OFF-Target Effects of Mycophenolic Acid on BK Polyomavirus Replication in Primary Human Renal Proximal Tubular Epithelial Cells

Abstract

Tacrolimus (TAC) and mycophenolic acid (MPA) effectively reduce allograft rejection in kidney transplant recipients by inhibiting donor-specific lymphocyte activation and proliferation, respectively. However, this desired on-target effect is associated with uncontrolled BK polyomavirus (BKPyV)-replication and premature kidney allograft failure in 10%-20% of recipients. Besides impairing BKPyV-specific immunity, TAC also stimulates BKPyV-replication directly in renal proximal tubule epithelial cells (RPTECs) as an off-target effect. We now investigated off-target effects of MPA on BKPyV-replication in RPTECs. Following BKPyV exposure for 2 h, MPA inhibited BKPyV replication as shown by reduced supernatant BKPyV loads (IC50 0.65 M), number of infected cells, and viral protein expression at 72 hours post-infection (hpi). Notably, MPA inhibition of the viral large tumor antigen (LTag) was associated with the appearance of a truncated Tag of [~]17 kDa (truncTag-17). Adding guanosine (GUO) reversed MPA inhibition of BKPyV replication and viral protein expression, while truncTag-17 disappeared. Time course studies indicated that MPA inhibition was operative from 24 h before up to 24 h after BKPyV infection. GUO reversed MPA inhibition when administered from 24 h before to 24 h after BKPyV infection. Deep sequencing of passaged supernatant BKPyV genomes revealed more guanine-replacement (C[->]T or G[->]A) mutations for MPA compared to MPA/GUO (35% versus 10%, read threshold >0.5%). Thus, MPA can exert direct off-target effects on BKPyV replication in RPTECs linked to truncTag-17 expression, which can be partially reversed by exogenous GUO. ImportanceReducing immunosuppression is currently recommended for kidney transplant recipients with BKPyV-DNAemia and nephropathy to improve virus-specific immune control, but increases the risk of T-cell and antibody-mediated rejection. Although TAC and MPA synergize in their on-target effects on lymphocytes, they differ in their off-target effects on RPTECs. Unlike TAC, MPA inhibits BKPyV replication in an uncompetitive dose-dependent manner. MPA inhibition can be partially reversed by exogenous GUO using the purine salvage pathway that is lacking in lymphocytes. MPA acts during the early viral replication phase that depends on LTag promoting cellular G1[->]S-phase progression. MPA-mediated GUO depletion not only reduces rapid cell proliferation similar to the well-known gastrointestinal or hematopoietic toxicities, but also slows viral replication, induces expression of a truncTag-17, and increases GC-mutation rates in progeny virus genomes. We discuss the relevance of these off-target characteristics of MPA in the optimized management of kidney transplant recipients with BKPyV-DNAemia and -nephropathy.