Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:3.6.3.44 (P-glycoprotein)
 13,344 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

In previous work (Weisburg, J. H., Curcio, M., Caron, P. C., Raghi, G., Mechetner, E. B., Roepe, P. D., and Scheinberg, D. A. (1996) J. Exp. Med. 183, 2699-2704), we showed that multidrug resistance (MDR) cells created by continuous selection with the vinca alkaloid vincristine (HL60 RV+) or by retroviral infection (K562/human MDR 1 cells) exhibited significant resistance to complement-mediated cytotoxicity (CMC). This resistance was due to the presence of overexpressed P-glycoprotein (P-GP). In this paper, we probe the molecular mechanism of this phenomenon. We test whether the significant elevated intracellular pH (pHi) that accompanies P-GP overexpression is sufficient to confer resistance to CMC and whether this resistance is related to effects on complement function in the cell membrane. Control HL60 cells not expressing P-GP, but comparably elevated in cytosolic pHi by two independent methods (CO2 "conditioning" or isotonic Cl- substitution), are tested for CMC using two different antibody-antigen systems (human IgG and murine IgM; protein and carbohydrate) and two complement sources (rabbit and human). Elevation of pHi by either of these methods or by expression of P-GP confers resistance to CMC. Resistance is not observed when the alkalinization mediated by reverse Cl-/HCO3- exchange upon Cl- substitution is blocked by treatment with dihydro-4,4'-diisothiocyanostilbene-2,2'-disulfonate. Continuous photometric monitoring of 2',7'-bis(carboxyethyl)-5, 6-carboxyfluorescein (BCECF), to assess changes in pHi or efflux of the probe through MAC pores, in single cells or cell populations, respectively, verifies changes in pHi upon CO2 conditioning and Cl- substitution and release of BCECF upon formation of MAC pores. Antibody binding and internalization kinetics are similar in both the parental and resistant cell lines as measured by radioimmunoassay, but flow cytometric data showed that net complement deposition in the cell membrane is both delayed and reduced in magnitude in the MDR cells and in the cells with increased pHi. This interpretation is supported by comparison of BCECF release data for the different cells. Dual isotopic labeling of key complement components shows no significant change in molecular stoichiometry of the MACs formed at different pHi. The results are relevant to understanding clinical implications of MDR, the physiology of P-GP, and the biochemistry of the complement cascade and further suggest that the "drug pump" model of P-GP action cannot account for all of its effects.
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PMID:Intracellular pH and multidrug resistance regulate complement-mediated cytotoxicity of nucleated human cells. 1019 65

The taxanes, paclitaxel (PTX) and docetaxel (DTX), belong to a novel class of anticancer drugs that stabilize microtubules and lead to tumor cell death. While both agents are widely used for the treatment of lung, breast, and ovarian cancer, many tumor types are refractory or develop resistance to these drugs. We describe here a novel analogue of DTX, designated MAC-321 [Microtubule/Apoptosis/Cytotoxic: 5beta, 20-epoxy-1, 2alpha-, 4-, 7beta-, 10beta-, 13alpha-hexahydroxytax-11-en-9-one 4 acetate 2 benzoate 7-propionate 13-ester with (2R,3S)-N-tertbutoxycarbonyl-3-(2-furyl)isoserine], that overcomes P-glycoprotein-mediated resistance to PTX and DTX in preclinical model systems. Similar to PTX or DTX, MAC-321 enhanced the rate of tubulin polymerization in vitro and caused the bundling of microtubules in cells. MAC-321 inhibited proliferation of a panel of 14 tumor cell lines with minimal variation in potency (IC(50) = 2.2 +/- 1.4 nM; range = 0.6-5.3 nM). Unlike PTX or DTX, the IC(50) of MAC-321 did not vary in cells that expressed low to moderate levels of P-glycoprotein. Even under extraordinary conditions in KB-V1 cells, which highly overexpress P-glycoprotein, resistance to MAC-321 was 80-fold compared with that of PTX (1400-fold) and DTX (670-fold). In addition, equivalent or less resistance to MAC-321 compared with PTX or DTX was observed in four cell lines that contain distinct point mutations within the taxane-binding site of beta-tubulin. Most importantly, MAC-321 displayed superior in vivo efficacy because: (a) MAC-321 either partially or completely inhibited tumor growth in three tumor models that overexpressed P-glycoprotein and were resistant to PTX; and (b) unlike PTX or DTX, MAC-321 was highly effective when given orally. MAC-321 was also highly effective when given as single i.v. dose. Our findings suggest that MAC-321, which is currently under clinical evaluation, may have broad therapeutic value.
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PMID:MAC-321, a novel taxane with greater efficacy than paclitaxel and docetaxel in vitro and in vivo. 1455 6

Resistance to paclitaxel-based therapy is frequently encountered in the clinic. The mechanisms of intrinsic or acquired paclitaxel resistance are not well understood. We sought to characterize the resistance mechanisms that develop upon chronic exposure of a cancer cell line to paclitaxel in the presence of the P-glycoprotein reversal agent, CL-347099. The epidermoid tumor line KB-3-1 was exposed to increasing concentrations of paclitaxel and 5 micromol/L CL-347099 for up to 1 year. Cells grown in 15 nmol/L paclitaxel plus CL-347099 (KB-15-PTX/099) developed 18-fold resistance to paclitaxel and were dependent upon paclitaxel for maximal growth. They grew well and retained resistance to paclitaxel when grown in athymic mice. Cross-resistance (3- to 5-fold) was observed in tissue culture to docetaxel, the novel taxane MAC-321, and epothilone B. Collateral sensitivity (approximately 3-fold) was observed to the depolymerizing agents vinblastine, dolastatin-10, and HTI-286. KB-15-PTX/099-resistant cells did not overexpress P-glycoprotein nor did they have an alteration of [14C]paclitaxel accumulation compared with parental cells. However, a novel point mutation (T to A) resulting in Asp26 to glutamate substitution in class I (M40) beta-tubulin was found. Based on an electron crystallography structure of Zn-stabilized tubulin sheets, the phenyl ring of C-3' NHCO-C6H5 of paclitaxel makes contact with Asp26 of beta-tubulin, suggesting a ligand-induced mutation. Optimized model complexes of paclitaxel, docetaxel, and MAC-321 in beta-tubulin show a novel hydrogen bonding pattern for the glutamate mutant and rationalize the observed resistance profiles. However, a mutation in the paclitaxel binding pocket does not explain the phenotype completely. KB-15-PTX/099 cells have impaired microtubule stability as determined by a reduced percentage of tubulin in microtubules and reflected by less acetylated tubulin. These results suggest that a mutation in tubulin might affect microtubule stability as well as drug binding and contribute to the observed resistance profile.
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PMID:Paclitaxel-resistant cells have a mutation in the paclitaxel-binding region of beta-tubulin (Asp26Glu) and less stable microtubules. 1650