In contrast, the combination treatment blocked the phosphorylation of EGFR and mTORC1 substrates ; thus, EGFR activation contributes to the incomplete inhibition of mTORC1 by PP242 and in combination with the EGFR inhibitor erlotinib, PP242 can completely block the mTORC1 kinase activity. The treatment of PP242 and erlotinib alone did not affect the p-AKT; but the combination treatment ablated the p-AKT. The data suggest that the inhibition of p-AKT is due to the synergistic effects; however, the mechanism remains to be elucidated. Next, we thought to examine the biological effects of the combination of EGFR and mTOR inhibitors on colorectal carcinoma cells. To this end, we Carfilzomib treated the carcinoma cell lines with erlotinib and PP242, alone and in combination, for 24 hours in the presence of EGF according to the experimental design as described in Figure 2. Cell viability assay revealed that the combination treatment produced a significant synergy in the cell growth inhibition. To examine this further, we treated the cells for 14 days in a colony formation assay and showed that the combination treatment synergistically eliminated the formation of colonies from each of these cell lines. Collectively, these results suggest that the combination of PP242 and erlotinib completely blocks both mTORC1 and mTORC2 kinase activity and synergistically inhibits the growth of colorectal carcinoma cells. PP242 has been reported to induce apoptosis in leukemia and breast cancer cells through inhibition of mTORC2 activity. The combination of P242 and erlotinib can inhibit the mTORC2 activity, which suggests that the combination may induce apoptosis in colorectal carcinoma cells. To text this notion, we treated DLD-1 cells with PP242 or erlotinib, alone and in combination in the presence of EGF. Flow cytometry detected approximately 5% sub-G1 apoptotic cells in the cells treated with PP242 or erlotinib alone. In contrast, approximately 15-20% cells underwent apoptotic cell death under the combination treatment of PP242 and erlotinib. To confirm the PP242 and erlotinib combination-induced apoptosis, we Foretinib side effects examined the treated cells on western blotting and thus revealed cleavage of caspase-3, DFF45 and PARP; the data confirmed the apoptotic cell death of the cells under the treatment of PP242 and erlotinib, in contrast, however, PP242 and erlotinib alone failed to induce apoptosis in the cells. These results suggest that the combination of PP242 and erlotinib synergistically inhibits the growth of colorectal carcinoma cells in part through induction of apoptosis. To evaluate therapeutic potential of the combination of PP242 and erlotinib in treating colorectal carcinoma, we generated subcutaneous xenografts by injecting subcutaneously DLD-1 cells in athymic mice.