Paclitaxel resistance: molecular mechanisms and pharmacologic manipulation.

It has been approximately ten years since the Food and Drug Administration (FDA) approved paclitaxel for the treatment of platinum resistant epithelial ovarian carcinoma. Since the approval, the drug has found therapeutic applications in a variety of schedules and in a wide variety of epithelial malignancies. Its novel mechanism of action provided the hope that it would demonstrate anti-neoplastic activity in multidrug resistant tumor cells. Unfortunately, as with other chemotherapeutic drugs, resistance is commonly seen. Laboratory investigation has defined a wide variety of resistance mechanisms including overexpression of multidrug resistance (MDR-1) gene, molecular changes in the target molecule (betatubulin), changes in apoptotic regulatory and mitosis checkpoint proteins, and more recently changes in lipid composition and potentially the overexpression of interleukin 6 (IL-6). This review describes the in vitro molecular data that define and support the various mechanisms of resistance and critically evaluates the evidence for the participation of these mechanisms in clinically relevant paclitaxel resistance. This review also explores pharmacologic attempts to modulate paclitaxel resistance, principally through inhibition of the MDR-1 drug efflux pump. Future avenues for drug resistance research and its pharmacologic manipulation in the clinic are discussed.

Overexpression of human phosphoglycerate kinase 1 (PGK1) induces a multidrug resistance phenotype.

BACKGROUND: Multidrug resistance is a significant barrier to the development of successful cancer treatment. To identify genetic alterations that are directly involved in paclitaxel resistance, a functional cloning strategy was developed. MATERIALS AND METHODS: Using mRNA from paclitaxel resistant human ovarian cancer cell line SW626TR, a cDNA library was established in a pCMV-Script vector that permits expression of cDNA inserts in mammalian cells. Transfection of the pCMV-Script/SW626TR cDNA library into the paclitaxel-sensitive human osteogenic sarcoma cell line, U-20S, resulted in several paclitaxel-resistant clones. RESULTS: DNA sequencing of clone C16 demonstrates complete homology to human phosphoglycerate kinase 1 (PGK1). Retransfection of the PGK1 insert into U-20S confers a multidrug resistant phenotype, characterized by a 30-fold increase in paclitaxel resistance, and cross-resistance to vincristine; adriamycin and mitoxantrone, but not methotrexate or cisplatin. Enzymatic analysis of the PGK1 transfectants demonstrates an increase in PGK1 activity as compared to the parental cell line, U-20S. Northern and Western analysis of PGK1 transfectants reveals no change in MDR-1 expression compared with the parental cell line. In addition, co-culture of PGK1 transfectants with verapamil only partially reverses the multidrug resistant phenotype. Rhodamine 123 studies are also consistent with an MDR-1 independent mechanism of increased drug efflux. CONCLUSION: Together this data suggests that PGK1 can induce a multidrug resistant phenotype through an MDR-1 independent mechanism.

Overexpression of IL-6 but not IL-8 increases paclitaxel resistance of U-2OS human osteosarcoma cells.

The cytokines IL-6, initially recognized as a regulator of immune and inflammatory response and IL-8, a potential regulator of angiogenesis, also regulate the growth of many tumor cells. Human cancer cells selected for multidrug resistance to common chemotherapeutic agents demonstrate increased expression of IL-6 and IL-8. To determine whether IL-6 or IL-8 overexpression contributes directly to the drug resistant phenotype, IL-6 or IL-8 cDNA were introduced into the paclitaxel sensitive human osteosarcoma cell line U-2OS using the pIRESneo bicistronic expression vector. Interleukin-6 and IL-8 transfectants were selected for either high IL-6 or IL-8 secretion and evaluated in drug resistance assays. Two IL-6 and two IL-8 secreting clones express IL-6 or IL-8 levels of 10 ng/ml and 1 ng/ml in culture, while parental U-2OS and pIRESneo vector transfected control cells express IL-6 and IL-8 levels of 0.005 ng/ml and 0.1 ng/ml, respectively. MTT cytotoxicity with IL-6 transfected cells demonstrates a five-fold increase in resistance to paclitaxel and a four-fold increase in resistance to doxorubicin as compared to U-2OS. There are no changes in mitoxantrone or topotecan resistance in the IL-6 transfectants as compared to parental U-2OS. Northern analysis of IL-6 transfectants demonstrates that the resistant phenotype is not related to increased levels of MDR-1, MRP-1, or LRP. Western analysis also confirms that P-glycoprotein levels are not altered in IL-6 transfectants. Further supporting an MDR-1 independent mechanism of drug resistance, verapamil cannot reverse paclitaxel resistance in transfected cells, findings further supported by rhodamine 123 exclusion data. Treatment of IL-6 transfected cells with paclitaxel, compared with drug-sensitive parental U-2OS, shows U-2OS(IL-6) are significantly more resistant to apoptosis induced by paclitaxel and exhibit decreased proteolytic activation of caspase-3. In contrast U-2OS(IL-8) transfectants demonstrate no appreciable increase in paclitaxel resistance when compared with parental cells. In summary, while both IL-6 and IL-8 are overexpressed in paclitaxel resistant cell lines, only IL-6 has the potential to contribute directly to paclitaxel and doxorubicin resistance in U-2OS. This resistance is through a non-MDR-1 pathway.