Cancer gene panel analysis of cultured circulating tumor cells and primary tumor tissue from patients with breast cancer

Hwang,Eunjoo; Uh,Ji‑Hyun; Lee,Hye  Seon; Lee,Cham  Han; Lee,Soo  Jeong; Ahn,Sei  Hyun; Son,Byung  Ho; Lee,Jong  Won; Yu,Jong  Han; Kwon,Nak‑Jung; Lee,Woo  Chung; Yang,Kap‑Seok; Choi,Sung  Ho; Kim,Myoung  Shin; Lee,Jinseon; Jeon,Byung  Hee

doi:10.3892/ol.2017.6077

Cancer gene panel analysis of cultured circulating tumor cells and primary tumor tissue from patients with breast cancer

Authors:
- Eunjoo Hwang
- Ji‑Hyun Uh
- Hye Seon Lee
- Cham Han Lee
- Soo Jeong Lee
- Sei Hyun Ahn
- Byung Ho Son
- Jong Won Lee
- Jong Han Yu
- Nak‑Jung Kwon
- Woo Chung Lee
- Kap‑Seok Yang
- Sung Ho Choi
- Myoung Shin Kim
- Jinseon Lee
- Byung Hee Jeon
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Affiliations: Cytogen Incorporated, Songpa, Seoul 138‑961, Republic of Korea, Department of Surgery, Asan Medical Center, College of Medicine, University of Ulsan, Songpa, Seoul 138‑736, Republic of Korea, Macrogen Incorporated, Geumcheon, Seoul 153‑781, Republic of Korea
Published online on: April 24, 2017 https://doi.org/10.3892/ol.2017.6077
Pages: 4627-4632
Copyright: © Hwang et al. This is an open access article distributed under the terms of Creative Commons Attribution License.

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Abstract

Although numerous effective therapies have improved the survival rate of patients with breast cancer, a number of patients present with tumor recurrence and metastasis. A liquid biopsy of circulating tumor cells (CTC) is a non‑invasive method to obtain tumor cells and may be used as substitute for a tumor tissue biopsy. The present study focuses on determining whether CTC culture is an optimal method of obtaining sufficient amounts of CTCs for molecular analysis. The current study demonstrates a method of isolating and culturing CTCs from patients with breast cancer and the construction of a molecular profile of cultured cells using the Ion AmpliSeq Cancer Gene Panel V2. Gene mutations that were observed in cultured CTCs were compared with those observed in primary tumor tissues. CTCs were isolated and cultured from the blood of six patients with breast cancer. Mutations from the Catalogue Of Somatic Mutation In Cancer (COSMIC) were detected in Platelet‑Derived Growth Factor Receptor Alpha, MET (also known as Hepatocyte Growth Factor Receptor), Phosphatase and Tensin Homolog, Harvey Rat Sarcoma Viral Oncogene Homolog, SWI/SNF Related, Matrix Associated, Actin Dependent Regulator of Chromatin Subfamily B Member 1, Cyclin Dependent Kinase Inhibitor 2A and MutL Homolog 1 genes in 5/6 samples. A comparison between mutations detected in cultured CTCs and mutations detected in primary tumor tissues demonstrated that a large number of mutations that were identified in CTCs were also detected in primary tumor tissues. The results from the current study describe a novel cell culture approach that may be used to obtain an optimal number of CTCs for molecular analysis. This novel approach is able to be used as a tool for liquid biopsy during breast cancer treatment.

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1	Ashworth TR: A case of cancer in which cells similar to those in the tumors were seen the blood after death. Aust Med J. 14:146–149. 1869.
2	Aceto N, Bardia A, Miyamoto DT, Donaldson MC, Wittner BS, Spencer JA, Yu M, Pely A, Engstrom A, Zhu H, et al: Circulating tumor cell clusters are oligoclonal precursors of breast cancer metastasis. Cell. 158:1110–1122. 2014. View Article : Google Scholar : PubMed/NCBI
3	Cristofanilli M, Hayes DF, Budd GT, Ellis MJ, Stopeck A, Reuben JM, Doyle GV, Matera J, Allard WJ, Miller MC, et al: Circulating tumor cells: A novel prognostic factor for newly diagnosed metastatic breast cancer. J Clin Oncol. 23:1420–1430. 2005. View Article : Google Scholar : PubMed/NCBI
4	Markiewicz A, Książkiewicz M, Wełnicka-Jaśkiewicz M, Seroczyńska B, Skokowski J, Szade J and Żaczek AJ: Mesenchymal phenotype of CTC-enriched blood fraction and lymph node metastasis formation potential. PLoS One. 9:e939012014. View Article : Google Scholar : PubMed/NCBI
5	Marchetti A, Del Grammastro M, Felicioni L, Malatesta S, Filice G, Centi I, de Pas T, Santoro A, Chella A, Brandes AA, et al: Assessment of EGFR mutations in circulating tumor cell preparations from NSCLC patients by next generation sequencing: Toward a real-time liquid biopsy for treatment. PLoS One. 9:e1038832014. View Article : Google Scholar : PubMed/NCBI
6	Marrinucci D, Bethel K, Luttgen M, Bruce RH, Nieva J and Kuhn P: Circulating tumor cells from well-differentiated lung adenocarcinoma retain cytomorphologic features of primary tumor type. Arch Pathol Lab Med. 133:1468–1471. 2009.PubMed/NCBI
7	van de Stolpe A, Pantel K, Sleijfer S, Terstappen LW and den Toonder JM: Circulating tumor cell isolation and diagnostics: Toward routine clinical use. Cancer Res. 71:5955–5960. 2011. View Article : Google Scholar : PubMed/NCBI
8	Giuliano M, Giordano A, Jackson S, de Giorgi U, Mego M, Cohen EN, Gao H, Anfossi S, Handy BC, Ueno NT, et al: Circulating tumor cells as early predictors of metastatic spread in breast cancer patients with limited metastatic dissemination. Brest Cancer Res. 16:4402014. View Article : Google Scholar
9	Torre LA, Bray F, Siegel RL, Ferlay J, Lortet-Tieulent J and Jemal A: Global cancer statistics, 2012. CA Cancer J Clin. 65:87–108. 2015. View Article : Google Scholar : PubMed/NCBI
10	Lorusso G and Rüegg C: New insights into the mechanisms of organ-specific breast cancer metastasis. Semin Cancer Biol. 22:226–233. 2012. View Article : Google Scholar : PubMed/NCBI
11	Cardoso F, Harbeck N, Fallowfield L, Kyriakides S and Senkus E; ESMO Guidelines Working Group, : Locally recurrent or metastatic breast cancer: ESMO clinical practice guidelines for diagnosis, treatment and follow-up. Ann Oncol. 23 Suppl 7:vii11–vii19. 2012. View Article : Google Scholar : PubMed/NCBI
12	Suzuki M and Tarin D: Gene expression profiling of human lymph node metastases and matched primary breast carcinomas: Clinical implications. Mol Oncol. 1:172–180. 2007. View Article : Google Scholar : PubMed/NCBI
13	Giuliano M, Giordano A, Jackson S, de Giorgi U, Mego M, Cohen EN, Gao H, Anfossi S, Handy BC, Ueno NT, et al: Circulating tumor cells as early predictors of metastatic spread in breast cancer patients with limited metastatic dissemination. Breast Cancer Res. 16:4402014. View Article : Google Scholar : PubMed/NCBI
14	Rack B, Schindlbeck C, Jückstock J, Andergassen U, Hepp P, Zwingers T, Friedl TW, Lorenz R, Tesch H, et al: Circulating tumor cells predict survival in early average-to-high risk breast cancer patients. J Natl Cancer Inst. 106:pii: dju066. 2014. View Article : Google Scholar : PubMed/NCBI
15	Bidard FC, Fehm T, Ignatiadis M, Smerage JB, Alix-Panabières C, Janni W, Messina C, Paoletti C, Müller V, Hayes DF, et al: Clinical application of circulating tumor cells in breast cancer: Overview of the current interventional trials. Cancer Metastasis Rev. 32:179–188. 2013. View Article : Google Scholar : PubMed/NCBI
16	Nadal R, Lorente JA, Rosell R and Serrano MJ: Relevance of molecular characterization of circulating tumor cells in breast cancer in the era of targeted therapies. Expert Rev Mol Diagn. 13:295–307. 2013. View Article : Google Scholar : PubMed/NCBI
17	Franken B, de Groot MR, Mastboom WJ, Vermes I, van der Palen J, Tibbe AG and Terstappen LW: Circulating tumor cells, disease recurrence and survival in newly diagnosed breast cancer. Breast Cancer Res. 14:R1332012. View Article : Google Scholar : PubMed/NCBI
18	Cristofanilli M, Budd GT, Ellis MJ, Stopeck A, Matera J, Miller MC, Reuben JM, Doyle GV, Allard WJ, Terstappen LW and Hayes DF: Circulating tumor cells, disease progression, and survival in metastatic breast cancer. N Engl J Med. 351:781–791. 2004. View Article : Google Scholar : PubMed/NCBI
19	Fernandez SV, Bingham C, Fittipaldi P, Austin L, Palazzo J, Palmer G, Alpaugh K and Cristofanilli M: TP53 mutations detected in circulating tumor cells present in the blood of metastatic triple negative breast cancer patients. Breast Cancer Res. 16:4452014. View Article : Google Scholar : PubMed/NCBI
20	Yu M, Stott S, Toner M, Maheswaran S and Haber DA: Circulating tumor cells: Approaches to isolation and characterization. J Cell Biol. 192:373–382. 2011. View Article : Google Scholar : PubMed/NCBI
21	Edge SB and Compton CC: The American Joint Committee on Cancer: The 7th Edition of the AJCC Cancer Staging Manual and the Future of TNM. Ann Surg Oncol. 17:14712010. View Article : Google Scholar : PubMed/NCBI
22	Kim EH, Lee JK, Kim BC, Rhim SH, Kim JW, Kim KH, Jung SM, Park PS, Park HC, Lee J and Jeon BH: Enrichment of cancer cells from whole blood using a microfabricated porous filter. Anal Biochem. 440:114–116. 2013. View Article : Google Scholar : PubMed/NCBI
23	Gorges TM, Tinhofer I, Drosch M, Röse L, Zollner TM, Krahn T and von Ahsen O: Circulating tumour cells escape from EpCAM based detection due to epithelial-to mesenchymal transition. BMC Cancer. 12:1782012. View Article : Google Scholar : PubMed/NCBI
24	Tung N, Battelli C, Allen B, Kaldate R, Bhatnagar S, Bowles K, Timms K, Garber JE, Herold C, Ellisen L, et al: Frequency of mutations in individuals with breast cancer referred for BRCA1 and BRCA2 testing using next-generation sequencing with a 25-gene panel. Cancer. 121:25–33. 2015. View Article : Google Scholar : PubMed/NCBI
25	Inês C, Fernanda M, Albino M, Rui MR and Fernando S: Overexpression of platelet-derived growth factor receptor alpha in breast cancer is associated with tumour progression. Breast Cancer Res. 7:R788–R795. 2005. View Article : Google Scholar : PubMed/NCBI
26	Fernández-Medarde A and Santos E: Ras in cancer and developmental diseases. Genes Cancer. 2:344–358. 2011. View Article : Google Scholar : PubMed/NCBI
27	Yong HY, Hwang JS, Son H, Park HI, Oh ES, Kim HH, Kim DK, Choi WS, Lee BJ, Kim HR and Moon A: Identification of H-Ras-specific motif for the activation of invasive signaling program in human breast epithelial cells. Neoplasia. 13:98–107. 2011. View Article : Google Scholar : PubMed/NCBI
28	Watson DM, Elton RA, Jack WJ, Dixon JM, Chetty U and Miller WR: The H-ras oncogene product p21 and prognosis in human breast cancer. Breast Cancer Res Treat. 17:161–169. 1991. View Article : Google Scholar : PubMed/NCBI