Skip to main content
PMC home page
The Journal of Clinical Investigation logoLink to The Journal of Clinical Investigation
. 1996 Dec 1;98(11):2539–2548. doi: 10.1172/JCI119072

Selective transgene expression for detection and elimination of contaminating carcinoma cells in hematopoietic stem cell sources.

L Chen 1, M Pulsipher 1, D Chen 1, C Sieff 1, A Elias 1, H A Fine 1, D W Kufe 1
PMCID: PMC507711  PMID: 8958216

Abstract

Tumor contamination of bone marrow (BM) and peripheral blood (PB) may affect the outcome of patients receiving high dose chemotherapy with autologous transplantation of hematopoietic stem cell products. In this report, we demonstrate that replication defective adenoviral vectors containing the cytomegalovirus (CMV) or DF3/MUC1 carcinoma-selective promoter can be used to selectively transduce contaminating carcinoma cells. Adenoviral-mediated reporter gene expression in breast cancer cells was five orders of magnitude higher than that found in BM, PB, and CD34+ cells. Our results demonstrate that CD34+ cells have low to undetectable levels of integrins responsible for adenoviral internalization. We show that adenoviral-mediated transduction of a reporter gene can detect one breast cancer cell in 5 x 10(5) BM or PB cells with a vector containing the DF3/MUC1 promoter. We also show that transduction of the HSV-tk gene for selective killing by ganciclovir can be exploited for purging cancer cells from hematopoietic stem cell populations. The selective expression of TK followed by ganciclovir treatment resulted in the elimination of 6-logs of contaminating cancer cells. By contrast, there was little effect on CFU-GM and BFU-E formulation or on long term culture initiating cells. These results indicate that adenoviral vectors with a tumor-selective promoter provide a highly efficient and effective approach for the detection and purging of carcinoma cells in hematopoietic stem cell preparations.

Full Text

The Full Text of this article is available as a PDF (536.0 KB).

Selected References

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

  1. Abe M., Kufe D. Characterization of cis-acting elements regulating transcription of the human DF3 breast carcinoma-associated antigen (MUC1) gene. Proc Natl Acad Sci U S A. 1993 Jan 1;90(1):282–286. doi: 10.1073/pnas.90.1.282. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Anderson I. C., Shpall E. J., Leslie D. S., Nustad K., Ugelstad J., Peters W. P., Bast R. C., Jr Elimination of malignant clonogenic breast cancer cells from human bone marrow. Cancer Res. 1989 Aug 15;49(16):4659–4664. [PubMed] [Google Scholar]
  3. Antman K., Ayash L., Elias A., Wheeler C., Hunt M., Eder J. P., Teicher B. A., Critchlow J., Bibbo J., Schnipper L. E. A phase II study of high-dose cyclophosphamide, thiotepa, and carboplatin with autologous marrow support in women with measurable advanced breast cancer responding to standard-dose therapy. J Clin Oncol. 1992 Jan;10(1):102–110. doi: 10.1200/JCO.1992.10.1.102. [DOI] [PubMed] [Google Scholar]
  4. Chen L., Chen D., Manome Y., Dong Y., Fine H. A., Kufe D. W. Breast cancer selective gene expression and therapy mediated by recombinant adenoviruses containing the DF3/MUC1 promoter. J Clin Invest. 1995 Dec;96(6):2775–2782. doi: 10.1172/JCI118347. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Clarke M. F., Apel I. J., Benedict M. A., Eipers P. G., Sumantran V., González-García M., Doedens M., Fukunaga N., Davidson B., Dick J. E. A recombinant bcl-x s adenovirus selectively induces apoptosis in cancer cells but not in normal bone marrow cells. Proc Natl Acad Sci U S A. 1995 Nov 21;92(24):11024–11028. doi: 10.1073/pnas.92.24.11024. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Cote R. J., Rosen P. P., Lesser M. L., Old L. J., Osborne M. P. Prediction of early relapse in patients with operable breast cancer by detection of occult bone marrow micrometastases. J Clin Oncol. 1991 Oct;9(10):1749–1756. doi: 10.1200/JCO.1991.9.10.1749. [DOI] [PubMed] [Google Scholar]
  7. Datta Y. H., Adams P. T., Drobyski W. R., Ethier S. P., Terry V. H., Roth M. S. Sensitive detection of occult breast cancer by the reverse-transcriptase polymerase chain reaction. J Clin Oncol. 1994 Mar;12(3):475–482. doi: 10.1200/JCO.1994.12.3.475. [DOI] [PubMed] [Google Scholar]
  8. Diel I. J., Kaufmann M., Goerner R., Costa S. D., Kaul S., Bastert G. Detection of tumor cells in bone marrow of patients with primary breast cancer: a prognostic factor for distant metastasis. J Clin Oncol. 1992 Oct;10(10):1534–1539. doi: 10.1200/JCO.1992.10.10.1534. [DOI] [PubMed] [Google Scholar]
  9. Friedman E. L., Hayes D. F., Kufe D. W. Reactivity of monoclonal antibody DF3 with a high molecular weight antigen expressed in human ovarian carcinomas. Cancer Res. 1986 Oct;46(10):5189–5194. [PubMed] [Google Scholar]
  10. Gerhard M., Juhl H., Kalthoff H., Schreiber H. W., Wagener C., Neumaier M. Specific detection of carcinoembryonic antigen-expressing tumor cells in bone marrow aspirates by polymerase chain reaction. J Clin Oncol. 1994 Apr;12(4):725–729. doi: 10.1200/JCO.1994.12.4.725. [DOI] [PubMed] [Google Scholar]
  11. Graham F. L., Smiley J., Russell W. C., Nairn R. Characteristics of a human cell line transformed by DNA from human adenovirus type 5. J Gen Virol. 1977 Jul;36(1):59–74. doi: 10.1099/0022-1317-36-1-59. [DOI] [PubMed] [Google Scholar]
  12. Harris J. D., Gutierrez A. A., Hurst H. C., Sikora K., Lemoine N. R. Gene therapy for cancer using tumour-specific prodrug activation. Gene Ther. 1994 May;1(3):170–175. [PubMed] [Google Scholar]
  13. Herz J., Gerard R. D. Adenovirus-mediated transfer of low density lipoprotein receptor gene acutely accelerates cholesterol clearance in normal mice. Proc Natl Acad Sci U S A. 1993 Apr 1;90(7):2812–2816. doi: 10.1073/pnas.90.7.2812. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Huber B. E., Richards C. A., Krenitsky T. A. Retroviral-mediated gene therapy for the treatment of hepatocellular carcinoma: an innovative approach for cancer therapy. Proc Natl Acad Sci U S A. 1991 Sep 15;88(18):8039–8043. doi: 10.1073/pnas.88.18.8039. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Jain V. K., Magrath I. T. A chemiluminescent assay for quantitation of beta-galactosidase in the femtogram range: application to quantitation of beta-galactosidase in lacZ-transfected cells. Anal Biochem. 1991 Nov 15;199(1):119–124. doi: 10.1016/0003-2697(91)90278-2. [DOI] [PubMed] [Google Scholar]
  16. Kennedy M. J., Beveridge R. A., Rowley S. D., Gordon G. B., Abeloff M. D., Davidson N. E. High-dose chemotherapy with reinfusion of purged autologous bone marrow following dose-intense induction as initial therapy for metastatic breast cancer. J Natl Cancer Inst. 1991 Jul 3;83(13):920–926. doi: 10.1093/jnci/83.13.920. [DOI] [PubMed] [Google Scholar]
  17. Kufe D., Inghirami G., Abe M., Hayes D., Justi-Wheeler H., Schlom J. Differential reactivity of a novel monoclonal antibody (DF3) with human malignant versus benign breast tumors. Hybridoma. 1984 Fall;3(3):223–232. doi: 10.1089/hyb.1984.3.223. [DOI] [PubMed] [Google Scholar]
  18. Maimonis P., Hayes D. F., O'Hara C., Kufe D. Detection and characterization of a high molecular weight human lung carcinoma-associated glycoprotein. Cancer Res. 1990 Oct 15;50(20):6738–6743. [PubMed] [Google Scholar]
  19. Manome Y., Abe M., Hagen M. F., Fine H. A., Kufe D. W. Enhancer sequences of the DF3 gene regulate expression of the herpes simplex virus thymidine kinase gene and confer sensitivity of human breast cancer cells to ganciclovir. Cancer Res. 1994 Oct 15;54(20):5408–5413. [PubMed] [Google Scholar]
  20. Mathias P., Wickham T., Moore M., Nemerow G. Multiple adenovirus serotypes use alpha v integrins for infection. J Virol. 1994 Oct;68(10):6811–6814. doi: 10.1128/jvi.68.10.6811-6814.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Molino A., Colombatti M., Bonetti F., Zardini M., Pasini F., Perini A., Pelosi G., Tridente G., Veneri D., Cetto G. L. A comparative analysis of three different techniques for the detection of breast cancer cells in bone marrow. Cancer. 1991 Feb 15;67(4):1033–1036. doi: 10.1002/1097-0142(19910215)67:4<1033::aid-cncr2820670428>3.0.co;2-h. [DOI] [PubMed] [Google Scholar]
  22. Myklebust A. T., Godal A., Juell S., Pharo A., Fodstad O. Comparison of two antibody-based methods for elimination of breast cancer cells from human bone marrow. Cancer Res. 1994 Jan 1;54(1):209–214. [PubMed] [Google Scholar]
  23. Nabel E. G., Gordon D., Yang Z. Y., Xu L., San H., Plautz G. E., Wu B. Y., Gao X., Huang L., Nabel G. J. Gene transfer in vivo with DNA-liposome complexes: lack of autoimmunity and gonadal localization. Hum Gene Ther. 1992 Dec;3(6):649–656. doi: 10.1089/hum.1992.3.6-649. [DOI] [PubMed] [Google Scholar]
  24. Nabel E. G., Plautz G., Nabel G. J. Site-specific gene expression in vivo by direct gene transfer into the arterial wall. Science. 1990 Sep 14;249(4974):1285–1288. doi: 10.1126/science.2119055. [DOI] [PubMed] [Google Scholar]
  25. Nolan G. P., Fiering S., Nicolas J. F., Herzenberg L. A. Fluorescence-activated cell analysis and sorting of viable mammalian cells based on beta-D-galactosidase activity after transduction of Escherichia coli lacZ. Proc Natl Acad Sci U S A. 1988 Apr;85(8):2603–2607. doi: 10.1073/pnas.85.8.2603. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Nolta J. A., Smogorzewska E. M., Kohn D. B. Analysis of optimal conditions for retroviral-mediated transduction of primitive human hematopoietic cells. Blood. 1995 Jul 1;86(1):101–110. [PubMed] [Google Scholar]
  27. Pasqualini R., Bodorova J., Ye S., Hemler M. E. A study of the structure, function and distribution of beta 5 integrins using novel anti-beta 5 monoclonal antibodies. J Cell Sci. 1993 May;105(Pt 1):101–111. doi: 10.1242/jcs.105.1.101. [DOI] [PubMed] [Google Scholar]
  28. Passos-Coelho J., Ross A. A., Davis J. M., Huelskamp A. M., Clarke B., Noga S. J., Davidson N. E., Kennedy M. J. Bone marrow micrometastases in chemotherapy-responsive advanced breast cancer: effect of ex vivo purging with 4-hydroperoxycyclophosphamide. Cancer Res. 1994 May 1;54(9):2366–2371. [PubMed] [Google Scholar]
  29. Peters W. P., Ross M., Vredenburgh J. J., Meisenberg B., Marks L. B., Winer E., Kurtzberg J., Bast R. C., Jr, Jones R., Shpall E. High-dose chemotherapy and autologous bone marrow support as consolidation after standard-dose adjuvant therapy for high-risk primary breast cancer. J Clin Oncol. 1993 Jun;11(6):1132–1143. doi: 10.1200/JCO.1993.11.6.1132. [DOI] [PubMed] [Google Scholar]
  30. Peters W. P., Shpall E. J., Jones R. B., Olsen G. A., Bast R. C., Gockerman J. P., Moore J. O. High-dose combination alkylating agents with bone marrow support as initial treatment for metastatic breast cancer. J Clin Oncol. 1988 Sep;6(9):1368–1376. doi: 10.1200/JCO.1988.6.9.1368. [DOI] [PubMed] [Google Scholar]
  31. Ross A. A., Cooper B. W., Lazarus H. M., Mackay W., Moss T. J., Ciobanu N., Tallman M. S., Kennedy M. J., Davidson N. E., Sweet D. Detection and viability of tumor cells in peripheral blood stem cell collections from breast cancer patients using immunocytochemical and clonogenic assay techniques. Blood. 1993 Nov 1;82(9):2605–2610. [PubMed] [Google Scholar]
  32. Seth P., Brinkmann U., Schwartz G. N., Katayose D., Gress R., Pastan Ira, Cowan K. Adenovirus-mediated gene transfer to human breast tumor cells: an approach for cancer gene therapy and bone marrow purging. Cancer Res. 1996 Mar 15;56(6):1346–1351. [PubMed] [Google Scholar]
  33. Shpall E. J., Bast R. C., Jr, Joines W. T., Jones R. B., Anderson I., Johnston C., Eggleston S., Tepperberg M., Edwards S., Peters W. P. Immunomagnetic purging of breast cancer from bone marrow for autologous transplantation. Bone Marrow Transplant. 1991 Feb;7(2):145–151. [PubMed] [Google Scholar]
  34. Shpall E. J., Jones R. B., Bast R. C., Jr, Rosner G. L., Vandermark R., Ross M., Affronti M. L., Johnston C., Eggleston S., Tepperburg M. 4-Hydroperoxycyclophosphamide purging of breast cancer from the mononuclear cell fraction of bone marrow in patients receiving high-dose chemotherapy and autologous marrow support: a phase I trial. J Clin Oncol. 1991 Jan;9(1):85–93. doi: 10.1200/JCO.1991.9.1.85. [DOI] [PubMed] [Google Scholar]
  35. Simpson S. J., Vachula M., Kennedy M. J., Kaizer H., Coon J. S., Ghalie R., Williams S., Van Epps D. Detection of tumor cells in the bone marrow, peripheral blood, and apheresis products of breast cancer patients using flow cytometry. Exp Hematol. 1995 Sep;23(10):1062–1068. [PubMed] [Google Scholar]
  36. Sutherland H. J., Lansdorp P. M., Henkelman D. H., Eaves A. C., Eaves C. J. Functional characterization of individual human hematopoietic stem cells cultured at limiting dilution on supportive marrow stromal layers. Proc Natl Acad Sci U S A. 1990 May;87(9):3584–3588. doi: 10.1073/pnas.87.9.3584. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Taswell C. Limiting dilution assays for the determination of immunocompetent cell frequencies. I. Data analysis. J Immunol. 1981 Apr;126(4):1614–1619. [PubMed] [Google Scholar]
  38. Tondini C., Pap S. A., Hayes D. F., Elias A. D., Kufe D. W. Evaluation of monoclonal antibody DF3 conjugated with ricin as a specific immunotoxin for in vitro purging of human bone marrow. Cancer Res. 1990 Feb 15;50(4):1170–1175. [PubMed] [Google Scholar]
  39. Vile R. G., Hart I. R. In vitro and in vivo targeting of gene expression to melanoma cells. Cancer Res. 1993 Mar 1;53(5):962–967. [PubMed] [Google Scholar]
  40. Vile R. G., Hart I. R. Use of tissue-specific expression of the herpes simplex virus thymidine kinase gene to inhibit growth of established murine melanomas following direct intratumoral injection of DNA. Cancer Res. 1993 Sep 1;53(17):3860–3864. [PubMed] [Google Scholar]
  41. Wickham T. J., Mathias P., Cheresh D. A., Nemerow G. R. Integrins alpha v beta 3 and alpha v beta 5 promote adenovirus internalization but not virus attachment. Cell. 1993 Apr 23;73(2):309–319. doi: 10.1016/0092-8674(93)90231-e. [DOI] [PubMed] [Google Scholar]
  42. Wu G. Y., Wilson J. M., Shalaby F., Grossman M., Shafritz D. A., Wu C. H. Receptor-mediated gene delivery in vivo. Partial correction of genetic analbuminemia in Nagase rats. J Biol Chem. 1991 Aug 5;266(22):14338–14342. [PubMed] [Google Scholar]
  43. Wu G. Y., Wu C. H. Receptor-mediated gene delivery and expression in vivo. J Biol Chem. 1988 Oct 15;263(29):14621–14624. [PubMed] [Google Scholar]
  44. Zhang W. W., Koch P. E., Roth J. A. Detection of wild-type contamination in a recombinant adenoviral preparation by PCR. Biotechniques. 1995 Mar;18(3):444–447. [PubMed] [Google Scholar]

Articles from Journal of Clinical Investigation are provided here courtesy of American Society for Clinical Investigation

RESOURCES