Abstract
Small-cell lung cancer (SCLC), an aggressive neuroendocrine tumor with early dissemination and dismal prognosis, accounts for 15–20% of lung cancer cases and ∼200,000 deaths each year. Most cases are inoperable, and biopsies to investigate SCLC biology are rarely obtainable. Circulating tumor cells (CTCs), which are prevalent in SCLC, present a readily accessible 'liquid biopsy'. Here we show that CTCs from patients with either chemosensitive or chemorefractory SCLC are tumorigenic in immune-compromised mice, and the resultant CTC-derived explants (CDXs) mirror the donor patient's response to platinum and etoposide chemotherapy. Genomic analysis of isolated CTCs revealed considerable similarity to the corresponding CDX. Most marked differences were observed between CDXs from patients with different clinical outcomes. These data demonstrate that CTC molecular analysis via serial blood sampling could facilitate delivery of personalized medicine for SCLC. CDXs are readily passaged, and these unique mouse models provide tractable systems for therapy testing and understanding drug resistance mechanisms.
This is a preview of subscription content, access via your institution
Access options
Subscribe to this journal
Receive 12 print issues and online access
$209.00 per year
only $17.42 per issue
Buy this article
- Purchase on Springer Link
- Instant access to full article PDF
Prices may be subject to local taxes which are calculated during checkout
Similar content being viewed by others
References
Einhorn, L.H., Fee, W.H., Farber, M.O., Livingston, R.B. & Gottlieb, J.A. Improved chemotherapy for small-cell undifferentiated lung cancer. J. Am. Med. Assoc. 235, 1225–1229 (1976).
Evans, W.K. et al. VP-16 and cisplatin as first-line therapy for small-cell lung cancer. J. Clin. Oncol. 3, 1471–1477 (1985).
Sierocki, J.S. et al. cis-Dichlorodiammineplatinum(II) and VP-16–213: an active induction regimen for small cell carcinoma of the lung. Cancer Treat. Rep. 63, 1593–1597 (1979).
Gazdar, A.F. et al. Establishment of continuous, clonable cultures of small-cell carcinoma of lung which have amine precursor uptake and decarboxylation cell properties. Cancer Res. 40, 3502–3507 (1980).
Oboshi, S., Tsugawa, S., Seido, T., Shimosato, Y. & Koide, T. A new floating cell line derived from human pulmonary carcinoma of oat cell type. Gann 62, 505–514 (1971).
Pleasance, E.D. et al. A small-cell lung cancer genome with complex signatures of tobacco exposure. Nature 463, 184–190 (2010).
Joshi, M., Ayoola, A. & Belani, C.P. Small-cell lung cancer: an update on targeted therapies. Adv. Exp. Med. Biol. 779, 385–404 (2013).
William, W.N. Jr. & Glisson, B.S. Novel strategies for the treatment of small-cell lung carcinoma. Nat. Rev. Clin. Oncol. 8, 611–619 (2011).
Meuwissen, R. et al. Induction of small cell lung cancer by somatic inactivation of both Trp53 and Rb1 in a conditional mouse model. Cancer Cell 4, 181–189 (2003).
Kwon, M.C. & Berns, A. Mouse models for lung cancer. Mol. Oncol. 7, 165–177 (2013).
Daniel, V.C. et al. A primary xenograft model of small-cell lung cancer reveals irreversible changes in gene expression imposed by culture in vitro. Cancer Res. 69, 3364–3373 (2009).
Poupon, M.F. et al. Response of small-cell lung cancer xenografts to chemotherapy: multidrug resistance and direct clinical correlates. J. Natl. Cancer Inst. 85, 2023–2029 (1993).
Davenport, R.D. Diagnostic value of crush artifact in cytologic specimens. Occurrence in small cell carcinoma of the lung. Acta Cytol. 34, 502–504 (1990).
Peifer, M. et al. Integrative genome analyses identify key somatic driver mutations of small-cell lung cancer. Nat. Genet. 44, 1104–1110 (2012).
Rudin, C.M. et al. Comprehensive genomic analysis identifies SOX2 as a frequently amplified gene in small-cell lung cancer. Nat. Genet. 44, 1111–1116 (2012).
Cristofanilli, M. et al. Circulating tumor cells, disease progression, and survival in metastatic breast cancer. N. Engl. J. Med. 351, 781–791 (2004).
de Bono, J.S. et al. Circulating tumor cells predict survival benefit from treatment in metastatic castration-resistant prostate cancer. Clin. Cancer Res. 14, 6302–6309 (2008).
Hayes, D.F. et al. Circulating tumor cells at each follow-up time point during therapy of metastatic breast cancer patients predict progression-free and overall survival. Clin. Cancer Res. 12, 4218–4224 (2006).
Hou, J.M. et al. Clinical significance and molecular characteristics of circulating tumor cells and circulating tumor microemboli in patients with small-cell lung cancer. J. Clin. Oncol. 30, 525–532 (2012).
Krebs, M.G. et al. Evaluation and prognostic significance of circulating tumor cells in patients with non-small-cell lung cancer. J. Clin. Oncol. 29, 1556–1563 (2011).
Hou, J.M. et al. Evaluation of circulating tumor cells and serological cell death biomarkers in small cell lung cancer patients undergoing chemotherapy. Am. J. Pathol. 175, 808–816 (2009).
Evans, W.K. et al. VP-16 alone and in combination with cisplatin in previously treated patients with small cell lung cancer. Cancer 53, 1461–1466 (1984).
Swanton, C. Intratumor heterogeneity: evolution through space and time. Cancer Res. 72, 4875–4882 (2012).
Martinez, P. et al. Parallel evolution of tumour subclones mimics diversity between tumours. J. Pathol. 230, 356–364 (2013).
Arriola, E. et al. Genetic changes in small cell lung carcinoma. Clin. Transl. Oncol. 10, 189–197 (2008).
Mori, N. et al. Variable mutations of the RB gene in small-cell lung carcinoma. Oncogene 5, 1713–1717 (1990).
Wistuba, I.I. & Gazdar, A.F. & Minna, J.D. Molecular genetics of small cell lung carcinoma. Semin. Oncol. 28, 3–13 (2001).
Baccelli, I. et al. Identification of a population of blood circulating tumor cells from breast cancer patients that initiates metastasis in a xenograft assay. Nat. Biotechnol. 31, 539–544 (2013).
Allard, W.J. et al. Tumor cells circulate in the peripheral blood of all major carcinomas but not in healthy subjects or patients with nonmalignant diseases. Clin. Cancer Res. 10, 6897–6904 (2004).
Pretlow, T.G. et al. Prostate cancer and other xenografts from cells in peripheral blood of patients. Cancer Res. 60, 4033–4036 (2000).
Ni, X. et al. Reproducible copy number variation patterns among single circulating tumor cells of lung cancer patients. Proc. Natl. Acad. Sci. USA 110, 21083–21088 (2013).
Klein, C.A. Selection and adaptation during metastatic cancer progression. Nature 501, 365–372 (2013).
Gasch, C. et al. Heterogeneity of epidermal growth factor receptor status and mutations of KRAS/PIK3CA in circulating tumor cells of patients with colorectal cancer. Clin. Chem. 59, 252–260 (2013).
Fabbri, F. et al. Detection and recovery of circulating colon cancer cells using a dielectrophoresis-based device: KRAS mutation status in pure CTCs. Cancer Lett. 335, 225–231 (2013).
Heitzer, E. et al. Complex tumor genomes inferred from single circulating tumor cells by array-CGH and next-generation sequencing. Cancer Res. 73, 2965–2975 (2013).
Klein, C.A. et al. Genetic heterogeneity of single disseminated tumour cells in minimal residual cancer. Lancet 360, 683–689 (2002).
Klein, C.A. et al. Combined transcriptome and genome analysis of single micrometastatic cells. Nat. Biotechnol. 20, 387–392 (2002).
Morelli, M.P. et al. Prioritizing phase I treatment options through preclinical testing on personalized tumorgraft. J. Clin. Oncol. 30, e45–e48 (2012).
Board, R.E. et al. Isolation and extraction of circulating tumor DNA from patients with small cell lung cancer. Ann. NY Acad. Sci. 1137, 98–107 (2008).
Bettegowda, C. et al. Detection of circulating tumor DNA in early- and late-stage human malignancies. Sci .Transl. Med. 6, 224ra224 (2014).
Dawson, S.J. et al. Analysis of circulating tumor DNA to monitor metastatic breast cancer. N. Engl. J. Med. 368, 1199–1209 (2013).
Krebs, M.G. et al. Molecular analysis of circulating tumour cells—biology and biomarkers. Nat. Rev. Clin. Oncol. 11, 129–144 (2014).
Lohmann, D.R. et al. Constitutional RB1-gene mutations in patients with isolated unilateral retinoblastoma. Am. J. Hum. Genet. 61, 282–294 (1997).
Szijan, I., Lohmann, D.R., Parma, D.L., Brandt, B. & Horsthemke, B. Identification of RB1 germline mutations in Argentinian families with sporadic bilateral retinoblastoma. J. Med. Genet. 32, 475–479 (1995).
Joerger, A.C. & Fersht, A.R. The tumor suppressor p53: from structures to drug discovery. Cold Spring Harb. Perspect. Biol. 2, a000919 (2010).
Alcoser, S.Y. et al. Real-time PCR-based assay to quantify the relative amount of human and mouse tissue present in tumor xenografts. BMC Biotechnol. 11, 124 (2011).
Thierry, A.R. et al. Origin and quantification of circulating DNA in mice with human colorectal cancer xenografts. Nucleic Acids Res. 38, 6159–6175 (2010).
Peeters, D.J. et al. Semiautomated isolation and molecular characterisation of single or highly purified tumour cells from CellSearch enriched blood samples using dielectrophoretic cell sorting. Br. J. Cancer 108, 1358–1367 (2013).
McKenna, A. et al. The Genome Analysis Toolkit: a MapReduce framework for analyzing next-generation DNA sequencing data. Genome Res. 20, 1297–1303 (2010).
DePristo, M.A. et al. A framework for variation discovery and genotyping using next-generation DNA sequencing data. Nat. Genet. 43, 491–498 (2011).
Wang, K., Li, M. & Hakonarson, H. ANNOVAR: functional annotation of genetic variants from high-throughput sequencing data. Nucleic Acids Res. 38, e164 (2010).
Boeva, V. et al. Control-free calling of copy number alterations in deep-sequencing data using GC-content normalization. Bioinformatics 27, 268–269 (2011).
Krzywinski, M. et al. Circos: an information aesthetic for comparative genomics. Genome Res. 19, 1639–1645 (2009).
Yates, T., Okoniewski, M.J. & Miller, C.J. X:Map: annotation and visualization of genome structure for Affymetrix exon array analysis. Nucleic Acids Res. 36, D780–D786 (2008).
Acknowledgements
We are indebted to the patients who agreed to donate their blood samples for this study. We thank R. Marais, N. Jones and D. Ogilvie for their constructive comments on the manuscript. We thank M. Dawson, M. Lancashire, S. Bramley, J. Halstead and J. Castle, who enumerated CTCs using CellSearch. We thank A. Jardine for administrative support and M. Greaves, our laboratory manager. This research was supported by Cancer Research UK via core funding to the Cancer Research UK Manchester Institute (C5759/A12328), the Manchester Experimental Cancer Medicine Centre (C1467/A15578), the Manchester Cancer Research Centre (A12197) and their Translational Research Award for 2012. Funding to support this work was also provided via the European Union CHEMORES FP6 (contract number LSHG-CT-2007-037665). R.L.M. and L.C. were supported by education grants from Cancer Research UK and AstraZeneca.
Author information
Authors and Affiliations
Contributions
C.L.H., P.K. and B.B. performed in vivo studies, F.T., R.P., K.L.S. and D.N. conducted histopathological examinations, D.G.R., D.J.B., S.D.P., A.G., J.A., M.G.K., M.A., L.C. and S.F. conducted the genomic analyses, Y.L., C.T., C.J. Miller and G.B. performed the bioinformatic analysis, K.M. oversaw CTC enumeration by CellSearch, R.L.M., L.C., L.P. and F.B. recruited and consented patients and collected blood samples, C.J. Morrow, C.J. Miller, G.B., F.B. and C.D. conceived and directed the study, interpreted the data and wrote the manuscript. All authors discussed the results and commented on the manuscript.
Corresponding author
Ethics declarations
Competing interests
The authors declare no competing financial interests.
Supplementary information
Supplementary Text and Figures
Supplementary Figures 1–4 and Supplementary Tables 1–7 (PDF 432 kb)
Source data
Rights and permissions
About this article
Cite this article
Hodgkinson, C., Morrow, C., Li, Y. et al. Tumorigenicity and genetic profiling of circulating tumor cells in small-cell lung cancer. Nat Med 20, 897–903 (2014). https://doi.org/10.1038/nm.3600
Received:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1038/nm.3600