Epithelial and fibroblast cell lines derived from a spontaneous mammary carcinoma in a MMTV/neu transgenic mouse

  • Michael J. Campbell
  • Wendy S. Wollish
  • Margaret Lobo
  • Laura J. Esserman
Articles Cell and Tissue Models

Summary

Female murine mammary tumor virus (MMTV)/neu transgenic mice, expressing a wild-type rat neu oncogene driven by an MMTV promoter, develop focal mammary adenocarcinomas that are pathologically very similar to human breast tumors. Two new cell lines were established from a mammary tumor that arose in a female MMTV/neu transgenic mouse. One of these lines, mammary carcinoma from Neu transgenic mouse A (MCNeuA), has an epithelial morphology, is cytokeratin positive, and expresses high levels of the neu transgene. Karyotyping and comparative genomic hybridization analyses demonstrated genomic alterations in the MCNeuA cell line. The other line, N202Fb3, has a fibroblast morphology, is cytokeratin negative, and expresses the neu transgene at a very low level. This cell line also expresses smooth muscle α-actin, suggesting that it is a myofibroblast line. The MCNeuA cell line is tumorigenic when injected into syngeneic MMTV/neu transgenic mice, with an in vivo doubling time of about 14 d. The rationale for establishing this tumor cell line was to provide a tumor transplantation system for rapidly assessing immunotherapeutic interventions before testing in the more cumbersome model of spontaneous tumor development in the MMTV/neu transgenic mice. Mice immunized with a Neu extracellular domain protein vaccine were protected against a subsequent inoculation of MCNeuA cells, indicating that this cell line will be useful for evaluating cancer vaccine strategies. This tumor cell line may also prove useful in studying the biological properties of the neu oncogene and its role in the malignant process. In addition, the tumor-derived fibroblast line may be useful for studying tumor-stromal cell interactions.

Key words

breast cancer cell line HER2/neu transgenic mice 

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References

  1. Adams, J. M.; Cory, S. Transgenic models of tumor development. Science 254:1161–1167; 1991.PubMedCrossRefGoogle Scholar
  2. Amici, A.; Venanzi, F. M.; Concetti, A. Genetic immunization against neu/erbB2 transgenic breast cancer. Cancer Immunol. Immunother. 47:183–190; 1998.PubMedCrossRefGoogle Scholar
  3. Baselga, J.; Tripathy, D.; Mendelsohn, J., et al. Phase II study of weekly intravenous recombinant humanized anti-p185HER2 monoclonal antibody in patients with HER2/neu-overexpressing metastatic breast cancer. J. Clin. Oncol. 14:737–744; 1996.PubMedGoogle Scholar
  4. Bellacosa, A.; Franke, T. F.; Gonzalez-Portal, M. E., et al. Structure, expression and chromosomal mapping of c-akt: relationship to v-akt and its implications. Oncogene 8:745–754; 1993.PubMedGoogle Scholar
  5. Berx, G.; Cleton-Jansen, A. M.; Nollet, F.; de Leeuw, W. J.; van de Vijver, M.; Corneliss, C.; van Roy, F. E-cadherin is a tumour/invasion suppressor gene mutated in human lobular breast cancers. Embo. J. 14:6107–6115; 1995.PubMedGoogle Scholar
  6. Bieche, I.; Champeme, M. H.; Lidereau, R. A tumor suppressor gene on chromosome 1p32-pter controls the amplification of MYC family genes in breast cancer. Cancer Res. 54:4274–4276; 1994.PubMedGoogle Scholar
  7. Bieche, I.; Champeme, M. H.; Matifas, F.; Cropp, C. S.; Callahan, R.; Lidereau, R. Two distinct regions involved in 1p deletion in human primary breast cancer. Cancer Res. 53:1990–1994; 1993.PubMedGoogle Scholar
  8. Brenner, A. J.; Aldaz, C. M. Chromosome 9p allelic loss and p16/CDKN2 in breast cancer and evidence of p16 inactivation in immortal breast epithelial cells. Cancer Res. 55:2892–2895; 1995.PubMedGoogle Scholar
  9. Campbell, M. J.; Esserman, L.; Byars, N. E.; Allison, A. C.; Levy, R. Idiotype vaccination against murine B cell lymphoma. Humoral and cellular requirements for the full expression of antitumor immunity. J. Immunol. 145:1029–1036; 1990.PubMedGoogle Scholar
  10. Cardiff, R. D.; Muller, W. J. Transgenic mouse models of mammary tumorigenesis. Cancer Surv. 16:97–113; 1993.PubMedGoogle Scholar
  11. Cefai, D.; Morrison, B. W.; Sckell, A.; Favre, L.; Balli, M.; Leunig, M.; Gimmi, C. D. Targeting HER-2/neu for active-specific immunotherapy in a mouse model of spontaneous breast cancer. Int. J. Cancer 83:393–400; 1999.PubMedCrossRefGoogle Scholar
  12. Chen, Y.; Hu, D.; Eling, D. J.; Robbins, J.; Kipps, T. J. DNA vaccines encoding full-length or truncated Neu induce protective immunity against Neu-expressing mammary tumors. Cancer Res. 58:1965–1971; 1998.PubMedGoogle Scholar
  13. Concetti, A.; Amici, A.; Petrelli, C.; Tibaldi, A.; Provinciali, M.; Venanzi, F. M. Autoantibody to p185erbB2/neu oncoprotein by vaccination with xenogenic DNA. Cancer Immunol. Immunother. 43:307–315; 1996.PubMedCrossRefGoogle Scholar
  14. Cool, M.; Jolicoeur, P. Elevated frequency of loss of heterozygosity in mammary tumors arising in mouse mammary tumor virus/neu transgenic mice. Cancer Res. 59:2438–2444; 1999.PubMedGoogle Scholar
  15. Devereux, T. R.; Anna, C. H.; Patel, A. C.; White, C. M.; Festing, M. F.; You, M. Smad4 (homolog of human DPC4) and Smad2 (homolog of human JV18-1): candidates for murine lung tumor resistance and suppressor genes. Carcinogenesis 18:1751–1755; 1997.PubMedCrossRefGoogle Scholar
  16. Disis, M. L.; Grabstein, K. H.; Sleath, P. R.; Cheever, M. A. Generation of immunity to the HER-2/neu oncogenic protein in patients with breast and ovarian cancer using a peptide-based vaccine. Clin. Cancer Res. 5:1289–1297; 1999.PubMedGoogle Scholar
  17. Disis, M. L.; Gralow, J. R.; Bernhard, H.; Hand, S. L.; Rubin, W. D.; Cheever, M. A. Peptide-based, but not whole protein, vaccines, elicit immunity to HER-2/neu, oncogenic self-protein. J. Immunol. 156:3151–3158; 1996.PubMedGoogle Scholar
  18. Driouch, K.; Dorion-Bonnet, F.; Briffod, M.; Champeme, M. H.; Longy M.; Lidereau, R. Loss of heterozygosity on chromosome arm 16q in breast cancer metastases. Genes Chromosomes Cancer 19:185–191; 1997.PubMedCrossRefGoogle Scholar
  19. Edwards, B. K.; Howe, H. L.; Ries, L. A. G. Annual report to the nation on the status of cancer, 1973–1999, featuring implications of age and aging on the US cancer burden. Cancer 94:2766–2792; 2002.PubMedCrossRefGoogle Scholar
  20. Esserman, L. J.; Lopez, T.; Montes, R.; Bald, L. N.; Fendly, B. M.; Campbell, M. J. Vaccination with the extracellular domain of p185neu prevents mammary tumor development in neu transgenic mice. Cancer Immunol. Immunother. 47:337–342; 1999.PubMedCrossRefGoogle Scholar
  21. Fendly, B. M.; Kotts, C.; Vetterlein, D., et al. The extracellular domain of HER2/neu is a potential immunogen for active specific immunotherapy of breast cancer. J. Biol. Response Modif. 9:449–455; 1990.Google Scholar
  22. Foster, B. A.; Gingrich, J. R.; Kwon, E. D.; Madias, C.; Greenberg, N. M. Characterization of prostatic epithelial cell lines derived from transgenic adenocarcinoma of the mouse prostate (TRAMP) model. Cancer Res. 57:3325–3330; 1997.PubMedGoogle Scholar
  23. Freshney, R. I. Culture of animal cells: a manual of basic techniques. New York: John Wiley & Sons; 1994:166–169.Google Scholar
  24. Geradts, J.; Wilson, P. A. High frequency of aberrant p16(INK4A) expression in human breast cancer. Am. J. Pathol. 149:15–20; 1996.PubMedGoogle Scholar
  25. Guy, C. T.; Webster, M. A.; Schaller, M.; Parsons, T. J.; Cardiff, R. D.; Muller, W. J. Expression of the neu protooncogene in the mammary epithelium of transgenic mice induces metastatic disease. Proc. Natl. Acad. Sci. USA 89:10578–10582; 1992.PubMedCrossRefGoogle Scholar
  26. Habuchi, T.; Luscombe, M.; Elder, P. A.; Knowles, M. A. Structure and methylation-based silencing of a gene (DBCCR1) within a candidate bladder cancer tumor suppressor region at 9q32-q33. Genomics 48:277–288; 1998.PubMedCrossRefGoogle Scholar
  27. Hegi, M. E.; Devereux, T. R.; Dietrich, W. F. Allelotype analysis of mouse lung carcinomas reveals frequent allelic losses on chromosome 4 and an association between allelic imbalances on chromosome 6 and K-ras activation. Cancer Res 54:6257–6264; 1994.PubMedGoogle Scholar
  28. Hendrich, B.; Abbott, C.; McQueen, H.; Chambers, D.; Cross, S.; Bird, A. Genomic structure and chromosomal mapping of the murine and human Mbd1, Mbd2, Mbd3, and Mbd4 genes. Mamm. Genome 10:906–912; 1999.PubMedCrossRefGoogle Scholar
  29. Herman, J. G.; Jen, J.; Merlo, A.; Baylin, S. B. Hypermethylation-associated inactivation indicates a tumor suppressor role for p15INK4B. Cancer Res. 56:722–727; 1996.PubMedGoogle Scholar
  30. Herman, J. G.; Merlo, A.; Mao, L.; Lapidus, R. G.; Issa, J. P.; Davidson, N. E.; Sidransky, D.; Baylin, S. B. Inactivation of the CDKN2/p16/MTS1 gene is frequently associated with aberrant DNA methylation in all common human cancers. Lancet Res. 55:4525–4530; 1995.Google Scholar
  31. Herzog, C. R.; Wiseman, R. W.; You, M. Deletion mapping of a putative tumor suppressor gene on chromosome 4 in mouse lung tumors. Cancer Res. 54:4007–4010; 1994.PubMedGoogle Scholar
  32. Kremmidiotis, C.; Baker, E.; Crawford, J.; Eyre, H. J.; Nahmias, J.; Callen, D. F. Localization of human cadherin genes to chromosome regions exhibiting cancer-related loss of heterozygosity. Genomics 49:467–471; 1998.PubMedCrossRefGoogle Scholar
  33. Lahti, J. M.; Valentine, M.; Xiang, J., et al. Alterations in the PITSLRE protein kinase gene complex on chromosome 1p36 in childhood neuroblastoma. Nat. Genet. 7:370–375; 1994.PubMedCrossRefGoogle Scholar
  34. Lazard, D.; Sastre, X.; Frid, M. G.; Glukhova, M. A.; Thiery, J. P.; Koteliansky, V. E. Expression of smooth muscle-specific proteins in myoepithelium and stromal myofibroblasts of normal and malignant human breast tissue. Proc. Natl. Acad. Sci. USA 90:999–1003; 1993.PubMedCrossRefGoogle Scholar
  35. Li, R.; Yerganian, G.; Duesberg, P.; Kraemer, A.; Willer, A.; Rausch, C.; Hehlmann, R. Aneuploidy correlated 100% with chemical transformation of Chinese hamster cells. Proc. Natl. Acad. Sci. USA 94:14506–14511; 1997.PubMedCrossRefGoogle Scholar
  36. Liu, E.; Thor, A.; He, M.; Barcos, M.; Ljung, B. M.; Benz, C. The HER2 (c-erbB-2) oncogene is frequently amplified in in situ carcinomas of the breast. Oncogene 7:1027–1032; 1992.PubMedGoogle Scholar
  37. Mosmann, T. Rapid colorimetric assay for cellular growth and survival: application to proliferation and cytotoxicity assays. J. Immunol. Methods 65:55–63; 1983.PubMedCrossRefGoogle Scholar
  38. Muller, W. J.; Sinn, E.; Pattengale, P. K.; Wallace, R.; Leder, P. Single-step induction of mammary adenocarcinoma in transgenic mice bearing the activated c-neu oncogene. Cell 54:105–115; 1988.PubMedCrossRefGoogle Scholar
  39. Nagai, H.; Negrini, M.; Carter, S. L.; Gillum, D. R.; Rosenberg, A. L.; Schwartz, G. F.; Croce, C. M. Detection and cloning of a common region of loss of heterozygosity at chromosome 1p in breast cancer. Cancer Res. 55:1752–1757; 1995.PubMedGoogle Scholar
  40. Nagata, Y.; Furugen, R.; Hiasa, A., et al. Peptides derived from a wild-type murine proto-oncogene c-erbB-2/HER2/neu can induce, CTL and tumor suppression in syngeneic hosts. J. Immunol. 159:1336–1343; 1997.PubMedGoogle Scholar
  41. Narducci, M. G.; Virgilio, L.; Engiles, J. B., et al. The murine Tcll, oncogene: embryonic and lymphoid cell expression. Oncogene 15:919–926; 1997.PubMedCrossRefGoogle Scholar
  42. Phelan, C. M.; Larsson, C.; Baird, S., et al. The human mammary-derived growth inhibitor (MDGI) gene: genomic structure and mutation analysis in human breast tumors. Genomics 34:63–68; 1996.PubMedCrossRefGoogle Scholar
  43. Quelle, D. E.; Ashmun, R. A.; Hannon, G. J., et al. Cloning and characterization of murine p16INK4a and p15INK4b genes. Oncogene 11:635–645; 1995.PubMedGoogle Scholar
  44. Recio, J. A.; Zambrano, N.; de La Pena, L.; Powers, C.; Siwarski, D.; Huppi, K.; Notario, V. cDNA isolation, expression, and chromosomal localization of the mouse pcph proto-oncogene. Mol. Carcinog. 26:130–136; 1999.PubMedCrossRefGoogle Scholar
  45. Ritland, S. R.; Rowse, G. J.; Chang, Y.; Gendler, S. J. Loss of heterozygosity analysis in primary mammary tumors and lung metastases of MMTV-MTAg and MMTV-neu transgenic mice. Cancer Res. 57:3520–3525; 1997.PubMedGoogle Scholar
  46. Sacco, M. G.; Gribaldo, L.; Barbieri, O., et al. Establishment and characterization of a new mammary adenocarcinoma cell line derived from MMTV neu transgenic mice. Breast Cancer Res Treat. 47:171–180; 1998.PubMedCrossRefGoogle Scholar
  47. Sheng, Z. M.; Marchetti, A.; Buttitta, F.; Champeme, M. H.; Campani, D.; Bistocchi, M.; Lidereau, R.; Callahan, R. Multiple regions of chromosome 6q affected by loss of heterozygosity in primary human breast carcinomas. Br. J. Cancer 73:144–147; 1996.PubMedGoogle Scholar
  48. Shepard, H. M.; Lewis, G. D.; Sarup, J. C., et al. Monoclonal antibody therapy of human cancer: taking the HER2 protooncogene to the clinie. J. Clin. Immunol. 11:117–127; 1991.PubMedCrossRefGoogle Scholar
  49. Siegel, P. M.; Dankort, D. L.; Hardy, W. R.; Muller, W. J. Novel activating mutations in the neu proto-oncogene involved in induction of mammary tumors. Mol. Cell. Biol. 14:7068–7077; 1994.PubMedGoogle Scholar
  50. Siegel, P. M.; Muller, W. J. Mutations affecting conserved cysteine residues within the extracellular domain of Neu promote receptor dimerization and activation. Proc. Natl. Acad. Sci. USA 93:8878–8883; 1996.PubMedCrossRefGoogle Scholar
  51. Slamon, D.; Clark, G.; Wong, S.; Levin, W.; Ullrich, A.; McGuire, W. Human breast cancer: correlation of relapse and survival with amplification of the HER2/neu oncogene. Science 235:177–182; 1987.PubMedCrossRefGoogle Scholar
  52. Slamon, D. J.; Godolphin, W.; Jones, L. A., et al. Studies of the HER2/neu proto-oncogene in human breast and ovarian cancer. Science 244:707–712; 1989.PubMedCrossRefGoogle Scholar
  53. Sola, B.; Simon, D.; Mattei, M. G.; Fichelson, S.; Bordereaux, D.; Tambourin, P. E.; Guenet, J. L.; Gisselbrecht, S. Fim-1, Fim-2/c-fims, and Fim-3, three common integration sites of Friend murine leukemia virus in myeloblastic leukemias, map to mouse chromosomes 13, 18, and 3, respectively. J. Virol. 62:3973–3978; 1988.PubMedGoogle Scholar
  54. Solin, L. J.; Haffty, B.; Fourquet, A., et al. An international collaborative study: a 15-yr experience. In: Silverstein, M., ed. Ductal carcinoma in situ of the breast. Baltimore, MD: Williams and Wilkins; 1997:385–390.Google Scholar
  55. Tremblay, G. B.; Tremblay, A.; Copeland, N. G.; Gilbert D. J.; Jenkins, N. A.; Labrie, F.; Giguere, V. Cloning, chromosomal localization, and functional analysis of the murine estrogen receptor beta. Mol. Endocrinol. 11:353–365; 1997.PubMedCrossRefGoogle Scholar
  56. Tsukamoto, K.; Ito, N.; Yoshimoto, M.; Kasumi, F.; Akiyama, F.; Sakamoto, G.; Nakamura, Y.; Emi, M. Allelic loss on chromosome 1p is associated with progression and lymph node metastasis of primary breast carcinoma. Cancer 82:317–322; 1998.PubMedCrossRefGoogle Scholar
  57. Van Camp, G.; Coucke, P.; Speleman, F.; Van Roy N.; Beyer, E. C.; Oostra, B. A.; Willems, P. J. The gene for human gap junction protein connexin37 (GJA4) maps to chromosome 1p35.1, in the vicinity of DIS195. Genomics 30:402–403; 1995.PubMedGoogle Scholar
  58. Van de Vijver, M. J.; Peterse, J. L.; Mooi, W. J.; Wisman, P.; Lomans, J.; Dalesio, O.; Nusse, R. Neu-protein overexpression in breast cancer. Association with comedo-type ductal carcinoma in situ and limited prognostic value in stage II breast cancer. N. Engl. J. Med. 319:1239–1245; 1988.PubMedCrossRefGoogle Scholar
  59. Van Zee, K. J.; Calvano, J. E.; Bisogna, M. Hypomethylation and increased gene expression of p16INK4a in primary and metastatic breast carcinoma as compared to normal breast tissue. Oncogene 16:2723–2727; 1998.PubMedCrossRefGoogle Scholar
  60. Webster, M. A.; Cardiff, R. D.; Muller, W. J. Induction of mammary epithelial hyperplasias and mammary tumors in transgenic mice expressing a murine mammary tumor virus/activated c-src fusion gene. Proc. Natl. Acad. Sci. USA 92:7849–7853; 1995.PubMedCrossRefGoogle Scholar
  61. Wei, W. Z.; Shi, W. P.; Galy, A.; Lichlyter, D.; Hernandez, S.; Groner, B.; Heilbrun, L.; Jones, R. F. Protection against mammary tumor growth by vaccination with full-length, modified human ErbB-2 DNA. Int. J. Cancer 81:748–754; 1999.PubMedCrossRefGoogle Scholar
  62. Yamashita, J.; Ogawa, M.; Sakai, K. Prognostic significance of three novel biologic factors in a clinical trial of adjuvant therapy for node-negative breast cancer. Surgery 117:601–608; 1995.PubMedCrossRefGoogle Scholar
  63. Yamashita, S.; Yamashita, J.; Ogawa, M. Overexpression of group II phospholipase A2 in human breast cancer tissues in closely associated with their malignant potency. Br. J. Cancer 69:1166–1170; 1994.PubMedGoogle Scholar
  64. Zhuang, S. M.; Eklund, I. K.; Cochran, C.; Rao, G. N.; Wiseman, R. W.; Soderkvist, P. Allelotype analysis of 2′,3′-dideoxycytidine- and 1,3-butadiene-induced lymphomas in B6C3F1 mice. Cancer Res. 56:3338–3343; 1996.PubMedGoogle Scholar

Copyright information

© Society for In Vitro Biology 2002

Authors and Affiliations

  • Michael J. Campbell
    • 1
  • Wendy S. Wollish
    • 1
  • Margaret Lobo
    • 1
  • Laura J. Esserman
    • 1
  1. 1.Department of SurgeryUniversity of CaliforniaSan Francisco

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