JCT  Vol.3 No.4 A , September 2012
Therapy-Induced Changes of Gene Expression in a Matched Pair of Small Cell Lung Cancer (SCLC) Cell Lines
Abstract: Extended stage small cell lung cancer (SCLC) responds to platinum/vepeside-based first-line chemotherapy but relapses early as drug-resistant tumor associated with a dismal prognosis. A pair of SCLC cell lines obtained from a single patient at different time points during treatment allows for the investigation of the changes in gene expression prior to (GLC14) and following cycles of chemotherapy and irradiation (GLC19). GLC19 cells were reported to reveal an increased doubling time and exhibit increased chemoresistance to doxorubicin, etoposide, melphalan and vinblastine. Upregulated transcripts in GLC19, as assessed by microarray analysis, comprised proteins involved in regulation of cellular growth (NGFRAP1/BEX3), adhesion, glutathione metabolism and, in particular, WNT/Notch pathways and the putative cancer stem cell phenotype (CD44, ALDH1A1, and AKR1C1/13). Metallothioneins, tubulins TUBA3/4 and tumor protein p53 inducible protein 11 (TP53IP11) were downregulated in this cell line compared to GLC14. Except increased expression of glutathione transferases no classical markers of chemoresistance were found, pointing to a role of altered growth control/differentiation and reduced accessibility of this SCLC tumor cells growing as multicellular spheroids. In conclusion, treatment of this single SCLC with cyclophosphamide, doxorubicin and etoposide (CDE) followed by radiotherapy ultimately resulted in an enrichment of tumor cells displaying the typical signature of tumor-initiating or cancer stem cells (CIC/CSC).
Cite this paper: G. Hamilton, E. Ulsperger, K. Geissler and U. Olszewski, "Therapy-Induced Changes of Gene Expression in a Matched Pair of Small Cell Lung Cancer (SCLC) Cell Lines," Journal of Cancer Therapy, Vol. 3 No. 4, 2012, pp. 442-451. doi: 10.4236/jct.2012.324057.

[1]   D. R. Youlden, S. M. Cramb and P. D. Baade, “The International Epidemiology of Lung Cancer: Geographical Distribution and Secular Trends,” Journal of Thoracic Oncology, Vol. 3, No. 8, 2008, pp. 819-831. doi:10.1097/JTO.0b013e31818020eb

[2]   S. G. Spiro and G. A. Silvestri, “One Hundred Years of Lung Cancer,” American Journal of Respiratory Critical Care Medicine, Vol. 172, No. 5, 2005, pp. 523-529. doi:10.1164/rccm.200504-531OE

[3]   R. Califano, A. Z. Abidin, R. Peck, C. Faivre-Finn and P. Lorigan, “Management of Small Cell Lung Cancer: Recent Developments for Optimal Care,” Drugs, Vol. 72, No. 4, 2012, pp. 471-490. doi:10.2165/11597640-000000000-00000

[4]   R. Govindan, N. Page, D. Morgensztern, et al., “Changing Epidemiology of Small-Cell Lung Cancer in the United States over the Last 30 Years: Analysis of the Surveillance, Epidemiologic, and End Results Database,” Journal of Clinical Oncology, Vol. 24, 2006, pp. 4539-4544. doi:10.1200/JCO.2005.04.4859

[5]   W. N. William and B. S. Glisson, “Novel Strategies for the Treatment of Small-Cell Lung Carcinoma, “Nature Reviews Clinical Oncology, Vol. 8, No. 10, 2011, pp. 611-619. doi:10.1038/nrclinonc.2011.90

[6]   Y. Agra, M. Pelayo, M. Sacristan, et al., “Chemotherapy versus Best Supportive Care for Extensive Small Cell Lung Cancer,” Cochrane Database Systematic Review, Vol. 4, 2003, Article ID: CD001990.

[7]   M. Takada, M. Fukuoka, M. Kawahara, et al., “Phase III Study of Concurrent versus Sequential Thoracic Radiotherapy in Combination with Cisplatin and Etoposide for Limited-Stage Small-Cell Lung Cancer,” Journal of Clinical Oncology, Vol. 20, 2002, pp. 3054-3060. doi:10.1200/JCO.2002.12.071

[8]   B. Fischer and A. Arcaro, “Current Status of Clinical Trials for Small Cell Lung Cancer,” Review of Recent Clinical Trials, Vol. 3, 2008, pp. 40-61. doi:10.2174/157488708783330503

[9]   H. H. Berendsen, L. de Leij, E. G. de Vries, G. Mesander, N. H. Mulder, B. de Jong, C. H. Buys, P. E. Postmus, S. Poppema, H. J. Sluiter, et al., “Characterization of Three Small Cell Lung Cancer Cell Lines Established from One Patient during Longitudinal Follow-Up,” Cancer Research, Vol. 48, No. 23, 1988, pp. 6891-6899.

[10]   V. G. Tusher, R. Tibshirani and G. Chu, “Significance Analysis of Microarrays Applied to the Ionizing Radiation Response,” Proceedings of the National Academy of Science U S A., Vol. 98, No. 9, 2001, pp. 5116-5121. doi:10.1073/pnas.091062498

[11]   D. Croft, G. O’Kelly, G. Wu, R. Haw, M. Gillespie, L. Matthews, M. Caudy, P. Garapati, G. Gopinath, B. Jassal, S. Jupe, I. Kalatskaya, S. Mahajan, B. May, N. Ndegwa, E. Schmidt, V. Shamovsky, C. Yung, E. Birney, H. Hermjakob, P. D’Eustachio and L. Stein, “Reactome: A Database of Reactions, Pathways and Biological Processes,” Nucleic Acids Research, Vol. 39, 2011, pp. D691-D697. doi:10.1093/nar/gkq1018

[12]   E. G. de Vries, C. Meijer, H. Timmer-Bosscha, H. H. Berendsen, L. de Leij, R. J. Scheper and N. H. Mulder, “Resistance Mechanisms in Three Human Small Cell Lung Cancer Cell Lines Established from One Patient during Clinical Follow-Up,” Cancer Research, Vol. 49, No. 15, 1989, pp. 4175-4178.

[13]   G. Rapp, J. Freudenstein, J. Klaudiny, J. Mucha, F. Wempe, M. Zimmer and K. H. Scheit, “Characterization of Three Abundant mRNAs from Human Ovarian Granulosa Cells,” DNA Cell Biology, Vol. 9, No. 7, 1990, pp. 479-485.doi:10.1089/dna.1990.9.479

[14]   M. H. Lawson, N. M. Cummings, D. M. Rassl, R. Russell, J. D. Brenton, R. C. Rintoul and G. Murphy, “Two Novel Determinants of Etoposide Resistance in Small Cell Lung Cancer,” Cancer Research, Vol. 71, No. 14, 2011, pp. 4877-4887. doi:10.1158/0008-5472.CAN-11-0080

[15]   A. Naderi, A. E. Teschendorff, J. Beigel, M. Cariati, I. O. Ellis, J. D. Brenton and C. Caldas, “BEX2 Is Overexpressed in a Subset of Primary Breast Cancers and Mediates Nerve Growth Factor/Nuclear Factor-KappaB Inhibition of Apoptosis in Breast Cancer Cell Lines,” Cancer Research, Vol. 67, No. 14, 2007, pp. 6725-6736. doi:10.1158/0008-5472.CAN-06-4394

[16]   N. H. Molloy, D. E. Read and A. M. Gorman, “Nerve Growth Factor in Cancer Cell Death and Survival,” Cancers, Vol. 3, 2011, pp. 510-530. doi:10.3390/cancers3010510

[17]   X. Tong, D. Xie, W. Roth, J. Reed, H. P. Koeffler, “NADE (p75NTR-Associated Cell Death Executor) Suppress Cellular Growth in Vivo,” International Journal of Oncology, Vol. 22, 2003, pp. 1357-1362.

[18]   T. L. Romanuik, T. Ueda, N. Le, S. Haile, T. M. Yong, T. Thomson, R. L. Vessella and M. D. Sadar, “NGFRAP1 Novel Biomarkers for Prostate Cancer Including Noncoding Transcripts,” American Journal of Pathology, Vol. 175, No. 6, 2006, pp. 2264-2276. doi:10.2353/ajpath.2009.080868

[19]   Y. P. Ching, A. S. Pang, W. H. Lam, R. Z. Qi and J. H. Wang, “Identification of a Neuronal Cdk5 Activator- Binding Protein as Cdk5 Inhibitor,” Journal of Biological Chemistry, Vol. 277, No. 18, 2002, pp. 15237-15240. doi:10.1074/jbc.C200032200

[20]   K. Kok, J. Osinga, D. C. Schotanus, H. H. Berendsen, L. F. de Leij and C. H. Buys, “Amplification and Expression of Different Myc-Family Genes in a Tumor Specimen and 3 Cell Lines Derived from One Small-Cell Lung Cancer Patient during Longitudinal Follow-Up,” International Journal of Cancer, Vol. 44, No. 1, 1989, pp. 75-78. doi:10.1002/ijc.2910440114

[21]   I. I. Wistuba, A. F. Gazdar and J. D. Minna, “Molecular Genetics of Small Cell Lung Carcinoma,” Seminars in Oncology, Vol. 28, No. 2-4, 2001, pp. 3-13.

[22]   Z. Zhao, Q. Cheng, X. Li, X.Wang and K. Liu, “c-fos Antisense Oligodeoxynucleotide Reduces VIP-Induced Upregulation of VEGF Expression in Small Cell Lung Cancer Cells,” Chinese Journal of Lung Cancer, Vol. 9, No. 4, 2006, pp. 312-315.

[23]   C. A. Gilbert and A. H. Ross, “Cancer Stem Cells: Cell Culture, Markers and Targets for New Therapies,” Journal of Cellular Biochemistry, Vol. 108, No. 5, 2009, pp. 1031-1038. doi:10.1002/jcb.22350

[24]   T. W. Moody, D. Chan, J. Fahrenkrug and R. T. Jensen, “Neuropeptides as Autocrine Growth Factors in Cancer Cells,” Current Pharmaceutical Design, Vol. 9, No. 6, 2003, pp. 495-509. doi:10.2174/1381612033391621

[25]   U. Olszewski and G. Hamilton, “Neurotensin Signaling Induces Intracellular Alkalinization and Interleukin-8 Expression in Human Pancreatic Cancer Cells,” Molecular Oncology, Vol. 3, 2009, pp. 204-213. doi:10.1016/j.molonc.2009.01.006

[26]   H. Yao, E. Ashihara and T. Maekawa, “Targeting the Wnt/β-Catenin Signaling Pathway in Human Cancers,” Expert Opinion on Therapeutic Targets, Vol. 15, No. 7, 2011, pp. 873-887. doi:10.1517/14728222.2011.577418

[27]   B. Purow, “Notch Inhibition as a Promising New Approach to Cancer Therapy,” Advances in Experimental Medicine and Biology, Vol. 727, 2012, pp. 305-319. doi:10.1007/978-1-4614-0899-4_23

[28]   A. C. Chang, Y. Fu, V. C. Garside, K. Niessen, L. Chang, M. Fuller, A. Setiadi, J. Smrz, A. Kyle, A. Minchinton, M. Marra, P. A. Hoodless and A. Karsan, “Notch Initiates the Endothelial-to-Mesenchymal Transition in the Atrioventricular Canal through Autocrine Activation of Soluble Guanylyl Cyclase,” Developmental Cell, Vol. 21, No. 2, 2011, pp. 288-300. doi:10.1016/j.devcel.2011.06.022

[29]   J. Wang, B. A. Sullenger and J. N. Rich, “Notch Signaling in Cancer Stem Cells,” Advances in Experimental Medicine and Biology, Vol. 727, 2012, pp. 174-185. doi:10.1007/978-1-4614-0899-4_13

[30]   J. Lesley, R. Hyman and P. W. Kincade, “CD44 and Its Interaction with Extracellular Matrix,” Advances in Immunology, Vol. 54, 1993, pp. 271-335. doi:10.1016/S0065-2776(08)60537-4

[31]   M. Zoller, “CD44: Can a Cancer-Initiating Cell Profit from an Abundantly Expressed Molecule?” Nature Reviews Cancer, Vol. 11, No. 4, 2011, pp. 254-267. doi:10.1038/nrc3023

[32]   J. Huang, M. I. Che, Y. T. Huang, M. K. Shyu, Y. M. Huang, Y. M. Wu, W. C. Lin, P. H. Huang, J. T. Liang, P. H. Lee and M. C. Huang, “Overexpression of MUC15 Activates Extracellular Signal-Regulated Kinase 1/2 and Promotes the Oncogenic Potential of Human Colon Cancer Cells,” Carcinogenesis, Vol. 30, No. 8, 2009, pp. 1452-1458. doi:10.1093/carcin/bgp137

[33]   F. Siddiq, F. H. Sarkar, A. Wali, H. I. Pass and F. Lonardo, “Increased Osteonectin Expression Is Associated with Malignant Transformation and Tumor Associated Fibrosis in the Lung,” Lung Cancer, Vol. 45, No. 2, 2004, pp. 197-205.doi:10.1016/j.lungcan.2004.01.020

[34]   M. Fu, E. L. Maresh, R. A. Soslow, M. Alavi, V. Mah, Q. Zhou, A. Iasonos, L. Goodglick, L. K. Gordon, J. Braun and M. Wadehra, “Epithelial Membrane Protein-2 Is a Novel Therapeutic Target in Ovarian Cancer,” Clinical Cancer Research, Vol. 16, No. 15, 2010, pp. 3954-3963. doi:10.1158/1078-0432.CCR-10-0368

[35]   R. O. Hynes, “The Extracellular Matrix: Not Just Pretty Fibrils,” Science, Vol. 326, No. 5957, 2009, pp. 1216-1219. doi:10.1126/science.1176009

[36]   M. Jensen and F. Berthold, “Targeting the Neural Cell Adhesion Molecule in Cancer,” Cancer Letters, Vol. 258, 2007, pp. 9-21. doi:10.1016/j.canlet.2007.09.004

[37]   S. Takahashi, K. Kato, K. Nakamura, R. Nakano, K. Kubota and H. Hamada, “Neural Cell Adhesion Molecule 2 as a Target Molecule for Prostate and Breast Cancer Gene Therapy,” Cancer Science, Vol. 102, No. 4, 2011, pp. 808-814. doi:10.1111/j.1349-7006.2011.01855.x

[38]   M. Namdarghanbari, W. Wobig, S. Krezoski, N. M. Tabatabai and D. H. Petering, “Mammalian Metallothionein in Toxicology, Cancer, and Cancer Chemotherapy,” Journal of Biologic Inorganic Chemistry, Vol. 16, No. 7, 2011, pp. 1087-1101.

[39]   J. S. Moreb, “Aldehyde Dehydrogenase as a Marker for Stem Cells,” Current Stem Cell Research and Therapy, Vol. 3, No. 4, 2008, pp. 237-246. doi:10.2174/157488808786734006

[40]   J. Hilton, “Role of Aldehyde Dehydrogenase in Cyclophosphamide-Resistant L1210 Leukemia,” Cancer Research, Vol. 44, No. 11, 1984, pp. 5156-5160.

[41]   X. Li, L. Wan, J. Geng, C. L. W and X. Bai, “Aldehyde Dehydrogenase 1A1 Possesses Stem-Like Properties and Predicts Lung Cancer Patient Outcome,” Journal of Thoracic Oncology, 2012, in Press. doi:10.1097/JTO.0b013e318257cc6d

[42]   D. C. Seo, J. M. Sung, H. J. Cho, H. Yi, K. H. Seo, I. S. Choi, D. K. Kim, J. S. Kim, A. El-Aty and H. C. Shin, “Gene Expression Profiling of Cancer Stem Cell in Human Lung Adenocarcinoma A549 Cells,” Molecular Cancer, Vol. 6, No. 75, 2007. doi:10.1186/1476-4598-6-75

[43]   G. G. Wang, M. P. Pasillas and M. P. Kamps, “Meis1 Programs Transcription of FLT3 and Cancer Stem Cell Character, Using a Mechanism That Requires Interaction with Pbx and a Novel Function of the Meis1 C-Terminus,” Blood, Vol. 106, No. 1, 2005, pp. 254-264. doi:10.1182/blood-2004-12-4664

[44]   N. Pedersen, S. Mortensen, S. B. Sorensen, M. W. Pedersen, K. Rieneck, L. F. Bovin and H. S. Poulsen, “Transcriptional Gene Expression Profiling of Small Cell Lung Cancer Cells,” Cancer Research, Vol. 63, No. 8, 2003, pp. 1943-1953.

[45]   Koomagi, G. Stammler, C. Manegold, J. Mattern and M. Volm, “Expression of Resistance-Related Proteins in Tumoral and Peritumoral Tissues of Patients with Lung Cancer,” Cancer Letters, Vol. 110, No. 1-2, 1996, pp. 129-136. doi:10.1016/S0304-3835(96)04471-0

[46]   X. Q. Liang, E. H. Cao, Y. Zhang and J. F. Qin, “A P53 Target Gene, PIG11, Contributes to Chemosensitivity of Cells to Arsenic Trioxide,” FEBS Letters, Vol. 569, No. 1-3, 2004, pp. 94-98. doi:10.1016/j.febslet.2004.05.057

[47]   A. Cormier, M. Knossow, C. Wang and B. Gigant, “The Binding of Vinca Domain Agents to Tubulin: Structural and Biochemical Studies,” Methods in Cell Biology, Vol. 95, 2010, pp. 373-390. doi:10.1016/S0091-679X(10)95020-6

[48]   B. Ogretmen, “Sphingolipids in Cancer: Regulation of Pathogenesis and Therapy,” FEBS Letters, Vol. 580, No. 23, 2006, pp. 5467-5476. doi:10.1016/j.febslet.2006.08.052

[49]   G. A. Patwardhan and Y. Y. Liu, “Sphingolipids and Expression Regulation of Genes in Cancer,” Progess in Lipid Research, Vol. 50, No. 1, 2011, pp. 104-114. doi:10.1016/j.plipres.2010.10.003

[50]   M. Kohno, M. L. Momoi, J. H. Oo, Y. M. Paik, K. Lee, Venkataraman, et al., “Intracellular Role for Sphingosine Kinase 1 in Intestinal Adenoma Cell Proliferation,” Molecular Cell Biology, Vol. 26, 2006, pp. 7211-7223. doi:10.1128/MCB.02341-05

[51]   B. Malik and D. Nie, “Cancer Stem Cells and Resistance to Chemo and Radio Therapy,” Frontiers of Bioscience, Vol. 4, 2012, pp. 2142-2149. doi:10.2741/E531

[52]   M. R. García Campelo, G. A. Curbera, G. Aparicio Gallego, E. G. Pulido and L. M. A. Aparicio, “Stem Cell and Lung Cancer Development: Blaming the Wnt, Hh and Notch Signalling Pathway,” Clinical and Translational Oncology, Vol. 13, No. 2, 2011, pp. 77-83. doi:10.1007/s12094-011-0622-0

[53]   C. D. Salcido, A. Larochelle, B. J. Taylor, C. E. Dunbar and L. Varticovski, “Molecular Characterisation of Side Population Cells with Cancer Stem Cell-Like Characteristics in Small-Cell Lung Cancer,” British Journal of Cancer, Vol. 102, No. 11, 2010, pp. 1636-1644. doi:10.1038/sj.bjc.6605668

[54]   R. E. Durand and P. L. Olive, “Resistance of Tumor Cells to Chemo- and Radiotherapy Modulated by the Three-Dimensional Architecture of Solid Tumors and Spheroids,” Methods in Cell Biology, Vol. 64, 2001, pp. 211-233. doi:10.1016/S0091-679X(01)64015-9