Polycystin-1 but not polycystin-2 deficiency causes upregulation of the mTOR pathway and can be synergistically targeted with rapamycin and metformin

Abstract

Autosomal dominant polycystic kidney disease (ADPKD) is caused by loss-of-function mutations in either PKD1 or PKD2 genes, which encode polycystin-1 (TRPP1) and polycystin-2 (TRPP2), respectively. Increased activity of the mammalian target of rapamycin (mTOR) pathway has been shown in PKD1 mutants but is less documented for PKD2 mutants. Clinical trials using mTOR inhibitors were disappointing, while the AMP-activated kinase (AMPK) activator, metformin is not yet tested in patients. Here, we studied the mTOR activity and its upstream pathways in several human and mouse renal cell models with either siRNA or stable knockdown and with overexpression of TRPP2. Our data reveal for the first time differences between TRPP1 and TRPP2 deficiency. In contrast to TRPP1 deficiency, TRPP2-deficient cells did neither display excessive activation of the mTOR-kinase complex nor inhibition of AMPK activity, while ERK1/2 and Akt activity were similarly affected among TRPP1- and TRPP2-deficient cells. Furthermore, cell proliferation was more pronounced in TRPP1 than in TRPP2-deficient cells. Interestingly, combining low concentrations of rapamycin and metformin was more effective for inhibiting mTOR complex 1 activity in TRPP1-deficient cells than either drug alone. Our results demonstrate a synergistic effect of a combination of low concentrations of drugs suppressing the increased mTOR activity in TRPP1-deficient cells. This novel insight can be exploited in future clinical trials to optimize the efficiency and avoiding side effects of drugs in the treatment of ADPKD patients with PKD1 mutations. Furthermore, as TRPP2 deficiency by itself did not affect mTOR signaling, this may underlie the differences in phenotype, and genetic testing has to be considered for selecting patients for the ongoing trials.

Vasopressin and disruption of calcium signalling in polycystic kidney disease

“…Yamaguchi et al showed that inhibition of calcium-dependent phosphatidylinositol 3-kinase activity and downstream inhibition of Akt in mouse collecting duct epithelial cells allows B rapidly accelerated fibrosarcoma (B-Raf) and ERK to be activated in a PKA-, Src-, and Ras-dependent manner and underlie the proliferative response to cAMP in the setting of reduced calcium. 119 Consistent with these observations other studies have shown that knockdown of PC1 or PC2, reduction in intracellular calcium and downregulation of calcium calmodulin kinase II (CaMKII) inhibit Akt and activate ERK, 137,138 and that knockdown of CaMKII induces pronephric cysts in zebrafish. 139 Similarly, inhibition of PKA-associated proliferation by elevation of intracellular calcium in cholangiocytes derived from PCK rats has been shown to be dependent on the activation of PI3K and Akt.…”

Double inhibition of cAMP and mTOR signalling may potentiate the reduction of cell growth in ADPKD cells

“…Consistently, it has been reported that the double treatment with Tolvaptan and Pasireotide, adenylyl cyclase 6 inhibitors, induces an additive beneficial effect on renal function in a hypomorphic Pkd1 model [24]. Moreover, the double treatment with rapamycin and metformin in renal cells is more effective for inhibiting mTOR activity than molecules used alone [25]. …”

The combination of metformin and 2‐deoxyglucose significantly inhibits cyst formation in miniature pigs with polycystic kidney disease

“…The glucose analogue 2DG acts as a competitive inhibitor of the glycolysis rate-limiting enzyme hexokinase (Brown, 1962), leading to an inhibition of cell cycle progression and cell death in vitro (Maher et al, 2004). Metformin is an AMPK activator, and studies have explored the use of metformin in the context of ADPKD, both in vivo and in vitro (Takiar et al, 2011;Mekahli et al, 2014). In addition, metformin can inhibit the mitochondrial oxidative phosphorylation activity of Complex I (El-Mir et al, 2000).…”