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. 1989 Oct;9(10):4357–4363. doi: 10.1128/mcb.9.10.4357

Expression of a human multidrug resistance cDNA (MDR1) in the bone marrow of transgenic mice: resistance to daunomycin-induced leukopenia.

H Galski 1, M Sullivan 1, M C Willingham 1, K V Chin 1, M M Gottesman 1, I Pastan 1, G T Merlino 1
PMCID: PMC362516  PMID: 2573831

Abstract

The human multidrug resistance gene (MDR1) encodes a drug efflux pump glycoprotein (P-glycoprotein) responsible for resistance to multiple cytotoxic drugs. A plasmid carrying a human MDR1 cDNA under the control of a chicken beta-actin promoter was used to generate transgenic mice in which the transgene was mainly expressed in bone marrow and spleen. Immunofluorescence localization studies showed that P-glycoprotein was present on bone marrow cells. Furthermore, leukocyte counts of the transgenic mice treated with daunomycin did not fall, indicating that their bone marrow was resistant to the cytotoxic effect of the drug. Since bone marrow suppression is a major limitation to chemotherapy, these transgenic mice should serve as a model to determine whether higher doses of drugs can cure previously unresponsive cancers.

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Selected References

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

  1. Akiyama S., Fojo A., Hanover J. A., Pastan I., Gottesman M. M. Isolation and genetic characterization of human KB cell lines resistant to multiple drugs. Somat Cell Mol Genet. 1985 Mar;11(2):117–126. doi: 10.1007/BF01534700. [DOI] [PubMed] [Google Scholar]
  2. Beck W. T., Mueller T. J., Tanzer L. R. Altered surface membrane glycoproteins in Vinca alkaloid-resistant human leukemic lymphoblasts. Cancer Res. 1979 Jun;39(6 Pt 1):2070–2076. [PubMed] [Google Scholar]
  3. Biedler J. L., Riehm H. Cellular resistance to actinomycin D in Chinese hamster cells in vitro: cross-resistance, radioautographic, and cytogenetic studies. Cancer Res. 1970 Apr;30(4):1174–1184. [PubMed] [Google Scholar]
  4. Brinster R. L., Allen J. M., Behringer R. R., Gelinas R. E., Palmiter R. D. Introns increase transcriptional efficiency in transgenic mice. Proc Natl Acad Sci U S A. 1988 Feb;85(3):836–840. doi: 10.1073/pnas.85.3.836. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Chen C. J., Chin J. E., Ueda K., Clark D. P., Pastan I., Gottesman M. M., Roninson I. B. Internal duplication and homology with bacterial transport proteins in the mdr1 (P-glycoprotein) gene from multidrug-resistant human cells. Cell. 1986 Nov 7;47(3):381–389. doi: 10.1016/0092-8674(86)90595-7. [DOI] [PubMed] [Google Scholar]
  6. Chomczynski P., Sacchi N. Single-step method of RNA isolation by acid guanidinium thiocyanate-phenol-chloroform extraction. Anal Biochem. 1987 Apr;162(1):156–159. doi: 10.1006/abio.1987.9999. [DOI] [PubMed] [Google Scholar]
  7. Cornwell M. M., Safa A. R., Felsted R. L., Gottesman M. M., Pastan I. Membrane vesicles from multidrug-resistant human cancer cells contain a specific 150- to 170-kDa protein detected by photoaffinity labeling. Proc Natl Acad Sci U S A. 1986 Jun;83(11):3847–3850. doi: 10.1073/pnas.83.11.3847. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Cornwell M. M., Tsuruo T., Gottesman M. M., Pastan I. ATP-binding properties of P glycoprotein from multidrug-resistant KB cells. FASEB J. 1987 Jul;1(1):51–54. doi: 10.1096/fasebj.1.1.2886389. [DOI] [PubMed] [Google Scholar]
  9. Dano K. Active outward transport of daunomycin in resistant Ehrlich ascites tumor cells. Biochim Biophys Acta. 1973 Oct 25;323(3):466–483. doi: 10.1016/0005-2736(73)90191-0. [DOI] [PubMed] [Google Scholar]
  10. DePonti-Zilli L., Seiler-Tuyns A., Paterson B. M. A 40-base-pair sequence in the 3' end of the beta-actin gene regulates beta-actin mRNA transcription during myogenesis. Proc Natl Acad Sci U S A. 1988 Mar;85(5):1389–1393. doi: 10.1073/pnas.85.5.1389. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Elder P. K., French C. L., Subramaniam M., Schmidt L. J., Getz M. J. Evidence that the functional beta-actin gene is single copy in most mice and is associated with 5' sequences capable of conferring serum- and cycloheximide-dependent regulation. Mol Cell Biol. 1988 Jan;8(1):480–485. doi: 10.1128/mcb.8.1.480. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Fojo A. T., Shen D. W., Mickley L. A., Pastan I., Gottesman M. M. Intrinsic drug resistance in human kidney cancer is associated with expression of a human multidrug-resistance gene. J Clin Oncol. 1987 Dec;5(12):1922–1927. doi: 10.1200/JCO.1987.5.12.1922. [DOI] [PubMed] [Google Scholar]
  13. Fojo A. T., Ueda K., Slamon D. J., Poplack D. G., Gottesman M. M., Pastan I. Expression of a multidrug-resistance gene in human tumors and tissues. Proc Natl Acad Sci U S A. 1987 Jan;84(1):265–269. doi: 10.1073/pnas.84.1.265. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Gabizon A., Meshorer A., Barenholz Y. Comparative long-term study of the toxicities of free and liposome-associated doxorubicin in mice after intravenous administration. J Natl Cancer Inst. 1986 Aug;77(2):459–469. [PubMed] [Google Scholar]
  15. Goldstein L. J., Galski H., Fojo A., Willingham M., Lai S. L., Gazdar A., Pirker R., Green A., Crist W., Brodeur G. M. Expression of a multidrug resistance gene in human cancers. J Natl Cancer Inst. 1989 Jan 18;81(2):116–124. doi: 10.1093/jnci/81.2.116. [DOI] [PubMed] [Google Scholar]
  16. Gottesman M. M., Pastan I. The multidrug transporter, a double-edged sword. J Biol Chem. 1988 Sep 5;263(25):12163–12166. [PubMed] [Google Scholar]
  17. Gros P., Ben Neriah Y. B., Croop J. M., Housman D. E. Isolation and expression of a complementary DNA that confers multidrug resistance. Nature. 1986 Oct 23;323(6090):728–731. doi: 10.1038/323728a0. [DOI] [PubMed] [Google Scholar]
  18. Gros P., Croop J., Housman D. Mammalian multidrug resistance gene: complete cDNA sequence indicates strong homology to bacterial transport proteins. Cell. 1986 Nov 7;47(3):371–380. doi: 10.1016/0092-8674(86)90594-5. [DOI] [PubMed] [Google Scholar]
  19. Guild B. C., Mulligan R. C., Gros P., Housman D. E. Retroviral transfer of a murine cDNA for multidrug resistance confers pleiotropic drug resistance to cells without prior drug selection. Proc Natl Acad Sci U S A. 1988 Mar;85(5):1595–1599. doi: 10.1073/pnas.85.5.1595. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Gunning P., Leavitt J., Muscat G., Ng S. Y., Kedes L. A human beta-actin expression vector system directs high-level accumulation of antisense transcripts. Proc Natl Acad Sci U S A. 1987 Jul;84(14):4831–4835. doi: 10.1073/pnas.84.14.4831. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Hamada H., Tsuruo T. Functional role for the 170- to 180-kDa glycoprotein specific to drug-resistant tumor cells as revealed by monoclonal antibodies. Proc Natl Acad Sci U S A. 1986 Oct;83(20):7785–7789. doi: 10.1073/pnas.83.20.7785. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Horio M., Gottesman M. M., Pastan I. ATP-dependent transport of vinblastine in vesicles from human multidrug-resistant cells. Proc Natl Acad Sci U S A. 1988 May;85(10):3580–3584. doi: 10.1073/pnas.85.10.3580. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Jaenisch R. Transgenic animals. Science. 1988 Jun 10;240(4858):1468–1474. doi: 10.1126/science.3287623. [DOI] [PubMed] [Google Scholar]
  24. Juliano R. L., Ling V. A surface glycoprotein modulating drug permeability in Chinese hamster ovary cell mutants. Biochim Biophys Acta. 1976 Nov 11;455(1):152–162. doi: 10.1016/0005-2736(76)90160-7. [DOI] [PubMed] [Google Scholar]
  25. Maral R. J., Jouanne M. Toxicology of daunorubicin in animals and man. Cancer Treat Rep. 1981;65 (Suppl 4):9–18. [PubMed] [Google Scholar]
  26. Ng S. Y., Gunning P., Eddy R., Ponte P., Leavitt J., Shows T., Kedes L. Evolution of the functional human beta-actin gene and its multi-pseudogene family: conservation of noncoding regions and chromosomal dispersion of pseudogenes. Mol Cell Biol. 1985 Oct;5(10):2720–2732. doi: 10.1128/mcb.5.10.2720. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Palmiter R. D., Brinster R. L. Germ-line transformation of mice. Annu Rev Genet. 1986;20:465–499. doi: 10.1146/annurev.ge.20.120186.002341. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Pastan I., Gottesman M. M., Ueda K., Lovelace E., Rutherford A. V., Willingham M. C. A retrovirus carrying an MDR1 cDNA confers multidrug resistance and polarized expression of P-glycoprotein in MDCK cells. Proc Natl Acad Sci U S A. 1988 Jun;85(12):4486–4490. doi: 10.1073/pnas.85.12.4486. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Pollard T. D., Cooper J. A. Actin and actin-binding proteins. A critical evaluation of mechanisms and functions. Annu Rev Biochem. 1986;55:987–1035. doi: 10.1146/annurev.bi.55.070186.005011. [DOI] [PubMed] [Google Scholar]
  30. Quitschke W. W., Lin Z. Y., DePonti-Zilli L., Paterson B. M. The beta actin promoter. High levels of transcription depend upon a CCAAT binding factor. J Biol Chem. 1989 Jun 5;264(16):9539–9546. [PubMed] [Google Scholar]
  31. Rigby P. W., Dieckmann M., Rhodes C., Berg P. Labeling deoxyribonucleic acid to high specific activity in vitro by nick translation with DNA polymerase I. J Mol Biol. 1977 Jun 15;113(1):237–251. doi: 10.1016/0022-2836(77)90052-3. [DOI] [PubMed] [Google Scholar]
  32. Riordan J. R., Deuchars K., Kartner N., Alon N., Trent J., Ling V. Amplification of P-glycoprotein genes in multidrug-resistant mammalian cell lines. 1985 Aug 29-Sep 4Nature. 316(6031):817–819. doi: 10.1038/316817a0. [DOI] [PubMed] [Google Scholar]
  33. Scotto K. W., Biedler J. L., Melera P. W. Amplification and expression of genes associated with multidrug resistance in mammalian cells. Science. 1986 May 9;232(4751):751–755. doi: 10.1126/science.2421411. [DOI] [PubMed] [Google Scholar]
  34. Shen D. W., Cardarelli C., Hwang J., Cornwell M., Richert N., Ishii S., Pastan I., Gottesman M. M. Multiple drug-resistant human KB carcinoma cells independently selected for high-level resistance to colchicine, adriamycin, or vinblastine show changes in expression of specific proteins. J Biol Chem. 1986 Jun 15;261(17):7762–7770. [PubMed] [Google Scholar]
  35. Shen D. W., Fojo A., Chin J. E., Roninson I. B., Richert N., Pastan I., Gottesman M. M. Human multidrug-resistant cell lines: increased mdr1 expression can precede gene amplification. Science. 1986 May 2;232(4750):643–645. doi: 10.1126/science.3457471. [DOI] [PubMed] [Google Scholar]
  36. Shen D. W., Fojo A., Roninson I. B., Chin J. E., Soffir R., Pastan I., Gottesman M. M. Multidrug resistance of DNA-mediated transformants is linked to transfer of the human mdr1 gene. Mol Cell Biol. 1986 Nov;6(11):4039–4045. doi: 10.1128/mcb.6.11.4039. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Southern E. M. Detection of specific sequences among DNA fragments separated by gel electrophoresis. J Mol Biol. 1975 Nov 5;98(3):503–517. doi: 10.1016/s0022-2836(75)80083-0. [DOI] [PubMed] [Google Scholar]
  38. Stiles C. D., Lee K. L., Kenney F. T. Differential degradation of messenger RNAs in mammalian cells. Proc Natl Acad Sci U S A. 1976 Aug;73(8):2634–2638. doi: 10.1073/pnas.73.8.2634. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. Ueda K., Cardarelli C., Gottesman M. M., Pastan I. Expression of a full-length cDNA for the human "MDR1" gene confers resistance to colchicine, doxorubicin, and vinblastine. Proc Natl Acad Sci U S A. 1987 May;84(9):3004–3008. doi: 10.1073/pnas.84.9.3004. [DOI] [PMC free article] [PubMed] [Google Scholar]
  40. Ueda K., Clark D. P., Chen C. J., Roninson I. B., Gottesman M. M., Pastan I. The human multidrug resistance (mdr1) gene. cDNA cloning and transcription initiation. J Biol Chem. 1987 Jan 15;262(2):505–508. [PubMed] [Google Scholar]
  41. Ueda K., Cornwell M. M., Gottesman M. M., Pastan I., Roninson I. B., Ling V., Riordan J. R. The mdr1 gene, responsible for multidrug-resistance, codes for P-glycoprotein. Biochem Biophys Res Commun. 1986 Dec 30;141(3):956–962. doi: 10.1016/s0006-291x(86)80136-x. [DOI] [PubMed] [Google Scholar]
  42. Van der Bliek A. M., Van der Velde-Koerts T., Ling V., Borst P. Overexpression and amplification of five genes in a multidrug-resistant Chinese hamster ovary cell line. Mol Cell Biol. 1986 May;6(5):1671–1678. doi: 10.1128/mcb.6.5.1671. [DOI] [PMC free article] [PubMed] [Google Scholar]
  43. Willingham M. C., Cornwell M. M., Cardarelli C. O., Gottesman M. M., Pastan I. Single cell analysis of daunomycin uptake and efflux in multidrug-resistant and -sensitive KB cells: effects of verapamil and other drugs. Cancer Res. 1986 Nov;46(11):5941–5946. [PubMed] [Google Scholar]
  44. Willingham M. C., Richert N. D., Cornwell M. M., Tsuruo T., Hamada H., Gottesman M. M., Pastan I. H. Immunocytochemical localization of P170 at the plasma membrane of multidrug-resistant human cells. J Histochem Cytochem. 1987 Dec;35(12):1451–1456. doi: 10.1177/35.12.2890686. [DOI] [PubMed] [Google Scholar]

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