Skip to main content
PMC home page
British Journal of Cancer logoLink to British Journal of Cancer
. 1996 Mar;73(5):596–602. doi: 10.1038/bjc.1996.103

Low-level doxorubicin resistance in P-glycoprotein-negative human pancreatic tumour PSN1/ADR cells implicates a brefeldin A-sensitive mechanism of drug extrusion.

V N Verovski 1, D L Van den Berge 1, M M Delvaeye 1, R J Scheper 1, W J De Neve 1, G A Storme 1
PMCID: PMC2074337  PMID: 8605092

Abstract

The human pancreatic tumour cell line PSN1/ADR, stepwise selected in 17-510 nM doxorubicin, displayed a multidrug resistance not conferred by P-glycoprotein (P-gp). Resistance to 17-51 nM doxorubicin was accompanied by overexpression of the vesicular marker lung resistance-related protein (LRP). Further selection in 170 nM doxorubicin led to the activation of multidrug resistance-associated protein (MRP) and to the development of drug accumulation/retention defects sensitive to verapamil. In addition, these defects were reversible by the vesicular traffic inhibitors brefeldin A, fluoroaluminate and nocodazole. In contrast, in human ovarian H134AD cells that are resistant to 1700 nM doxorubicin and used as P-gp-positive controls, the drug efflux was inhibited only by verapamil. The tyrosine kinase inhibitor genistein was a potent blocker of doxorubicin efflux in the PSN1/ADR cells but showed no activity in the H134 AD cells. The doxorubicin cytotoxicity in the PSN1/ADR cells was enhanced both by verapamil and brefeldin A, whereas in the parental PSN1 cells they demonstrated the opposite effects, being respectively sensitising and protecting. The P-gp-negative PSN1/ADR cells adapted to 510 nM doxorubicin retained brefeldin A-sensitive doxorubicin accumulation defects while MRP declined. The persistence of brefeldin A-responsive phenotype on the background of variable MRP expression suggests this agent as a useful functional probe for non-P-gp-mediated resistance to plasma-achievable doxorubicin concentrations.

Full text

PDF
596

Images in this article

598Figure 2
on p.598

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Barr F. A., Leyte A., Mollner S., Pfeuffer T., Tooze S. A., Huttner W. B. Trimeric G-proteins of the trans-Golgi network are involved in the formation of constitutive secretory vesicles and immature secretory granules. FEBS Lett. 1991 Dec 9;294(3):239–243. doi: 10.1016/0014-5793(91)81438-e. [DOI] [PubMed] [Google Scholar]
  2. Barrand M. A., Rhodes T., Center M. S., Twentyman P. R. Chemosensitisation and drug accumulation effects of cyclosporin A, PSC-833 and verapamil in human MDR large cell lung cancer cells expressing a 190k membrane protein distinct from P-glycoprotein. Eur J Cancer. 1993;29A(3):408–415. doi: 10.1016/0959-8049(93)90397-x. [DOI] [PubMed] [Google Scholar]
  3. Beck W. T. The cell biology of multiple drug resistance. Biochem Pharmacol. 1987 Sep 15;36(18):2879–2887. doi: 10.1016/0006-2952(87)90198-5. [DOI] [PubMed] [Google Scholar]
  4. Breitfeld P. P., McKinnon W. C., Mostov K. E. Effect of nocodazole on vesicular traffic to the apical and basolateral surfaces of polarized MDCK cells. J Cell Biol. 1990 Dec;111(6 Pt 1):2365–2373. doi: 10.1083/jcb.111.6.2365. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Cole S. P., Bhardwaj G., Gerlach J. H., Mackie J. E., Grant C. E., Almquist K. C., Stewart A. J., Kurz E. U., Duncan A. M., Deeley R. G. Overexpression of a transporter gene in a multidrug-resistant human lung cancer cell line. Science. 1992 Dec 4;258(5088):1650–1654. doi: 10.1126/science.1360704. [DOI] [PubMed] [Google Scholar]
  6. Cole S. P., Sparks K. E., Fraser K., Loe D. W., Grant C. E., Wilson G. M., Deeley R. G. Pharmacological characterization of multidrug resistant MRP-transfected human tumor cells. Cancer Res. 1994 Nov 15;54(22):5902–5910. [PubMed] [Google Scholar]
  7. Coley H. M., Amos W. B., Twentyman P. R., Workman P. Examination by laser scanning confocal fluorescence imaging microscopy of the subcellular localisation of anthracyclines in parent and multidrug resistant cell lines. Br J Cancer. 1993 Jun;67(6):1316–1323. doi: 10.1038/bjc.1993.244. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Delvaeye M., Verovski V., De Neve W., Storme G. DNA breakage, cytotoxicity, drug accumulation and retention in two human ovarian tumor cell lines AZ224 and AZ364 treated with adriamycin, modulated by verapamil. Anticancer Res. 1993 Sep-Oct;13(5A):1533–1538. [PubMed] [Google Scholar]
  9. Donaldson J. G., Finazzi D., Klausner R. D. Brefeldin A inhibits Golgi membrane-catalysed exchange of guanine nucleotide onto ARF protein. Nature. 1992 Nov 26;360(6402):350–352. doi: 10.1038/360350a0. [DOI] [PubMed] [Google Scholar]
  10. Duan R. D., Wagner A. C., Yule D. I., Williams J. A. Multiple inhibitory effects of genistein on stimulus-secretion coupling in rat pancreatic acini. Am J Physiol. 1994 Feb;266(2 Pt 1):G303–G310. doi: 10.1152/ajpgi.1994.266.2.G303. [DOI] [PubMed] [Google Scholar]
  11. Eijdems E. W., Zaman G. J., de Haas M., Versantvoort C. H., Flens M. J., Scheper R. J., Kamst E., Borst P., Baas F. Altered MRP is associated with multidrug resistance and reduced drug accumulation in human SW-1573 cells. Br J Cancer. 1995 Aug;72(2):298–306. doi: 10.1038/bjc.1995.328. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Flens M. J., Izquierdo M. A., Scheffer G. L., Fritz J. M., Meijer C. J., Scheper R. J., Zaman G. J. Immunochemical detection of the multidrug resistance-associated protein MRP in human multidrug-resistant tumor cells by monoclonal antibodies. Cancer Res. 1994 Sep 1;54(17):4557–4563. [PubMed] [Google Scholar]
  13. Gervasoni J. E., Jr, Fields S. Z., Krishna S., Baker M. A., Rosado M., Thuraisamy K., Hindenburg A. A., Taub R. N. Subcellular distribution of daunorubicin in P-glycoprotein-positive and -negative drug-resistant cell lines using laser-assisted confocal microscopy. Cancer Res. 1991 Sep 15;51(18):4955–4963. [PubMed] [Google Scholar]
  14. Gottesman M. M., Pastan I. Biochemistry of multidrug resistance mediated by the multidrug transporter. Annu Rev Biochem. 1993;62:385–427. doi: 10.1146/annurev.bi.62.070193.002125. [DOI] [PubMed] [Google Scholar]
  15. Klausner R. D., Donaldson J. G., Lippincott-Schwartz J. Brefeldin A: insights into the control of membrane traffic and organelle structure. J Cell Biol. 1992 Mar;116(5):1071–1080. doi: 10.1083/jcb.116.5.1071. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Ktistakis N. T., Linder M. E., Roth M. G. Action of brefeldin A blocked by activation of a pertussis-toxin-sensitive G protein. Nature. 1992 Mar 26;356(6367):344–346. doi: 10.1038/356344a0. [DOI] [PubMed] [Google Scholar]
  17. Marquardt D., Center M. S. Drug transport mechanisms in HL60 cells isolated for resistance to adriamycin: evidence for nuclear drug accumulation and redistribution in resistant cells. Cancer Res. 1992 Jun 1;52(11):3157–3163. [PubMed] [Google Scholar]
  18. McGrath T., Latoud C., Arnold S. T., Safa A. R., Felsted R. L., Center M. S. Mechanisms of multidrug resistance in HL60 cells. Analysis of resistance associated membrane proteins and levels of mdr gene expression. Biochem Pharmacol. 1989 Oct 15;38(20):3611–3619. doi: 10.1016/0006-2952(89)90134-2. [DOI] [PubMed] [Google Scholar]
  19. Rhodes T., Barrand M. A., Twentyman P. R. Modification by brefeldin A, bafilomycin A1 and 7-chloro-4-nitrobenz-2-oxa-1,3-diazole (NBD) of cellular accumulation and intracellular distribution of anthracyclines in the non-P-glycoprotein-mediated multidrug-resistant cell line COR-L23/R. Br J Cancer. 1994 Jul;70(1):60–66. doi: 10.1038/bjc.1994.250. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Scheper R. J., Broxterman H. J., Scheffer G. L., Kaaijk P., Dalton W. S., van Heijningen T. H., van Kalken C. K., Slovak M. L., de Vries E. G., van der Valk P. Overexpression of a M(r) 110,000 vesicular protein in non-P-glycoprotein-mediated multidrug resistance. Cancer Res. 1993 Apr 1;53(7):1475–1479. [PubMed] [Google Scholar]
  21. Scheper R. J., Bulte J. W., Brakkee J. G., Quak J. J., van der Schoot E., Balm A. J., Meijer C. J., Broxterman H. J., Kuiper C. M., Lankelma J. Monoclonal antibody JSB-1 detects a highly conserved epitope on the P-glycoprotein associated with multi-drug-resistance. Int J Cancer. 1988 Sep 15;42(3):389–394. doi: 10.1002/ijc.2910420314. [DOI] [PubMed] [Google Scholar]
  22. Sehested M., Skovsgaard T., van Deurs B., Winther-Nielsen H. Increased plasma membrane traffic in daunorubicin resistant P388 leukaemic cells. Effect of daunorubicin and verapamil. Br J Cancer. 1987 Dec;56(6):747–751. doi: 10.1038/bjc.1987.282. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Supino R., Binaschi M., Capranico G., Gambetta R. A., Prosperi E., Sala E., Zunino F. A study of cross-resistance pattern and expression of molecular markers of multidrug resistance in a human small-cell lung-cancer cell line selected with doxorubicin. Int J Cancer. 1993 May 8;54(2):309–314. doi: 10.1002/ijc.2910540224. [DOI] [PubMed] [Google Scholar]
  24. Takeda Y., Nishio K., Niitani H., Saijo N. Reversal of multidrug resistance by tyrosine-kinase inhibitors in a non-P-glycoprotein-mediated multidrug-resistant cell line. Int J Cancer. 1994 Apr 15;57(2):229–239. doi: 10.1002/ijc.2910570217. [DOI] [PubMed] [Google Scholar]
  25. Twentyman P. R., Rhodes T., Rayner S. A comparison of rhodamine 123 accumulation and efflux in cells with P-glycoprotein-mediated and MRP-associated multidrug resistance phenotypes. Eur J Cancer. 1994;30A(9):1360–1369. doi: 10.1016/0959-8049(94)90187-2. [DOI] [PubMed] [Google Scholar]
  26. Versantvoort C. H., Broxterman H. J., Lankelma J., Feller N., Pinedo H. M. Competitive inhibition by genistein and ATP dependence of daunorubicin transport in intact MRP overexpressing human small cell lung cancer cells. Biochem Pharmacol. 1994 Sep 15;48(6):1129–1136. doi: 10.1016/0006-2952(94)90149-x. [DOI] [PubMed] [Google Scholar]
  27. Versantvoort C. H., Schuurhuis G. J., Pinedo H. M., Eekman C. A., Kuiper C. M., Lankelma J., Broxterman H. J. Genistein modulates the decreased drug accumulation in non-P-glycoprotein mediated multidrug resistant tumour cells. Br J Cancer. 1993 Nov;68(5):939–946. doi: 10.1038/bjc.1993.458. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Vichi P. J., Tritton T. R. Protection from adriamycin cytotoxicity in L1210 cells by brefeldin A. Cancer Res. 1993 Nov 1;53(21):5237–5243. [PubMed] [Google Scholar]

Articles from British Journal of Cancer are provided here courtesy of Cancer Research UK

RESOURCES