Synergistic Inhibition of Kinase Pathways Overcomes Resistance of Colorectal Cancer Spheroids to Cyclic Targeted Therapies.

Cancer cells often adapt to single-agent treatments with chemotherapeutics. Activation of alternative survival pathways is a major mechanism of drug resistance. A potential approach to block this feedback signaling is using combination treatments of a pair of drugs, although toxicity has been a limiting factor. Preclinical tumor models to identify mechanisms of drug resistance and determine low but effective combination doses are critical to effectively suppress tumor growth with reduced toxicity to patients. Using our aqueous two-phase system microtechnology, we developed colorectal tumor spheroids in high-throughput and evaluated resistance of cancer cells to three mitogen-activated protein kinase inhibitors (MAPKi) in long-term cyclic treatments. Our quantitative analysis showed that the efficacy of MAPKi significantly reduced over time, leading to an increase in proliferation of HCT116 colorectal cancer cells and growth of spheroids. We established that resistance was due to feedback activation of PI3K/AKT/mTOR pathway. Using high-throughput, dose-dependent combinations of each MAPKi and a PI3K/mTOR inhibitor, we identified low-dose, synergistic combinations that blocked resistance to MAPKi and effectively suppressed the growth of colorectal tumor spheroids in long-term treatments. Our approach to study drug resistance offers the potential to determine high priority treatments to test in animal models.