Elucidating the effect of caffeine on NHEJ repair pathway

Genome stability and maintenance of its integrity is of utmost importance for the survival of any organism. Human genome is under constant threat by several endogenous and exogenous factors. In order to maintain genomic stability, cells have evolved different DNA repair pathways. Caffeine, an analogue of adenosine, is one of the commonly consumed stimulants across the world. It has been reported that caffeine inhibits the protein kinase activity of ATM and ATR (PI3KK family) although certain other studies show that caffeine failed to affect ATM autophosphorylation, which is a crucial step in ATM activation. Further, Caffeine treatment disrupted the formation of Rad51 filaments and thereby inhibiting the repair by HR. Thus, although certain studies suggest that Caffeine could play a role in DNA repair, its effect on NHEJ and MMEJ are poorly understood. It is also unclear, whether such an effect could contribute towards oncogenesis/cancer therapy. Using an array of biophysical and biochemical methods, we show that incubation with caffeine lead to significant reduction in DNA end joining when oligomeric DNA substrate harbouring DSBs were incubated with cell-free extract prepared from rat testes. The observed inhibition was consistent when joining was performed using sticky, blunt and non-compatible ends. Further, its impact on MMEJ is being evaluated. Using immunofluorescence assay, we show that there is a dose dependent increase in DNA breaks and cell death in mammalian cells. Studies using mice model showed that upon caffeine treatment (50 mg/kg body weight), there was a significant regression of tumor development as compared to the control. In summary, it appears that intake of Caffeine at certain concentration may lead to inhibition of DNA repair leading to accumulation of DNA breaks in cancer cells resulting in tumor regression. (author)