Figure 5. Anti-BRAF^V600E (vemurafenib) and anti-BCL2/MCL1 (obatoclax) combined therapy increases cell death in patient-derived angioinvasive papillary thyroid carcinoma (PTC) cells harboring BRAF^V600E and with somatic copy number alterations (SCNAs) of MCL1 or P16.

A. Flow cytometry analysis of non-synchronized cells normalized to the number of events in each condition shows that 10 μM vemurafenib or 100 nM obatoclax at 48 hours post-treatment partially increased sub-G1 (apoptosis) in representative patient-derived angioinvasive PTC cells (i.e. PTC7) harboring the BRAF^WT/V600E mutation and with MCL1 copy number gain/amplifications compared to vehicle-treated (DMSO, control) cells. Five μM vemurafenib and 100 nM obatoclax combined treatment significantly increased the percent of PTC cells in sub-G1 compared to vehicle-treated (DMSO, control) cells. These data represent the average ± standard deviation (error bars) of 2 independent experiments replicate measurements (*p < 0.05, **p < 0.01). B. Arrows highlight change of cell shape (rounded up and detached) in angioinvasive BRAF^WT/V600E-PTC cells (i.e. PTC7) with MCL1 copy number gain/amplifications treated for about 48 hours with vemurafenib (10 μM), obatoclax (100 nM), or vemurafenib (5 μM) + obatoclax (100 nM) compared to vehicle-treated PTC cells (DMSO, control). All scale bars are=400 μ. C. Flow cytometry analysis of non-synchronized cells normalized to the number of events in each condition at 48 hours post-treatment shows that 10 μM vemurafenib or 100 nM obatoclax significantly increased sub-G1 in metastatic PTC patient-derived KTC1 cells harboring the BRAF^WT/V600E mutation and with a lower copy number of MCL1 and with P16 homozygous loss compared to vehicle-treated (DMSO, control) cells. Five μM vemurafenib and 100 nM obatoclax combined treatment strongly increased the percent of KTC1 cells in sub-G1 compared to vehicle-treated (DMSO, control) cells. These data represent the average ± standard deviation (error bars) of 2 independent experiments replicate measurements (*p < 0.05, **p < 0.01, ***p < 0.01). D. Arrows highlight change of cell shape (rounded up and detached) in metastatic BRAF^WT/V600E-KTC1 cells in the presence of vemurafenib (10 μM), obatoclax (100 nM), or vemurafenib (5 μM) + obatoclax (100 nM) compared to vehicle-treated PTC cells (DMSO, control). All scale bars are=400 μ.