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Study of Cancer Stem Cell Subpopulations in Breast Cancer Models

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Abstract

The heterogeneous nature of tumor populations and the presence of tumor stem cells is one of the causes for resistance of malignant neoplasms to anticancer therapies and emergence of recurrences and metastases as well as for complexities in the management of this pathology. The aim of this study was to enrich multicellular tumor spheroids (MCTSs) of the mammary adenocarcinoma MCF-7 line cells with cancer stem cells (CSCs) and study the obtained CSC subpopulation using biochemical, immunological, and cytological methods. The results of our study have shown that the percentage of CSCs within a population of multicellular tumor spheroids in a nutrient-depleted growth medium increases significantly with certain growth factor supplements. For example, the percentage of the cells expressing CD133 and Nestin increased, respectively, from 12.47 to 82.08% and from 31.3 to 82.58%. The data of immunocytochemical staining showed that the count of cells expressing the CSC markers, such as CD44, CD133, and bmi1, also increased. The aldehyde dehydrogenase activity reached 0.07 mol/mg protein per min in the MCF7 line cells under the monolayer growth conditions and increased up to 1.58 mol/mg protein per min in CSCs-enriched multicellular tumor spheroids (eMCTSs). The activity of glucose-6-phosphate dehydrogenase (G6PDH) in the tumor cells was 934.6 ± 148.3 × 10–6 mol/mg protein per min under the monolayer growth conditions and increased more than 1.5 times with enriching MCTSs with CSCs. The activity of lactate dehydrogenase (LDH) in MCF-7 cells was 65.12 ± 1.28 μmol/mg protein per min under the monolayer growth conditions and decreased by 5.5 times due to a growth in the CTCs-enriched MCTSs. Thus, based on the authors’ data, one can assume that the MCF-7 receptor and energy profile change due to enriching a tumor cell population with CSCs under growth conditions, thus bringing the CTCs-enriched spheroids closer to the characteristics of metastatic micronodules and the tumor cells to those of cancer stem cells.

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REFERENCES

  1. Baccelli, I. and Trumpp, A., The evolving concept of cancer and metastasis stem cells, J. Cell Biol., 2012, vol. 198, pp. 281–293. https://doi.org/10.1083/jcb.201202014

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  2. Bapat, S., Mali, A., Koppikar, C., et al., Stem and progenitor-like cells contribute to the aggressive behavior of human epithelial ovarian cancer, Cancer Res., 2005, vol. 65, pp. 3025–3029. https://doi.org/10.1158/0008-5472.can-04-3931

    Article  CAS  PubMed  Google Scholar 

  3. Bjerkvig, R., Spheroid Culture in Cancer Research, Boca Raton: CRC Press, 1992.

    Google Scholar 

  4. Borlle, L., Dergham, A., Wund, Z., et al., Salinomycin decreases feline sarcoma and carcinoma cell viability when combined with doxorubicin, BMC Vet. Res., 2019, vol. 15, no. 1. https://doi.org/10.1186/s12917-019-1780-5

  5. Brugnoli, F., Grassilli, S., Al-Qassab, Y., et al., CD133 in breast cancer cells: more than a stem cell marker, J. Oncol., 2019, vol. 2019, art. ID 7512632. https://doi.org/10.1155/2019/7512632

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  6. Colak, S. and Medema, J., Cancer stem cells – important players in tumor therapy resistance, FEBS J., 2014, vol. 281, no. 21, pp. 4779–4791. https://doi.org/10.1111/febs.13023

    Article  CAS  PubMed  Google Scholar 

  7. Collins, A., Berry, P., Hyde, C., et al., Prospective identification of tumorigenic prostate cancer stem cells, Cancer Res., 2005, vol. 65, pp. 10946–10951. https://doi.org/10.1158/0008-5472.can-05-2018

    Article  CAS  PubMed  Google Scholar 

  8. Cui, J., Shi, M., Xie, D., et al., FOXM1 promotes the Warburg effect and pancreatic cancer progression via transactivation of LDHA expression, Clin. Cancer Res., 2014, vol. 20, no. 10, pp. 2595–2606. https://doi.org/10.1158/1078-0432.ccr-13-2407

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  9. Ehrmann, J., Kolar, Z., and Mokry, J., Nestin as a diagnostic and prognostic marker: immunohistochemical analysis of its expression in different tumours, J. Clin. Pathol., 2005, vol. 58, no. 2, pp. 222–223. https://doi.org/10.1136/jcp.2004.021238

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  10. Feng, Y., Xiong, Y., and Qiao, T., Lactate dehydrogenase A: A key player in carcinogenesis and potential target in cancer therapy, Cancer Med., 2018, vol. 7, no. 12. https://doi.org/10.1002/cam4.1820

  11. Ghanbari Movahed, Z., Rastegari-Pouyani, M., Mohammadi, M., et al., Cancer cells change their glucose metabolism to overcome increased ROS: One step from cancer cell to cancer stem cell?, Biomed. Pharmacother., 2019, vol. 112, art. ID 108690. https://doi.org/10.1016/j.biopha.2019.108690

    Article  CAS  PubMed  Google Scholar 

  12. Giatromanolaki, A., Sivridis, E., Gatter, K., et al., Lactate dehydrogenase 5 (LDH-5) expression in endometrial cancer relates to the activated VEGF/VEGFR2(KDR) pathway and prognosis, Gynecol. Oncol., 2006, vol. 103, no. 3, pp. 912–918. https://doi.org/10.1016/j.ygyno.2006.05.043

    Article  CAS  PubMed  Google Scholar 

  13. He, Q., Luo, X., Wang, K., et al., Isolation and characterization of cancer stem cells from high-grade serous ovarian carcinomas, Cell. Physiol. Biochem., 2014, vol. 33, no. 1, pp. 173–184. https://doi.org/10.1159/000356660

    Article  CAS  PubMed  Google Scholar 

  14. Herheliuk, T., Perepelytsina, O., Ugnivenko, A., et al., Investigation of multicellular tumor spheroids enriched for a cancer stem cell phenotype, Stem Cell Invest., 2019, vol. 6, art. ID 21. https://doi.org/10.21037/sci.2019.06.07

    Article  CAS  Google Scholar 

  15. Hermann, P., Huber, S., Herrler, T., et al., Distinct populations of cancer stem cells determine tumor growth and metastatic activity in human pancreatic cancer, Cell Stem Cell, 2007, vol. 1, no. 3, pp. 313–323. https://doi.org/10.1016/j.stem.2007.06.002

    Article  CAS  PubMed  Google Scholar 

  16. Hockmair, M., Rath, B., Klameth, L., et al., Effects of salinomycin and niclosamide on small cell lung cancer and small cell lung cancer circulating tumor cell lines, Invest. New Drugs, 2020, vol. 38, no. 4, pp. 46–955. https://doi.org/10.1007/S10637-019-00847-8

    Article  Google Scholar 

  17. Jiang, P., Du, W., and Wu, M., Regulation of the pentose phosphate pathway in cancer, Protein Cell, 2014, vol. 5, pp. 592–602. https://doi.org/10.1007/s13238-014-0082-8

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  18. Jiang, W., Zhou, F., Li, N., et al., FOXM1-LDHA signaling promoted gastric cancer glycolytic phenotype and progression, Int. J. Clin. Exp. Pathol., 2015, vol. 8, no. 6, pp. 6756–6763.

    PubMed  PubMed Central  Google Scholar 

  19. Karakaya, H. and Ozkul, K., Measurement of glucose-6-phosphate dehydrogenase activity in bacterial cell-free extracts, Bio-Protoc., 2016, vol. 6, no. 19, art. ID e1949. https://doi.org/10.21769/BioProtoc.1949

  20. Ketola, K., Hilvo, M., Hyötyläinen, T., Vuoristo, A., et al., Salinomycin inhibits prostate cancer growth and migration via induction of oxidative stress, Brit. J. Cancer, 2012, vol. 106, pp. 99–106. https://doi.org/10.1038/bjc.2011.530

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  21. Kim, Y., Siegler, E., Siriwon, N., and Wang, P., Therapeutic strategies for targeting cancer stem cells, J. Cancer Metastasis Treat., 2016, vol. 2, pp. 233–242. https://doi.org/10.20517/2394-4722.2016.26

    Article  CAS  Google Scholar 

  22. Kleeberger, W., Bova, G.S., and Nielsen, M.E., Roles for the stem cell associated intermediate filament Nestinin prostate cancer migration and metastasis, Cancer Res., 2007, vol. 67, no. 19, pp. 9199–9206. https://doi.org/10.1158/0008-5472.CAN-07-0806

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  23. Koukourakis, M., Kakouratos, C., and Kalamida, D., Hypoxia-inducible proteins HIF1α and lactate dehydrogenase LDH5, key markers of anaerobic metabolism, relate with stem cell markers and poor post-radiotherapy outcome in bladder cancer, Int. J. Radiat. Biol., 2016, vol. 92, no. 7, pp. 353–363. https://doi.org/10.3109/09553002.2016.1162921

    Article  CAS  PubMed  Google Scholar 

  24. Krupkova, Jr., Loja, T., Zambo, I., and Veselska, R., Nestin expression in human tumors and tumor cell lines, Neoplasma, 2010, vol. 4, pp. 291–298. https://doi.org/10.4149/neo_2010_04_291

    Article  Google Scholar 

  25. Kumar, V. and Gill, K.D., Determination of total lactate dehydrogenase activity in serum sample, in Basic Concepts in Clinical Biochemistry, A Practical Guide, Springer-Verlag, 2018, pp. 129–130. https://doi.org/10.1007/978-981-10-8186-6_32

    Book  Google Scholar 

  26. Kurpinska, A., Suraj, J., Bonar, E., et al., Proteomic characterization of early lung response to breast cancer metastasis in mice, Exp. Mol. Pathol., 2019, vol. 407, pp. 129–140. https://doi.org/10.1016/j.yexmp.2019.02.001

    Article  CAS  Google Scholar 

  27. Ma, L., Lai, D., Liu, T., et al., Cancer stem-like cells can be isolated with drug selection in human ovarian cancer cell line SKOV3, Acta Biochim. Biophys. Sin., 2010, vol. 42, no. 9, pp. 593–602. https://doi.org/10.1093/abbs/gmq067

    Article  CAS  PubMed  Google Scholar 

  28. Mukherjee, D. and Ahmad, R., Glucose-6-phosphate dehydrogenase activity during N'-nitrosodiethylamine-induced hepatic damage, Ach. Life Sci., 2015, vol. 9, pp. 51–56. https://doi.org/10.1016/j.als.2015.05.007

    Article  Google Scholar 

  29. Naujokat, C., Salinomycin in cancer: A new mission for an old agent, Mol. Med. Rep., 2015, vol. 3, no. 4, pp. 555–559. https://doi.org/10.1155/2012/950658

    Article  Google Scholar 

  30. Neradil, J. and Veselska, R., Nestin as a marker of cancer stem cells, Cancer Sci., 2015, vol. 106, no. 7, pp. 803–811. https://doi.org/10.1111/cas.12691

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  31. Patra, K. and Hay, N., The phosphate pathway and cancer, Trends Biochem., 2014, vol. 39, pp. 347–354. https://doi.org/10.1016/j.tibs.2014.06.005

    Article  CAS  Google Scholar 

  32. Piras, F., Perra, M.T., Murtas, D., et al., The stem cell marker nestin predicts poor prognosis in human melanoma, Oncol. Rep., 2010, vol. 23, no. 1, pp. 17–24. https://doi.org/10.3892/or_00000601

    Article  PubMed  Google Scholar 

  33. Ramos-Martinez, J., The regulation of the pentose phosphate pathway: Remember Krebs, Arch. Biochem. Biophys., 2017, vol. 614, pp. 50–52. https://doi.org/10.1016/j.abb.2016.12.012

    Article  CAS  PubMed  Google Scholar 

  34. Rappa, G., Fodstad, O., and Lorico, A., The stem cell-associated antigen CD133 (Prominin-1) is a molecular therapeutic target for metastatic melanoma, Stem Cells, 2008, vol. 26, no. 12, pp. 3008–3017. https://doi.org/10.1634/stemcells.2008-0601

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  35. Resham, K., Patel, P., Thummuri, D., et al., Preclinical drug metabolism and pharmacokinetics of salinomycin, a potential candidate for targeting human cancer stem cells, Chem.-Biol. Interact., 2015, vol. 240, pp. 146–152. https://doi.org/10.1016/j.cbi.2015.08.007

    Article  CAS  PubMed  Google Scholar 

  36. Sant, S., Johnston, P., et al., The production of 3D tumor spheroids for cancer drug discovery, Drug Discovery Today: Technol., 2017, vol. 23, pp. 27–36. https://doi.org/10.1016/j.ddtec.2017.03.002

    Article  Google Scholar 

  37. Schneider, M., Huber, J., Hadaschik, B., et al., Characterization of colon cancer cells: a functional approach characterizing CD133 as a potential stem cell marker, BMC Cancer, 2012, vol. 12, art. ID 96. https://doi.org/10.1186/1471-2407-12-96

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  38. Singh, Sh., Clarke, I., Terasaki, M., et al., Identification of a cancer stem cell in human brain, Cancer Res., 2003, vol. 63, no. 18, pp. 5821–5828.

    CAS  PubMed  Google Scholar 

  39. Strojnik, T., Rosland, G.V., Sakariassen, P.O., et al., Neural stem cell markers, nestin and musashi proteins, in the progression of human glioma: correlation of nestin with prognosis of patient survival, Surg. Neurol., 2007, vol. 68, no. 2, pp. 133–143. https://doi.org/10.1089/scd.2008.0359

    Article  PubMed  Google Scholar 

  40. Su, Y., Yu, Q., Wang, X., et al., JMJD2A promotes the Warburg effect and nasopharyngeal carcinoma progression by transactivating LDHA expression, BMC Cancer, 2007, vol. 17, art. ID 477. https://doi.org/10.1186/s12885-017-3473-4

    Article  CAS  Google Scholar 

  41. Talaiezadeh, A., Shahriari, A., Tabandeh, M., et al., Kinetic characterization of lactate dehydrogenase in normal and malignant human breast tissues, Cancer Cell Int., 2015, vol. 15, art. ID 19. https://doi.org/10.1186/s12935-015-0171-7

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  42. Tang, Q., Zhao, Z.-Q., Li, J.-C., Liang, Y., et al., Salinomycin inhibits osteosarcoma by targeting its tumor stem cells, Cancer Lett., 2011, vol. 311, pp. 113–121. https://doi.org/10.1016/j.canlet.2011.07.016

    Article  CAS  PubMed  Google Scholar 

  43. Taniguchi, M., Mori, N., and Iramina, C., Elevation of glucose 6-phosphate dehydrogenase activity induced by amplified insulin response in low glutathione levels in rat liver, Sci. World J., 2016, vol. 2016, art. ID 6382467. https://doi.org/10.1155/2016/6382467

    Article  CAS  Google Scholar 

  44. Teranishi, N., Naito, Z., Ishiwata, T., et al., Identification of neovasculature using nestin in colorectal cancer, Int. J. Oncol., 2007, vol. 30, no. 3, pp. 593–603. https://doi.org/10.3892/ijo.30.3.593

    Article  CAS  PubMed  Google Scholar 

  45. Tropepe, V., Alton, K., Sachewsky, N., et al., Neurogenic potential of isolated precursor cells from early post-gastrula somitic tissue, Stem Cells Dev., 2009, vol. 18, no. 10, pp. 1533–1542. https://doi.org/10.1089/scd.2008.0359

    Article  CAS  PubMed  Google Scholar 

  46. Vassalli, G., Aldehyde Dehydrogenases: not just markers, but functional regulators of stem cells, Stem Cells Int., 2019, vol. 2019, art. ID 3904645. https://doi.org/10.1155/2019/3904645

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  47. Versini, A., Colombeau, L., and Hienzsch, A., Salinomycin derivatives kill breast cancer stem cells by lysosomal iron targeting, Chem. - Eur. J., 2020, vol. 26, no. 33. https://doi.org/10.1002/chem.202000335

  48. Wang, H., Zhang, H., Zhu, Y., et al., Anticancer mechanisms of salinomycin in breast cancer and its clinical applications, Front. Oncol., 2021. https://doi.org/10.3389/fonc.2021.654428

  49. Wang, Y., Effects of salinomycin on cancer stem cell in human lung adenocarcinoma A549 cells, Med. Chem., 2011, vol. 7, no. 2, pp. 106–111. https://doi.org/10.2174/157340611794859307

    Article  CAS  PubMed  Google Scholar 

  50. Wong, T., Che, N., and Ma, S., Reprogramming of central carbon metabolism in cancer stem cells, Biochim. Biophys. Acta, Mol. Basis Dis., 2017, vol. 1863, pp. 1728–1738. https://doi.org/10.1016/j.bbadis.2017.05.012

    Article  CAS  Google Scholar 

  51. Yin, A.H., Miraglia, S., Zanjani, E.D., et al., AC133, a novel marker for human hematopoietic stem and progenitor cells, Blood, 1997, vol. 90, no. 12, pp. 5002–5012.

    Article  CAS  Google Scholar 

  52. Zdralevic, M., Marchiq, I., Cunhade, P., et al., Metabolic plasiticy in cancers—distinct role of glycolytic enzymes GPI, LDHs or membrane transporters MCTs, Front. Oncol., 2017. https://doi.org/10.3389/fonc.2017.00313

  53. Zhang, C., Tian, Y., Song, F., et al., Salinomycin inhibits the growth of colorectal carcinoma by targeting tumor stem cells, Oncol. Rep., 2015. https://doi.org/10.3892/or.2015.4253

  54. Zhao, Z., Lu, P., and Zhang, H., Nestin positively regulates the Wnt/β-catenin pathway and the proliferation, survival and invasiveness of breast cancer stem cells, Breast Cancer Res., 2014, vol. 16, art. ID 408. https://doi.org/10.1186/s13058-014-0408-8

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  55. Zhi, Q., Chen, X., Ji, J., Zhang, J., et al., Salinomycin can effectively kill ALDHhigh stem-like cells on gastric cancer, Biomedicine & Pharmacotherapy, 2011, vol. 65, no. 7, pp. 509–515. https://doi.org/10.1016/j.biopha.2011.06.006

    Article  CAS  Google Scholar 

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This study has not received any particular grant from financial establishments in the state, commercial, or noncommercial sectors.

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Authors and Affiliations

  1. Department of Biotechnical Problems of Diagnostics, Institute for Problems of Cryobiology and Cryomedicine, National Academy of Sciences of Ukraine, 03028, Kyiv, Ukraine

    T. S. Herheliuk, O. M. Perepelytsina, Yu. M. Chmelnytska & M. V. Sydorenko

  2. Institute of Biology and Medicine, Taras Shevchenko Kyiv National University, 03187, Kyiv, Ukraine

    T. S. Herheliuk, Yu. M. Chmelnytska, G. M. Kuznetsova, N. V. Dzjubenko & N. G. Raksha

  3. National Cancer Institute of Ukraine, Ministry of Healthcare of Ukraine, 03022, Kyiv, Ukraine

    O. I. Gorbach

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  1. T. S. Herheliuk
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  2. O. M. Perepelytsina
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Correspondence to O. M. Perepelytsina or Yu. M. Chmelnytska.

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Translated by N. Tarasyuk

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Herheliuk, T.S., Perepelytsina, O.M., Chmelnytska, Y.M. et al. Study of Cancer Stem Cell Subpopulations in Breast Cancer Models. Cytol. Genet. 56, 331–342 (2022). https://doi.org/10.3103/S0095452722040041

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