Abstract
Multidrug-Resistant (MDR) cancers mitigate the action of chemotherapeutics through drug efflux that occurs via ABC (ATP-Binding Cassette) transporters, including P-glycoprotein 1 (P-gp or ABCB1/MDR1). Because Toll-Like Receptor (TLR) agonist immunotherapies elicit abscopal anti-tumoral effects by modulating the activity of bystander tumor infiltrating immune cells, they not only circumvent the neutralizing effects of drug efflux, but could also work in synergy with this process. However, the effect of drug resistance on TLR agonist efflux is largely unknown. We begin to address this by investigating P-gp mediated efflux of model TLR agonists in cancer cell lines before and after acquired drug resistance. First, we used functionalized liposomes to determine that imidazoquinoline TLR agonists Imiquimod, Resiquimod, and Gardiquimod are substrates for P-gp. Next, we created Doxorubicin-resistant cancer cell lines from B16 melanoma, TRAMP prostate, and 4T1 breast cancer and observed that each cell line increased P-gp expression in response to Doxorubicin. Comparing imidazoquinoline efflux in Doxorubicin-resistant cell lines, relative to parent cancer cell lines, we used P-gp competitive substrates and inhibitors to demonstrate that imidazoquinoline efflux occurs through P-gp and is enhanced as a consequence of acquired drug resistance. We found that the most hydrophobic, yet least potent imidazoquinoline (Imiquimod), was the best substrate for efflux. This suggests a new parameter, susceptibility to drug efflux, could be an important consideration in the rationale design of the next generation of TLR agonist immunotherapies that are targeted to cancer cells, yet effect their mechanisms of action by modulating the activity of tumor infiltrating immune cells.
Competing Interest Statement
AJB, AEN, and RJM are inventors on WSU non-provisional patent application 15/722,018; AEN and RJM are owners of Astante Therapeutics Inc. both of which use concepts related to those in this work.