This Article Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Dumontet, C. Articles by Sikic, B. I. Search for Related Content PubMed PubMed Citation Articles by Dumontet, C. Articles by Sikic, B. I.

Resistance Mechanisms in Human Sarcoma Mutants Derived by Single-Step Exposure to Paclitaxel (Taxol)¹

Divisions of Oncology and Clinical Pharmacology, Department of Medicine, Stanford University School of Medicine, Stanford, California 94305-5306

A fluctuation analysis experiment was performed by exposing 15 expanded populations of MES-SA sarcoma cells to paclitaxel (Taxol) at a concentration of 10 nM for 7 days. The mutation rate was approximately 8 x 10^-7/cell generation. ANOVA supports a stochastic cell survival mechanism of spontaneous mutation rather than induction of an adaptive response under these selection conditions. Surviving colonies were found in 12 populations, 9 of which had clones that remained resistant to paclitaxel after a 2-month period of propagation. Analysis of mdr1 gene expression by reverse transcription PCR demonstrated positive clones in 4 of the 9 populations with stable resistance. Accumulation of [³H]paclitaxel was decreased in these clones but not in the mdr1-negative clones compared with parental cells. A high degree of resistance to paclitaxel (36-to 93-fold) was selected by this single drug exposure in all 9 stably resistant mutants. Those with mdr1 activation demonstrated a broad cross-resistance to vinblastine, doxorubicin, and etoposide, whereas the other 6 mutants were cross-resistant only to the Vinca alkaloids. Because tubulins are the target molecules for paclitaxel cytotoxicity, we evaluated total tubulin content by immunoblotting and performed semiquantitative reverse transcription PCR analysis for expression of the -tubulin isotypes B1, K1 and H44, the ß-tubulin isotypes M40, ß9, 5ß, ß2 and ß4, and -tubulin. Total tubulin content was decreased significantly in one of the single-step mutants. All surviving clones, both resistant and sensitive to paclitaxel, displayed reduced expression of the 5ß and the ß4 ß-tubulin isotype transcripts in comparison with the parental cell line. These data suggest that stringent exposure to paclitaxel selected clones with reduced transcript levels of 5ß and ß4 ß-tubulin isotypes, but that these reduced levels were not directly involved in the resistance of the clones to paclitaxel. The results suggest an important role for non-multidrug-resistant mechanisms of resistance to paclitaxel. These mechanisms do not involve reduced drug accumulation and provide cross-resistance among both paclitaxel and tubulin depolymerizing agents.

¹ This work was supported by American Cancer Society Grant DHP-76, Department of the Army Grant DAMD 17-94-J-4352, NIH Grant CA-52168 (B. I. S.), the Association pour la Recherche contre le Cancer, the Ligue Nationale Contre le Cancer, the Ligue contre le Cancer de la Drôme, and the Philippe Foundation, Inc. (C. D.).

² To whom requests for reprints should be addressed, at Room M-211, Stanford University School of Medicine, Department of Medicine, Divisions of Oncology and Clinical Pharmacology, Stanford, CA 94305-5306. Phone: 415-725-6427; Fax: 415-725-1420.

Received 10/13/95. Accepted 1/ 2/96.

This article has been cited by other articles:

				C. Papadaki, D. Mavroudis, M. Trypaki, A. Koutsopoulos, E. Stathopoulos, D. Hatzidaki, E. Tsakalaki, V. Georgoulias, and J. Souglakos Tumoral Expression of TXR1 and TSP1 Predicts Overall Survival of Patients with Lung Adenocarcinoma Treated with First-line Docetaxel-Gemcitabine Regimen Clin. Cancer Res., June 1, 2009; 15(11): 3827 - 3833. [Abstract] [Full Text] [PDF]

				A.-M. Liberatore, H. Coulomb, D. Pons, O. Dutruel, P. G. Kasprzyk, M. Carlson, A. S. Nelson, S. P. Newman, C. Stengel, P. Auvray, et al. IRC-083927 is a new tubulin binder that inhibits growth of human tumor cells resistant to standard tubulin-binding agents Mol. Cancer Ther., August 1, 2008; 7(8): 2426 - 2434. [Abstract] [Full Text] [PDF]

				M. Hari, F. Loganzo, T. Annable, X. Tan, S. Musto, D. B. Morilla, J. H. Nettles, J. P. Snyder, and L. M. Greenberger Paclitaxel-resistant cells have a mutation in the paclitaxel-binding region of {beta}-tubulin (Asp26Glu) and less stable microtubules. Mol. Cancer Ther., February 1, 2006; 5(2): 270 - 278. [Abstract] [Full Text] [PDF]

				M. N. Fishman, C. R. Garrett, G. R. Simon, A. A. Chiappori, R. M. Lush, W. R. Dinwoodie, J. J. Mahany, A. M. Dellaportas, A. Cantor, A. Gollerki, et al. Phase I Study of the Taxane BMS-188797 in Combination with Carboplatin Administered Every 3 Weeks in Patients with Solid Malignancies Clin. Cancer Res., January 15, 2006; 12(2): 523 - 528. [Abstract] [Full Text] [PDF]

				L. Mickley Huff, Z. Wang, A. Iglesias, T. Fojo, and J.-S. Lee Aberrant Transcription from an Unrelated Promoter Can Result in MDR-1 Expression following Drug Selection In vitro and in Relapsed Lymphoma Samples Cancer Res., December 15, 2005; 65(24): 11694 - 11703. [Abstract] [Full Text] [PDF]

				K. G. Chen, Y. C. Wang, M. E. Schaner, B. Francisco, G. E. Duran, D. Juric, L. M. Huff, H. Padilla-Nash, T. Ried, T. Fojo, et al. Genetic and Epigenetic Modeling of the Origins of Multidrug-Resistant Cells in a Human Sarcoma Cell Line Cancer Res., October 15, 2005; 65(20): 9388 - 9397. [Abstract] [Full Text] [PDF]

				Z. Duan, Y. Duan, D. E. Lamendola, R. Z. Yusuf, R. Naeem, R. T. Penson, and M. V. Seiden Overexpression of MAGE/GAGE Genes in Paclitaxel/Doxorubicin-resistant Human Cancer Cell Lines Clin. Cancer Res., July 1, 2003; 9(7): 2778 - 2785. [Abstract] [Full Text] [PDF]

				L. A. Martello, P. Verdier-Pinard, H.-J. Shen, L. He, K. Torres, G. A. Orr, and S. B. Horwitz Elevated Levels of Microtubule Destabilizing Factors in a Taxol-resistant/dependent A549 Cell Line with an {alpha}-Tubulin Mutation Cancer Res., March 15, 2003; 63(6): 1207 - 1213. [Abstract] [Full Text] [PDF]

				S. H. Jang, M. G. Wientjes, and J. L.-S. Au Interdependent Effect of P-Glycoprotein-Mediated Drug Efflux and Intracellular Drug Binding on Intracellular Paclitaxel Pharmacokinetics: Application of Computational Modeling J. Pharmacol. Exp. Ther., February 1, 2003; 304(2): 773 - 780. [Abstract] [Full Text] [PDF]

				D. Toppmeyer, A. D. Seidman, M. Pollak, C. Russell, K. Tkaczuk, S. Verma, B. Overmoyer, V. Garg, E. Ette, M. W. Harding, et al. Safety and Efficacy of the Multidrug Resistance Inhibitor Incel (Biricodar; VX-710) in Combination with Paclitaxel for Advanced Breast Cancer Refractory to Paclitaxel Clin. Cancer Res., March 1, 2002; 8(3): 670 - 678. [Abstract] [Full Text] [PDF]

				E. Auzenne, N. J. Donato, C. Li, E. Leroux, R. E. Price, D. Farquhar, and J. Klostergaard Superior Therapeutic Profile of Poly-L-Glutamic Acid-Paclitaxel Copolymer Compared with Taxol in Xenogeneic Compartmental Models of Human Ovarian Carcinoma Clin. Cancer Res., February 1, 2002; 8(2): 573 - 581. [Abstract] [Full Text] [PDF]

				A. Goncalves, D. Braguer, K. Kamath, L. Martello, C. Briand, S. Horwitz, L. Wilson, and M. A. Jordan Resistance to Taxol in lung cancer cells associated with increased microtubule dynamics PNAS, September 13, 2001; (2001) 191388598. [Abstract] [Full Text] [PDF]

				S. H. Jang, M. G. Wientjes, and J. L.-S. Au Kinetics of P-Glycoprotein-Mediated Efflux of Paclitaxel J. Pharmacol. Exp. Ther., September 1, 2001; 298(3): 1236 - 1242. [Abstract] [Full Text] [PDF]

				K. E. Luker, C. M. Pica, R. D. Schreiber, and D. Piwnica-Worms Overexpression of IRF9 Confers Resistance to Antimicrotubule Agents in Breast Cancer Cells Cancer Res., September 1, 2001; 61(17): 6540 - 6547. [Abstract] [Full Text] [PDF]

				J. T. Lin, R. Sharma, J. J. Grady, and S. Awasthi A Flow Cell Assay for Evaluation of Whole Cell Drug Efflux Kinetics: Analysis of Paclitaxel Efflux in CCRF-CEM Leukemia Cells Overexpressing P-Glycoprotein Drug Metab. Dispos., February 1, 2001; 29(2): 103 - 110. [Abstract] [Full Text]

				X. Lin, K. Ramamurthi, M. Mishima, A. Kondo, R. D. Christen, and S. B. Howell p53 Modulates the Effect of Loss of DNA Mismatch Repair on the Sensitivity of Human Colon Cancer Cells to the Cytotoxic and Mutagenic Effects of Cisplatin Cancer Res., February 1, 2001; 61(4): 1508 - 1516. [Abstract] [Full Text]

				H.-J. Kuh, S. H. Jang, M. G. Wientjes, and J. L.-S. Au Computational Model of Intracellular Pharmacokinetics of Paclitaxel J. Pharmacol. Exp. Ther., June 1, 2000; 293(3): 761 - 770. [Abstract] [Full Text]

				P. J. O'Dwyer, J. P. Stevenson, M. Gallagher, A. Cassella, I. Vasilevskaya, B. P. Monia, J. Holmlund, F. A. Dorr, and K.-S. Yao c-raf-1 Depletion and Tumor Responses in Patients Treated with the c-raf-1 Antisense Oligodeoxynucleotide ISIS 5132 (CGP 69846A) Clin. Cancer Res., December 1, 1999; 5(12): 3977 - 3982. [Abstract] [Full Text] [PDF]

				E. Harper, W. Dang, R. G. Lapidus, and R. I. Garver Jr. Enhanced Efficacy of a Novel Controlled Release Paclitaxel Formulation (PACLIMER Delivery System) for Local-Regional Therapy of Lung Cancer Tumor Nodules in Mice Clin. Cancer Res., December 1, 1999; 5(12): 4242 - 4248. [Abstract] [Full Text] [PDF]

				Z. Duan, A. J. Feller, R. T. Penson, Bruce. A. Chabner, and M. V. Seiden Discovery of Differentially Expressed Genes Associated with Paclitaxel Resistance Using cDNA Array Technology: Analysis of Interleukin (IL) 6, IL-8, and Monocyte Chemotactic Protein 1 in the Paclitaxel-resistant Phenotype Clin. Cancer Res., November 1, 1999; 5(11): 3445 - 3453. [Abstract] [Full Text] [PDF]

				X. Lin and S. B. Howell Effect of Loss of DNA Mismatch Repair on Development of Topotecan-, Gemcitabine-, and Paclitaxel-Resistant Variants after Exposure to Cisplatin Mol. Pharmacol., August 1, 1999; 56(2): 390 - 395. [Abstract] [Full Text]

				M. Monzo, R. Rosell, J. J. Sanchez, J. S. Lee, A. O'Brate, J. L. Gonzalez-Larriba, V. Alberola, J. C. Lorenzo, L. Nunez, J. Y. Ro, et al. Paclitaxel Resistance in Non-Small-Cell Lung Cancer Associated With Beta-Tubulin Gene Mutations J. Clin. Oncol., June 1, 1999; 17(6): 1786 - 1786. [Abstract] [Full Text] [PDF]

				P. L. Judson, J. M. Watson, P. A. Gehrig, W. C. Fowler Jr., and J. S. Haskill Cisplatin Inhibits Paclitaxel-induced Apoptosis in Cisplatin-resistant Ovarian Cancer Cell Lines: Possible Explanation for Failure ofCombination Therapy Cancer Res., May 1, 1999; 59(10): 2425 - 2432. [Abstract] [Full Text] [PDF]

				K Blade, D. Menick, and F Cabral Overexpression of class I, II or IVb beta-tubulin isotypes in CHO cells is insufficient to confer resistance to paclitaxel J. Cell Sci., January 7, 1999; 112(13): 2213 - 2221. [Abstract] [PDF]

				P. Giannakakou, D. L. Sackett, Y.-K. Kang, Z. Zhan, J. T. M. Buters, T. Fojo, and M. S. Poruchynsky Paclitaxel-resistant Human Ovarian Cancer Cells Have Mutant beta -Tubulins That Exhibit Impaired Paclitaxel-driven Polymerization J. Biol. Chem., July 4, 1997; 272(27): 17118 - 17125. [Abstract] [Full Text] [PDF]

				A. Goncalves, D. Braguer, K. Kamath, L. Martello, C. Briand, S. Horwitz, L. Wilson, and M. A. Jordan Resistance to Taxol in lung cancer cells associated with increased microtubule dynamics PNAS, September 25, 2001; 98(20): 11737 - 11742. [Abstract] [Full Text] [PDF]