Running Title: Apoptotic Effect of Mycolactone in SCC15 Cells

ABSTRACT

The effect of mycolactone, a recently reported apoptosis-inducing factor, was investigated in SCC15 oral squamous cell carcinoma (OSCC) cell line. Mycolactone rapidly induced cell death in OSCC cells in 2days, which was similar to that found in apoptotic cell such as detaching from culture plate and rounding-up of cells. Apoptotic cells were increased 4hrs after mycolactone treatment and more than half of cells showed apoptosis after 72hrs. Caspase 3 activation, a biochemical evidence of apoptosis, was determined by Western blotting. Caspase 3 activation was started at 2hrs that lasted until 8hrs after mycolactone treatment. The expression of bcl-2 family genes was determined to explain the mechanism of apoptosis found in OSCC cells. The expressions of bad, bak, and bax (pro-apoptotic genes) and bcl-w and bcl-2 genes (anti-apoptotic genes) were not changed by mycolactone treatment. The expression of bcl-XL was decreased 8 hrs after mycolactone treatment. Mcl-1 expression was initially increased at 2 hrs which was decreased 8 hrs after mycolactone treatment. The down-regulation of these two anti-apoptotic genes might explain the mycolactone-induced apoptosis in OSCC cells. In this study, mycolactone was revealed to induce cell death in OSCC cells via apoptosis and the apoptosis mechanism of OSCC cells was shown to be down-regulation of anti-apoptotic genes, bcl-XL and mcl-1. These results suggested the applicability of mycolactone for the development of an anti-cancer drug candidate by inducing apoptosis of OSCC cancer cell.