*4.5.1. Materials*


#### *4.5.2. Methods*


Cells were treated with LY294002, which is a cell-permeable inhibitor for phosphoinositide 3-kinase (PI3K) that acts on the enzyme ATP binding site (**Table 1**). As PI3K is strictly required for autophagy, PI3K inhibition sequentially leads to autophagy inhibition. As shown in **Figure 4**, both CCD112 and HT29 showed an increase in LC3B signal after starvation and LY294002 treatment. The signal is correlated to the increase in autophagosome formation after the treatment. However, such result was not obvious for HCT116, which has a high autophagosome formation in the untreated cell lines. This may be a result of continuous activation of autophagy pathway due to KRAS mutation.
