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. 2005 Jun 16;46(1):9–18. doi: 10.1007/s10616-005-1476-5

Identification of three gene candidates for multicellular resistance in colon carcinoma

Nicholas E Timmins 1, Tina L Maguire 2, Sean M Grimmond 2, Lars K Nielsen 1,✉
PMCID: PMC3449472  PMID: 19003254

Abstract

Solid tumours display elevated resistance to chemo- and radiotherapies compared to individual tumour derived cells. This so-called multicellular resistance (MCR) phenomenon can only be partly explained by reduced diffusion and altered cell cycle status; even fast growing cells on the surface of solid tumours display MCR. Multicellular spheroids (MCS) recapture this phenomenon ex vivo and here we compare gene expression in exponentially growing MCS with gene expression in monolayer culture. Using an 18,664 gene microarray, we identified 42 differentially expressed genes and three of these genes can be linked to potential mechanisms of MCR. A group of interferon response genes were also up-regulated in MCS, as were a number of genes that that are indicative of greater differentiation in three-dimensional cultures.

Keywords: 3D culture, Differential gene expression, MCR, MCS, Multicellular resistance, Multicellular spheroid, Tumour

Full Text

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Glossary

3D

Three-dimensional

5FU

5-Fluorouracil

MCR

Multicellular resistance

MCS

Multicellular spheroid

References

  1. Arany I., Chen S.H., Megyesi J.K., Adler-Storthz K., Chen Z., Rajaraman S., Ember I.A., Tyring S.K., Brysk M.M. Differentiation-dependent expression of signal transducers and activators of transcription (STATs) might modify responses to growth factors in the cancers of the head and neck. Cancer Lett. 2003;199(1):83–89. doi: 10.1016/S0304-3835(03)00345-8. [DOI] [PubMed] [Google Scholar]
  2. Baldwin A.S. Control of oncogenesis and cancer therapy resistance by the transcription factor NF-kappa B. J. Clin. Invest. 2001;107(3):241–246. doi: 10.1172/JCI11991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Courilleau D., Chastre E., Sabbah M., Redeuilh G., Atfi A., Mester J. B-ind1, a novel mediator of Rac1 signaling cloned from sodium butyrate-treated fibroblasts. J. Biol. Chem. 2000;275(23):17344–17348. doi: 10.1074/jbc.M000887200. [DOI] [PubMed] [Google Scholar]
  4. Debatin K.M. Apoptosis pathways in cancer and cancer therapy. Cancer Immunol. Immunother. 2004;53(3):153–159. doi: 10.1007/s00262-003-0474-8. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Desoize B., Jardillier J. Multicellular resistance: a paradigm for clinical resistance? Crit. Rev. Oncol. Hematol. 2000;36(2-3):193–207. doi: 10.1016/s1040-8428(00)00086-x. [DOI] [PubMed] [Google Scholar]
  6. Dubessy C., Merlin J.M., Marchal C., Guillemin F. Spheroids in radiobiology and photodynamic therapy. Crit. Rev. Oncol. Hematol. 2000;36(2-3):179–192. doi: 10.1016/s1040-8428(00)00085-8. [DOI] [PubMed] [Google Scholar]
  7. Flatmark K., Pedersen K., Nesland J., Rasmussen H., Aamodt G., Mikalsen S., Bjornland K., Fodstad O., Maelandsmo G. Nuclear localization of the metastasis-related protein s100a4 correlates with tumour stage in colorectal cancer. J. Pathol. 2003;200(5):589–595. doi: 10.1002/path.1381. [DOI] [PubMed] [Google Scholar]
  8. Fracasso G., Colombatti M. Effect of therapeutic macromolecules in spheroids. Crit. Rev. Oncol. Hematol. 2000;36(2-3):159–178. doi: 10.1016/s1040-8428(00)00084-6. [DOI] [PubMed] [Google Scholar]
  9. Hamilton G. Multicellular spheroids as an in vitro tumor model. Cancer Lett. 1998;131:29–34. doi: 10.1016/S0304-3835(98)00198-0. [DOI] [PubMed] [Google Scholar]
  10. Huang J., Teng L., Liu T., Li L., Chen D., Li F., Xu L.G., Zhai Z., Shu H.B. Identification of a novel serine/threonine kinase that inhibits TNF-induced NF-kappaB activation and p53-induced transcription. Biochem. Biophys. Res. Commun. 2003;309(4):774–778. doi: 10.1016/j.bbrc.2003.08.069. [DOI] [PubMed] [Google Scholar]
  11. Hwang B.J., Ford J.M., Hanawalt P.C., Chu G. Expression of the p48 xeroderma pigmentosum gene is p53-dependent and is involved in global genomic repair. Proc. Natl. Acad. Sci. USA. 1999;96(2):424–428. doi: 10.1073/pnas.96.2.424. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Kelm J.M., Timmins N.E., Brown C.J., Fussenegger M., Nielsen L.K. Method for generation of homogeneous multicellular tumor spheroids applicable to a wide variety of cell types. Biotechnol. Bioeng. 2003;83(2):173–180. doi: 10.1002/bit.10655. [DOI] [PubMed] [Google Scholar]
  13. Knowles H.J., Phillips R.M. Identification of differentially expressed genes in experimental models of the tumor microenvironment using differential display. Anticancer Res. 2001;21(4A):2305–2311. [PubMed] [Google Scholar]
  14. Kunz-Schughart L., Mueller-Klieser W. Three-dimensional culture. In: Masters J., editor. Animal Cell Culture: A Practical Approach. 3. New York: Oxford University Press; 2000. pp. 123–184. [Google Scholar]
  15. Kunz-Schughart L.A., Kreutz M., Knuechel R. Multicellular spheroids: a three-dimensional in vitro culture system to study tumour biology. Int. J. Exp. Pathol. 1998;79(1):1–23. doi: 10.1046/j.1365-2613.1998.00051.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Lazareff J.A., Suwinski R., De Rosa R., Olmstead C.E. Tumor volume and growth kinetics in hypothalamic-chiasmatic pediatric low grade gliomas. Pediatr. Neurosurg. 1999;30(6):312–319. doi: 10.1159/000028817. [DOI] [PubMed] [Google Scholar]
  17. Luker K., Pica C., Schreiber R., Piwnica-Worms D. Overexpression of irf9 confers resistance to antimicrotubule agents in breast cancer cells. Cancer Res. 2001;61(17):6540–6547. [PubMed] [Google Scholar]
  18. Marusic M., Bajzer Z., Vuk-Pavlovic, Freyer J. Tumour growth in vivoas multicellular spheroids compared by mathematical models. Bull. Math. Biol. 1994;56:617–631. doi: 10.1016/S0092-8240(05)80306-4. [DOI] [PubMed] [Google Scholar]
  19. Mayo M.W., Baldwin A.S. The transcription factor NF-kappaB: control of oncogenesis and cancer therapy resistance. Biochim. Biophys. Acta. 2000;1470(2):M55–62. doi: 10.1016/s0304-419x(00)00002-0. [DOI] [PubMed] [Google Scholar]
  20. Moriyama-Kita M., Endo Y., Yonemura Y., Heizmann C., Schafer B., Sasaki T., Yamamoto E. Correlation of s100a4 expression with invasion and metastasis in oral squamous cell carcinoma. Oral Oncol. 2004;40(5):496–500. doi: 10.1016/j.oraloncology.2003.10.003. [DOI] [PubMed] [Google Scholar]
  21. Mueller-Klieser W. Three-dimensional cell cultures: from molecular mechanisms to clinical applications. Am. J. Physiol. 1997;273(4 Pt 1):C1109–1123. doi: 10.1152/ajpcell.1997.273.4.C1109. [DOI] [PubMed] [Google Scholar]
  22. Nederman T., Twentyman P. Spheroids for studies of drug effects. Recent Results Cancer Res. 1984;95:84–102. doi: 10.1007/978-3-642-82340-4_5. [DOI] [PubMed] [Google Scholar]
  23. Olive P.L., Durand R.E. Drug and radiation resistance in spheroids: cell contact and kinetics. Cancer Metastasis Rev. 1994;13(2):121–138. doi: 10.1007/BF00689632. [DOI] [PubMed] [Google Scholar]
  24. Olive P.L., Banath J.P., Evans H.H. Cell killing and DNA damage by etoposide in Chinese hamster V79 monolayers and spheroids: influence of growth kinetics, growth environment and DNA packaging. Br. J. Cancer. 1993;67(3):522–530. doi: 10.1038/bjc.1993.97. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Oloumi A., MacPhail S.H., Johnston P.J., Banath J.P., Olive P.L. Changes in subcellular distribution of topoisomerase IIalpha correlate with etoposide resistance in multicell spheroids and xenograft tumors. Cancer Res. 2000;60(20):5747–5753. [PubMed] [Google Scholar]
  26. Oloumi A., Lam W., Banath J.P., Olive P.L. Identification of genes differentially expressed in v79 cells grown as multicell spheroids. Int. J. Radiation Biol. 2002;78(6):483–492. doi: 10.1080/09553000210122299. [DOI] [PubMed] [Google Scholar]
  27. Perou C.M., Jeffrey S.S., Rijn M., Rees C.A., Eisen M.B., Ross D.T., Pergamenschikov A., Williams C.F., Zhu S.X., Lee J.C., Lashkari D., Shalon D., Brown P.O., Botstein D. Distinctive gene expression patterns in human mammary epithelial cells and breast cancers. Proc. Natl. Acad. Sci. USA. 1999;96(16):9212–9217. doi: 10.1073/pnas.96.16.9212. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Poland J., Sinha P., Siegert A., Schnolzer M., Korf U., Hauptmann S. Comparison of protein expression profiles between monolayer and spheroid cell culture of ht-29 cells revealed fragmentation of ck18 in three-dimensional cell culture. Electrophoresis. 2002;23(78):1174–1184. doi: 10.1002/1522-2683(200204)23:7/8<1174::AID-ELPS1174>3.0.CO;2-O. [DOI] [PubMed] [Google Scholar]
  29. Rainaldi G., Calcabrini A., Arancia G., Santini M.T. Differential expression of adhesion molecules (CD44, ICAM-1 and LFA-3) in cancer cells grown in monolayer or as multicellular spheroids. Anticancer Res. 1999;19(3A):1769–1778. [PubMed] [Google Scholar]
  30. Ramakers C., Ruijter J.M., Deprez R.H., Moorman A.F. Assumptionfree analysis of quantitative real-time polymerase chain reaction (PCR) data. Neurosci. Lett. 2003;339(1):62–66. doi: 10.1016/S0304-3940(02)01423-4. [DOI] [PubMed] [Google Scholar]
  31. Rosty C., Ueki T., Argani P., Jansen M., Yeo C., Cameron J., Hruban R., Goggins M. Overexpression of S100A4 in pancreatic ductal adenocarcinomas is associated with poor differentiation and dna hypomethylation. Am. J. Pathol. 2002;160(1):45–50. doi: 10.1016/S0002-9440(10)64347-7. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Shiras A., Bhosale A., Patekar A., Shepal V., Shastry P. Differential expression of CD44(S) and variant isoforms v3, v10 in three-dimensional cultures of mouse melanoma cell lines. Clin. Exp. Metastasis. 2002;19(5):445–455. doi: 10.1023/A:1016305611858. [DOI] [PubMed] [Google Scholar]
  33. Sutherland R. Cell and environment interactions in tumor microregions: the multicell spheroids model. Science. 1988;240:177–184. doi: 10.1126/science.2451290. [DOI] [PubMed] [Google Scholar]
  34. Wartenberg M., Frey C., Diedershagen H., Ritgen J., Hescheler J., Sauer H. Development of an intrinsic P-glycoprotein-mediated doxorubicin resistance in quiescent cell layers of largemulticellular prostate tumor spheroids. Int. J. Cancer. 1998;75(6):855–863. doi: 10.1002/(SICI)1097-0215(19980316)75:6<855::AID-IJC7>3.0.CO;2-U. [DOI] [PubMed] [Google Scholar]
  35. Weaver V.M., Lelievre S., Lakins J.N., Chrenek M.A., Jones J.C., Giancotti F., Werb Z., Bissell M.J. Beta4 integrin-dependent formation of polarized three-dimensional architecture confers resistance to apoptosis in normal and malignant mammary epithelium. Cancer Cell. 2002;2(3):205–216. doi: 10.1016/S1535-6108(02)00125-3. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Wu M., Xu L.G., Zhai Z., Shu H.B. SINK is a p65-interacting negative regulator of NF-kappaB-dependent transcription. J. Biol. Chem. 2003;278(29):27072–27079. doi: 10.1074/jbc.M209814200. [DOI] [PubMed] [Google Scholar]

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