K, mTOR, PDK1 and Akt at 4 days post infection. In order to determine the mechanism of BK virus activation of the mTOR pathway, we have shown that BK virus large T antigen interacts with Protein Phosphatase 2A (PP2A), which regulates Akt phosphorylation upstream of the mTOR pathway. To inhibit this pathway, we employed the mTOR inhibitor Rapamycin, which repressed p70S6K phosphorylation and reduced the BK virus large T antigen expression in a dose dependent manner in cell lines. We then investigated the antiviral/immunosuppressive drug Leflunomide (using the active metabolite A77 1726) which decreased PDK1 and Akt phosphorylation, but did not affect p70S6K phosphorylation status. Nonetheless, BK virus genome replication and early gene expression were remarkably inhibited in primary and established cell lines by leflunomide in a dose dependent manner alone and in combination with rapamycin. The antiviral effect of leflunomide was not reversed by uridine addition, implicating leflunomide’s tyrosine kinase inhibition activity as the major anti-BK mechanism, rather than pyrimidine depletion. The combination of rapamycin and leflunomide completely inhibited BK virus genome replication and large T antigen expression, and PDK1, Akt, mTOR and p70S6K phosphorylation. Clinically, BK virus reactivation may be well established prior to antiviral therapy, so we developed a chronically infected model which consists of either primary cells (HPTC), CCD1103 or CCD1105 cell lines infected with BKV for 4 days. These cells express BK viral proteins constitutively for more than 2 weeks. These chronically infected cells, when exposed to rapamycin, leflunomide or both, 24-72 hours post infection, dramatically reduce the expression of BK large tumor antigen (large T Ag). Conclusions: Based on these results, we suggest that the inhibition of protein synthesis pathways with a combination of rapamycin and leflunomide may be an effective therapy for BK virus reactivation in the transplant population. Since both rapamycin and leflunomide posess immunosuppressive activity, combination therapy with these drugs may reduce BK pathogenesis while maintaining appropriate transplant immunosuppression.

Disclosure: All authors have declared no conflicts of interest.