Transcriptomic analysis of the effects of mTOR inhibitor everolimus, CDK8/19 inhibitor SNX631 and their combination in MDA-MB-468 xenografts

RNA-Seq analysis was carried out to investigate the transcriptomic changes associated with the development of in vivo everolimus resistance and the effects of CDK8/19 inhibition on the development of resistance in MDA-MB-468 TNBC xenografts. In one study (short-term), tumor samples were collected after 30-40 days of treatment with vehicle or everolimus, when the tumor growth was still suppressed by the mTOR inhibitor. In another study (long-term), tumor samples were collected after 60-153 days of treatment with vehicle, everolimus, CDK8/19 inhibitor SNX631 or their combination, when tumors treated with everolimus but not with the drug combination developed resistance. Overall design: MDA-MB-468 cells (2x10^6 cells in 0.1 mL 50% Matrigel per animal) were inoculated in female NSG mice orthotopically under fat pad. Once tumors reached ~80 mm^3, mice were randomized into groups with similar-size tumors for further treatments. For the short-term study, mice received vehicle (30 days) or 2 mg/kg everolimus (38 days) by daily oral gavage. For the long-term study, mice received vehicle (p.o., q.d., up to 78 days) or SNX631 (350 ppm in medicated food, up to 96 days) or 2 mg/kg everolimus (p.o., q.d., up to 148 days) or in everolimus-SNX631 combination (148 days) . At the endpoint for each tumor, tumors were dissected from euthanized animals using sterilized surgical tools. Pieces of non-necrotic tumor tissues 50~100mm^3 were excised at the margin near midline section for RNA extraction. At least seven biological replicates (xenograft tumors from different animals) per study group were analyzed.