fig3

Figure 3. MEKi withdrawal from BRAF^V600E-amplified HT6244-R cells causes cell death with features of apoptosis and authophagy and ultimately reversal of MEKi-resistance. BRAF^V600E amplification results in an enlarged p-MEK1/2 pool that reinstates p-ERK1/2 in selumetinib-resistant HT29 (HT6244-R) cells to parental HT29 levels in the presence of the MEKi selumetinib (left). This level of ERK1/2 activity maintains normal cell proliferation and survival. However, when selumetinib is withdrawn (middle), this enlarged pool of p-MEK1/2 is no longer restrained and levels of p-ERK1/2 increase to ~5 times those in parental cells. This ERK1/2 hyperactivation drives expression of pro-apoptotic NOXA and tBID, as well as processing of LC3, and cell death with features of apoptosis and autophagy. Prolonged MEKi withdrawal and cell death ultimately selects for reversal of selumetinib resistance (revertant HT6244-R(-), right). This reversal of MEKi resistance is due to loss of BRAF^V600E amplification in these revertant cells and a re-setting of BRAF and p-ERK1/2 back to parental HT29 levels. P: phosphate group