| Novel tropane alkaloids obtained from extracts of Erythroxylum pervillei Baillon and Erythroxylum rotundifolium Lanan (Erythroxylaceae) by means of bioassay-directed fraction were found to restore vinblastine sensitivity with cultured multidrug-resistant KB-V1 and CEM/VLB100 cells (pervilleine A was most active). Treatment of KB-V1 cells with modulators enhanced the intracellular accumulation of fluorescence dye. ClogP and molar refractivity values did not correlate with MDR-reversing activity. The mechanisms of the response were evaluated with a number of model systems. First, pervilleine A did not significantly affect the transcription of MDR1, as revealed by reverse transcriptional-PCR-based analysis of MDR1 mRNA, nor levels of Pgp, as shown by western blots. ATP-dependent binding of [3H]vinblastine observed with isolated KBVI cell membrane vesicles was inhibited by pervilleine A in a dose-dependent manner, and kinetic analysis indicted competitive inhibition with respect to vinblastine binding. Consistent with this effect, intracellular accumulation of [ 3H]vinblastine was increased significantly by pervilleine A. Visualization by confocal microscopy confirmed the intracellular accumulation of rhodamine 123 in drug resistant KB-V1 cells was significantly increased by pervilleine A. To explore the potential relevance of these responses, KB-V1 or KB-8-5 cells were placed in hollow fibers and implanted into NCr nu/ nu mice. Cell growth was significantly inhibited when vinblastine or pervilleine A were administered in combination. These data suggest that pervilleine A is an effective inhibitor of Pgp and should be further evaluated for clinical utility.; The hollow fiber test has been developed for the preliminary in vivo assessment of cancer chemotherapeutic efficacy. Using this model, we have established growth conditions with a panel of cells implanted at the i.p. and s.c. sites of athymic mice. Dioscin, 13-methoxy-15-oxozoapatlin, brusatol, deguelin, K9325-90-1, K9325-109-1, and (S)-coriolic acid function as chemotherapeutic agents by significantly inhibiting growth of cells implanted in mice. On the other hand, some in vitro active compounds and extracts did not mediate significant responses with the hollow fiber model. These studies illustrate the usefulness of the hollow fiber model in natural product drug discovery programs. Additional resources may be directed toward the most promising leads. |
