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. Author manuscript; available in PMC: 2020 Jul 31.
Published in final edited form as: Cell Rep. 2020 Jun 16;31(11):107764. doi: 10.1016/j.celrep.2020.107764

Figure 4. Concurrent RAF and ERK Inhibition Causes Apoptosis.

Figure 4.

(A) (left) Fold changes (log2) of RNA expression of pro-apoptosis and pro-survival genes as the averaged values of MIA PaCa-2, Pa02C, Pa14C, and Pa16C cell lines treated with RAFi/ERKi (0.3 μM and 0.04 μM, respectively) and compared to RAFi (0.3 μM) (24 h). Error bars are shown as standard error. All p values shown are in comparison to the vehicle control for individual graph; p values are from Wald test. Adjusted p values: BAX (0.2971), BAK1 (0.7226), BAD (*, 0.0332), BID (0.3363), BIK (0.2635), HRK (**, 0.003), BCL2L11 (****, 1.49E-5), PUMA/BBC3 (*, 0.0221), CASP1 (0.3642), CASP2 (**, 0.0034), CASP3 (0.6314), CASP4 (0.1526), CASP6 (0.5876), CASP7 (0.5114), CASP8 (0.9215), CASP9 (0.2887), BCL2 (***, 0.0014), BCL2L1 (0.1569), MCL1 (0.9338), BCL2L2 (0.9080). (Right) fold changes (log2) of the transcripts of the cells treated with RAFi/ERKi (0.3 μM and 0.04 μM, respectively) and compared to ERKi (0.04 μM) (24 h). Error bars are shown as standard error. All p values shown are in comparison to the vehicle control for individual graph; p values are determined from the Wald test. Adjusted p values: BAX (***, 0.0003), BAK1 (0.4834), BAD (0.6736), BID (0.1958), BIK (0.2171), HRK (***, 0.0011), BCL2L11 (****, 7.12E−6), PUMA/BBC3 (*, 0.0148), CASP1 (0.3406), CASP2 (*, 0.0250), CASP3 (0.7561), CASP4 (*, 0.0229), CASP6 (*, 0.0615), CASP7 (0.1599), CASP8 (0.4119), CASP9 (0.2582), BCL2 (****, 0.0001), BCL2L1 (**, 0.0032), MCL1 (0.7493), BCL2L2 (0.9250).

(B) Pa01C, Pa02C, or Pa14C cells were treated with vehicle, RAFi (0.3 μM), ERKi (0.04 μM), or the combination (72 h). Cell lysates were immunoblotted to determine levels of the indicated proteins.

(C) Percent apoptosis of Pa02C and Pa16C cells treated with the vehicle control, RAFi (0.3 or 0.6 μM), ERKi (0.04 or 0.08 μM), or the combinations (120 h). Error bars are shown as ± SEM.

(D) Representative images of percent apoptosis of the cell lines in (C). Fluorescence-activated cell sorting (FACS) analysis was used to measure apoptosis.