Abstract
The ability of the multidrug resistance modifiers R- and R,S-verapamil (VPL), cyclosporine A (CsA) and its non-immunosuppressive derivative SDZ PSC 833 (PSC 833) to inhibit P-glycoprotein (P-gp)-mediated transepithelial flux of tritiated vinblastine was investigated using tight and highly resistant (R>1,400 Ω cm2) monolayer cultures of intestinal adenocarcinoma-derived HCT-8 cells grown on permeable tissue-culture inserts. Apical addition of these chemosensitisers inhibited drug flux (137 pmol h−1 cm−2; range, 133–142 pmol h−1 cm−2) in the basal to apical secretory direction at clinically relevant concentrations, with PSC 833 showing the highest activity, exhibiting inhibition at concentrations as low as 10 ng/ml (9 nM). Acidification of the modulator-containing apical compartment to an extracellular pH (pHo) of 6.8 had no influence on MDR reversal by CsA at 1 μg/ml (0.9 μM; flux inhibition, 52%) or by PSC 833 at 100 ng/ml (0.09 μM; flux inhibition, 60%), in contrast to R,S- and R-VPL, which showed decreased inhibition and caused less accumulation of vinblastine in HCT-8 cells under this condition (flux inhibition of 35% and 23%, respectively, at pHo 6.8 vs 50% and 43%, respectively, at pHo 7.5). P-gp-mediated rhdamine 123 efflux from dye-loaded single-cell suspensions of HCT-8 cells as measured by flow cytometry was not impeded at pHo 6.8 in comparison with pHo 7.5 in standard medium, but at low pHo the inhibitory activity of r-VPL (29% vs 60% rhodamine 123 efflux inhibition) was diminished significantly, again without a reduction in the effect of PSC 833 (rhodamine 123 flux inhibition, 75%). In conclusion, drug extrusion across polarised monolayers, which offer a relevant model for normal epithelia and tumour border areas, is inhibited by the apical presence of R,S- and R-VPL, CsA and PSC 833 at similar concentrations described for single-cell suspensions, resulting in increased (2.2- to 3.7-fold) intracellular drug accumulation. Functional apical P-gp expression, the absence of paracellular leakage and modulator-sensitive rhodamine 123 efflux in single HCT-8 cells indicate a P-gp-mediated transcellular efflux in HCT-8 monolayers. In addition to its high MDR-reversing capacity, the inhibitory activity of PSC 833 is not affected by acidic extracellular conditions, which reduce the VPL-induced drug retention significantly. As far as MDR contributes to the overall cellular drug resistance of solid tumours with hypoxic and acidic microenvironments, PSC 833 holds the greatest promise for clinical reversal of unresponsiveness to the respective group of chemotherapeutics.
Article PDF
Similar content being viewed by others
Avoid common mistakes on your manuscript.
Abbreviations
- CsA:
-
cyclosporine A
- HEPES:
-
[N-(2-hydroxyethyl)piperazine-N′-(2-ethanesulfonic acid)]
- MDR:
-
multidrug resistance; PBS, phosphate-buffered saline
- P-gp:
-
P-glycoprotein
- pHo:
-
extracellular pH
- PSC 833:
-
SDZ PSC 833
- R:
-
resistance
- TEM:
-
transmission electron microscopy
- VPL:
-
verapamil
References
Bissett D, Kerr DJ, Cassidy J, Meredith P, Traugott U, Kaye SB (1991) Phase I and pharmacokinetic study ofd-verapamil and doxorubicin. Br J Cancer 64: 1168
Cordon-Cardo C, O'Brien JP, Boccia J, Casals D, Bertino JR, Melamed MR (1990) Expression of the multidrug resistance gene product (P-glycoprotein) in human normal and tumor tissues. J Histochem Cytochem 38: 1277
Ford JM, Hait WN (1990) Pharmacology of drugs that alter multidrug resistance in cancer. Pharmacol Rev 42: 155
Gaveriaux C, Boesch D, Jachez B, Bollinger P, Payne T, Loor F (1991) SDZ-PSC-833, a non-immunosuppressive cyclosporine analog, is a very potent multidrug-resistance modifier. J Cell Pharmacol 2: 225
Georges E, Bradley G, Gariepy J, Ling V (1990) Detection of P-glycoprotein isoforms by gene-specific monoclonal antibodies. Proc Natl Acad Sci USA 87: 152
Gottesman MM, Pastan I (1989) Clinical trials of agents that reverse multi-drug resistance. J Clin Oncol 7: 409
Gruber A, Peterson C, Reizenstein P (1988)d-Verapamil andl-verapamil are equally effective in increasing vincristine accumulation in leukemic cells in vitro. Int J Cancer 41: 224
Hamilton G, Cosentini EP, Teleky B, Koperna T, Zacherl J, Schiessel R, Wenzl E (1993) Chemosensitization effect of verapamil and cyclosporin A in vitro is reduced under acidic pH conditions. Eur J Cancer 29A: 1635
Hidalgo IJ, Raub RJ, Borchardt RT (1989) Characterisation of the human colon carcinoma cell line (Caco-2) as a model system for intestinal epithelial permeability. Gastroenterology 96: 736
Hunter J, Hirst BH, Simmons NL (1991) Epithelial secretion of vinblastine by human intestinal adenocareinoma cell (HCT-8 and T84) layers expressing P-glycoprotein. Br J Cancer 64: 437
Hunter J, Hirst BH, Simmons NL (1993) Drug absorption limited by P-glycoprotein-mediated secretory drug transport in human intestinal epithelial Caco-2 cell layers. Pharm Res 10: 743
Hunter J, Jepson MA, Tsuruo T, Simmons NL, Hirst BH (1993) Functional expression of P-glycoprotein in apical membranes of human intestinal Caco-2 cells. J Biol Chem 268: 1499
Inoue M, Kinne R, Tran T, Biempica L, Arias IM (1983) Rat liver canalicular membrane vesicles. J Biol Chem 258: 5183
Kaye SB (1993) P glycoprotein (P-gp) and drug resistance—time for reappraisal? Br J Cancer 67: 641
Kessel D, Beck WT, Kukuruga D, Schulz V (1991) Characterization of multidrug resistance by fluorescent dyes. Cancer Res 51: 4665
Klohs WD, Steinkampf RW (1988) Possible link between the intrinsic drug resistance of colon tumours and a detoxification mechanism of intestinal cells. Cancer Res 48: 3025
Madara JI, Dharmsathaphorn K (1985) Occluding junction structure-function relationships in a cultured epithelial monolayer. J Cell Biol 101:2124
Moscow JA, Fairchild CR, Madden MJ, Ransom DT, Wieand HS, O'Brien EE, Poplack DG, Cossman J, Myers CE, Cowan KH (1989) Expression of anionic glutathione-S-transferase and P-glycoprotein genes in human tissues and tumors. Cancer Res 49: 1422
Pastan I, Gottesman MM (1991) Multidrug resistance. Annu Rev Med 42: 277
Song CW, Lyons JC, Luo Y (1993) Intra- and extracellular pH in solid tumors: influence on therapeutic response. In: Teicher BA (ed) Drug resistance in oncology. Marcel Dekker, New York Basel Hong Kong, p 25
Tannock IF, Rotin D (1989) Acid pH in tumors and its potential for therapeutic exploitation. Cancer Res 52: 4373
Thiebaut F, Tsuruo T, Hamada H, Gottesman MM, Pastan I, Willingham MC (1987) Cellular localization of the multidrug-resistance gene product p-glycoprotein in normal human tissues. Proc Natl Acad Sci USA 84: 7735
Tompkins WAF, Watrach AM, Schmale JD, Schultz RM, Harris JA (1974) Cultural and antigenic properties of newly established cell strains derived from adenocarcinomas of the human colon and rectum. J Natl Cancer Inst 52: 1101
Tsuruo T, Iida H, Tsukagoshi S, Sakurai Y (1989) Overcoming of vincristine resistance in P388 leukemia in vivo and in vitro through enhanced cytotoxicity of vincristine and vinblastine by verapamil. Cancer Res 41: 1967
Twentyman PR (1988) Modification of cytotoxic drug resistance by non-immunosuppressive cyclosporins. Br J Cancer 57: 254
Vendrik CPJ, Bergers JJ, De Jong WH, Steerenberg PA (1992) Resistance to cytostatic drugs at the cellular level. Cancer Chemother Pharmacol 29: 413
Volm M, Mattern J, Efferth T, Pommerenke EW (1992) Expression of several resistance mechanisms in untreated human kidney and lung carcinomas. Anticancer Res 12: 1063
Yusa K, Tsuruo T (1989) Reversal mechanism of multidrug resistance by verapamil: direct binding of verapamil to P-glycoprotein on specific sites and transport of verapamil outward across the plasma membrane of K562/ADM cells. Cancer Res 49: 5002
Author information
Authors and Affiliations
Rights and permissions
About this article
Cite this article
Zacherl, J., Hamilton, G., Thalhammer, T. et al. Inhibition of P-glycoprotein-mediated vinblastine transport across HCT-8 intestinal carcinoma monolayers by verapamil, cyclosporine A and SDZ PSC 833 in dependence on extracellular pH. Cancer Chemother. Pharmacol. 34, 125–132 (1994). https://doi.org/10.1007/BF00685929
Received:
Accepted:
Issue Date:
DOI: https://doi.org/10.1007/BF00685929