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A sensitive polymerase chain reaction-based method for detection and quantification of metastasis in human xenograft mouse models

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Abstract

Tumor cell dissemination to distant organs accounts for the majority of cancer related deaths. Analysis of the stepwise process of metastasis formation and progression might provide novel therapeutic strategies for the treatment of disseminated cancer. However, studies with both biological and therapeutic endpoints would require highly sensitive and specific methods for precise quantification of the metastatic tumor burden in vivo. We have developed a quantitative real-time PCR-based assay for the detection and quantification of human tumor cells disseminated in mouse organs. The method relies on the parallel amplification of unique, species-specific, conserved and non-transcribed sequences in the mouse and human genomes. We tested the method in xenograft models to assess the metastatic potential of various cancer cell lines, the impact of injection modality and cell type on organ distribution, and the early stages of metastasis implantation and progression. With this method, we observed clear quantitative differences among colon cancer cell lines in terms of metastasis formation in the lung, consistent with the different in vitro growth properties. The mode of cell implantation and cell intrinsic properties strongly affected the metastatic pattern of prostate and breast cancer cell lines in mouse organs. The qPCR assay accurately determined the malignant cell burden even at early stages of metastasis progression in the lung. We describe a very sensitive assay for the highly reproducible detection and accurate quantification of human metastatic cells in mouse tissues and demonstrate its broad applicability to various experimental settings.

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References

  1. Hanahan D, Weinberg RA (2000) The hallmarks of cancer. Cell 100(1):57–70

    Article  CAS  PubMed  Google Scholar 

  2. Eccles SA, Welch DR (2007) Metastasis: recent discoveries and novel treatment strategies. Lancet 369(9574):1742–1757

    Article  CAS  PubMed  Google Scholar 

  3. Nguyen DX, Bos PD, Massague J (2009) Metastasis: from dissemination to organ-specific colonization. Nat Rev Cancer 9(4):274–284

    Article  CAS  PubMed  Google Scholar 

  4. Joyce JA, Pollard JW (2009) Microenvironmental regulation of metastasis. Nat Rev Cancer 9(4):239–252

    Article  CAS  PubMed  Google Scholar 

  5. Ramaswamy S, Ross KN, Lander ES et al (2003) A molecular signature of metastasis in primary solid tumors. Nat Genet 33(1):49–54

    Article  CAS  PubMed  Google Scholar 

  6. Giovanella BC, Yim SO, Stehlin JS et al (1972) Development of invasive tumors in the “nude” mouse after injection of cultured human melanoma cells. J Natl Cancer Inst 48(5):1531–1533

    CAS  PubMed  Google Scholar 

  7. Minn AJ, Gupta GP, Siegel PM et al (2005) Genes that mediate breast cancer metastasis to lung. Nature 436(7050):518–524

    Article  CAS  PubMed  Google Scholar 

  8. Bandyopadhyay A, Elkahloun A, Baysa SJ et al (2005) Development and gene expression profiling of a metastatic variant of the human breast cancer MDA-MB-435 cells. Cancer Biol Ther 4(2):168–174

    Article  CAS  PubMed  Google Scholar 

  9. Kang Y, Siegel PM, Shu W et al (2003) A multigenic program mediating breast cancer metastasis to bone. Cancer Cell 3(6):537–549

    Article  CAS  PubMed  Google Scholar 

  10. Minn AJ, Kang Y, Serganova I et al (2005) Distinct organ-specific metastatic potential of individual breast cancer cells and primary tumors. J Clin Investig 115(1):44–55

    CAS  PubMed  Google Scholar 

  11. Hedley BD, Vaidya KS, Phadke P et al (2008) BRMS1 suppresses breast cancer metastasis in multiple experimental models of metastasis by reducing solitary cell survival and inhibiting growth initiation. Clin Exp Metastasis 25(7):727–740

    Article  CAS  PubMed  Google Scholar 

  12. Chu JH, Sun ZY, Meng XL et al (2006) Differential metastasis-associated gene analysis of prostate carcinoma cells derived from primary tumor and spontaneous lymphatic metastasis in nude mice with orthotopic implantation of PC-3M cells. Cancer Lett 233(1):79–88

    Article  CAS  PubMed  Google Scholar 

  13. Lee H, Lin EC, Liu L et al (2003) Gene expression profiling of tumor xenografts: in vivo analysis of organ-specific metastasis. Int J Cancer 107(4):528–534

    Article  CAS  PubMed  Google Scholar 

  14. Nitsche A, Becker M, Junghahn I et al (2001) Quantification of human cells in NOD/SCID mice by duplex real-time polymerase-chain reaction. Haematologica 86(7):693–699

    CAS  PubMed  Google Scholar 

  15. Becker M, Nitsche A, Neumann C et al (2002) Sensitive PCR method for the detection and real-time quantification of human cells in xenotransplantation systems. Br J Cancer 87(11):1328–1335

    Article  CAS  PubMed  Google Scholar 

  16. Zubair AC, Ali SA, Rees RC et al (1996) Investigation of the effect of BB-94 (batimastat) on the colonization potential of human lymphoma cells in SCID mice. Cancer Lett 107(1):91–95

    Article  CAS  PubMed  Google Scholar 

  17. Kim J, Yu W, Kovalski K et al (1998) Requirement for specific proteases in cancer cell intravasation as revealed by a novel semiquantitative PCR-based assay. Cell 94(3):353–362

    Article  CAS  PubMed  Google Scholar 

  18. Schneider T, Osl F, Friess T et al (2002) Quantification of human Alu sequences by real-time PCR–an improved method to measure therapeutic efficacy of anti-metastatic drugs in human xenotransplants. Clin Exp Metastasis 19(7):571–582

    Article  CAS  PubMed  Google Scholar 

  19. Mira E, Lacalle RA, Gomez-Mouton C et al (2002) Quantitative determination of tumor cell intravasation in a real-time polymerase chain reaction-based assay. Clin Exp Metastasis 19(4):313–318

    Article  CAS  PubMed  Google Scholar 

  20. Bennett EA, Keller H, Mills RE et al (2008) Active Alu retrotransposons in the human genome. Genome Res 18(12):1875–1883

    Article  CAS  PubMed  Google Scholar 

  21. Eckhardt BL, Parker BS, van Laar RK et al (2005) Genomic analysis of a spontaneous model of breast cancer metastasis to bone reveals a role for the extracellular matrix. Mol Cancer Res 3(1):1–13

    CAS  PubMed  Google Scholar 

  22. Lombello CB, Malmonge SM, Wada ML (2000) PolyHEMA and polyHEMA-poly(MMA-co-AA) as substrates for culturing Vero cells. J Mater Sci Mater Med 11(9):541–546

    Article  CAS  PubMed  Google Scholar 

  23. Liang CC, Park AY, Guan JL (2007) In vitro scratch assay: a convenient and inexpensive method for analysis of cell migration in vitro. Nat Protoc 2(2):329–333

    Article  CAS  PubMed  Google Scholar 

  24. Franken NA, Rodermond HM, Stap J et al (2006) Clonogenic assay of cells in vitro. Nat Protoc 1(5):2315–2319

    Article  CAS  PubMed  Google Scholar 

  25. Levin LV, Griffin TW (1991) Specific adhesion of carcinoembryonic antigen-bearing colorectal cancer cells to immobilized carcinoembryonic antigen. Cancer Lett 60(2):143–152

    Article  CAS  PubMed  Google Scholar 

  26. Calleros L, Sánchez-Hernández I, Baquero P et al (2009) Oncogenic Ras, but not (V600E)B-RAF, protects from cholesterol depletion-induced apoptosis through the PI3K/AKT pathway in colorectal cancer cells. Carcinogenesis 30(10):1670–1677

    Article  CAS  PubMed  Google Scholar 

  27. Yao Y, Jia XY, Tian HY et al (2009) Comparative proteomic analysis of colon cancer cells in response to oxaliplatin treatment. Biochim Biophys Acta 1794(10):1433–1440

    CAS  PubMed  Google Scholar 

  28. Serganova I, Moroz E, Vider E et al (2009) Multimodality imaging of TGF beta signaling in breast cancer metastases. FASEB J 23:2662–2672

    Article  CAS  PubMed  Google Scholar 

  29. Zhang XH-F, Wang Q, Gerald W et al (2009) Latent bone metastasis in breast cancer tied to Src-dependent survival signals. Cancer Cell 16(1):67–78

    Article  CAS  PubMed  Google Scholar 

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Acknowledgments

This work was supported by a grant from the Deutsche Forschungsgemeinschaft (DFG Forschergruppe 627 ‘Nanohale’) to F.C. and A.A. (AI 24/6-1), and grants from the Canton Ticino DECS and the Fondazione Ticinese Ricerca sul Cancro to C.V.C.

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Authors and Affiliations

  1. Laboratory of Experimental Oncology, Oncology Institute of Southern Switzerland, 6500, Bellinzona, Switzerland

    Anastasia Malek & Carlo V. Catapano

  2. Department of Pharmacology and Toxicology, Philipps-University, School of Medicine, Marburg, Germany

    Frank Czubayko & Achim Aigner

Authors
  1. Anastasia Malek
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  2. Carlo V. Catapano
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  3. Frank Czubayko
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  4. Achim Aigner
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Correspondence to Anastasia Malek.

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Malek, A., Catapano, C.V., Czubayko, F. et al. A sensitive polymerase chain reaction-based method for detection and quantification of metastasis in human xenograft mouse models. Clin Exp Metastasis 27, 261–271 (2010). https://doi.org/10.1007/s10585-010-9324-1

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  • DOI: https://doi.org/10.1007/s10585-010-9324-1

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