POTENT INDUCTION OF HUMAN COLON-CANCER CELL UPTAKE OF CHEMOTHERAPEUTIC DRUGS BY N-MYRISTOYLATED PROTEIN-KINASE-C-ALPHA (PKC-ALPHA) PSEUDOSUBSTRATE PEPTIDES THROUGH A P-GLYCOPROTEIN-INDEPENDENT MECHANISM

Phorbol ester protein kinase C (PKC) activators and PKC isozyme over-e xpression have been shown to significantly reduce intracellular accumu lation of chemotherapeutic drugs, in association with the induction of multidrug resistance (MDR) in drug-sensitive cancer cells and enhance ment of drug resistance in MDR cancer cells, These observations consti tute solid evidence that PKC plays a significant role in the MDR pheno type of cancer cells. PKC-catalyzed phosphorylation of the drug-efflux pump P-glycoprotein was recently ruled out as a contributing factor i n MDR. At present, the sole drug transport-related event that has been identified as a component of the role of PKC in MDR is PKC-induced ex pression of the P-glycoprotein-encoding gene mdr1. The objective of th is study was to test the hypothesis that PKC can modulate the uptake o f chemotherapeutic drugs in cancer cells independently of P-glycoprote in. We analyzed the effects of selective PKC activators/inhibitors on the uptake of radiolabelled cytotoxic drugs by cultured human colon ca ncer cells that lacked P-glycoprotein activity and did not express the drug efflux pump at the level of message (mdr1) or protein. We found that the selective PKC activator 12-O-tetradecanoylphorbol-13-acetate (TPA) significantly reduced uptake of [C-14] Adriamycin and [H-3] vinc ristine in human colon cancer cells devoid of P-glycoprotein activity, and that PKC-inhibitory N-myristoylated PKC-alpha pseudosubstrate syn thetic peptides potently and selectively induced uptake of the cytotox ic drugs in the phorbol ester-treated and non-treated colon cancer cel ls. TPA treatment of the cells did not induce expression of either P-g lycoprotein or its message mdr1. In contrast with [C-14]Adriamycin and [H-3] vincristine uptake, [H-3] 5-fluorouracil uptake by the cells wa s unaffected by TPA and reduced by the PKC-inhibitory peptides. These results indicate that PKC activation can significantly reduce the upta ke of multiple cytotoxic drugs by cancer cells independently of P-glyc oprotein, and that N-myristoylated PKC-alpha pseudosubstrate peptides potently and selectively induce uptake of multiple cytotoxic drugs in cultured human colon cancer cells by a novel mechanism that does not i nvolve P-glycoprotein and may involve PKC isozyme inhibition. Thus, N- myristoylated PKC-alpha pseudosubstrate peptides may offer a basis for the development of agents that reverse intrinsic drug resistance in h uman colon cancer.