NUC-7738: A Promising ProTide Agent for AMPK Activation and Renal Cancer Cell Elimination In Vitro

Cordycepin, also known as 3′-deoxyadenosine, is recognized for its ability to inhibit cancer cell growth through various mechanisms. NUC-7738, a ProTide form of cordycepin, is developed to bypass resistance mechanisms by releasing 3’-deoxyadenosine monophosphate within cells. The hypothesis is that NUC-7738 could activate AMPK, a critical cellular energy sensor, and disrupt metabolic balance in cancer cells. Clear cell renal cell carcinoma (ccRCC), marked by lipid accumulation due to metabolic pathway dysregulation, is considered a suitable target for metabolic intervention. The activation of AMPK, indicated by the phosphorylation of AMPK (pAMPK), is linked to the downregulation of mTOR signaling, suggesting the potential of NUC-7738 to modulate this significant pathway in cancer.

The study analyzed pAMPK and AMPK expression in ccRCC from 293 patients using immunofluorescence and digital image capture, followed by analysis with QuPath software. The impact of NUC-7738 on the growth and confluence of nine renal cancer cell lines was evaluated under both low oxygen and normoxic conditions using the SRB assay and a Celigo scanner. Western blotting was utilized to assess the ratio changes of pAMPK:AMPK caused by NUC-7738, with results read using a LiCor Odyssey instrument. The effect of NUC-7738 on AMPK activation in ex vivo ccRCC tissue slices was also analyzed.

Findings indicated that while AMPK was broadly expressed in ccRCC, pAMPK expression was focal and heterogeneous. Cell lines strongly expressed pAMPK, but this expression was reduced under more physiologically relevant culture conditions, such as lower oxygen tension and glucose levels. NUC-7738 was found to inhibit the growth of renal cancer cell lines under both hypoxic and normoxic conditions, increasing pAMPK levels after various treatment durations and inhibiting mTOR activity primarily after 48 hours. NUC-7738 also increased pAMPK levels in ex vivo ccRCC tissue slices.

The study concludes that AMPK activation is generally low in primary ccRCC tissue and cell lines grown under conditions that mimic the physiological environment. NUC-7738 activates AMPK in ex vivo ccRCC tissue slices and cell lines and shows efficacy against cell lines under both oxygen conditions. The typically low expression of pAMPK in renal cancer tissue suggests that AMPK modulation may provide a therapeutic strategy for ccRCC. The results imply that the inhibition of the mTOR pathway might be one of the ways NUC-7738 exerts its anti-cancer effects.