Skip to main content
SpringerLink
Log in

Development of a Colorectal Cancer 3D Micro-tumor Construct Platform From Cell Lines and Patient Tumor Biospecimens for Standard-of-Care and Experimental Drug Screening

  • Bioengineering and Enabling Technologies
  • Published:
Annals of Biomedical Engineering Aims and scope Submit manuscript

Abstract

Colorectal cancer is subject to a high rate of mutations, with late stage tumors often containing many mutations. These tumors are difficult to treat, and even with the recently implemented methods of personalized medicine at modern hospitals aiming to narrow treatments, a gap still exists. Proper modeling of these tumors may help to recommend optimal treatments for individual patients, preferably utilizing a model that maintains proper signaling in respect to the derived parent tissue. In this study, we utilized an extracellular matrix-derived hydrogel to create a 3D micro-tumor construct platform capable of both supporting cells for long time durations and for high throughput drug screening. Experiments with cell lines demonstrated long-term viability with maintenance of cell proliferation. Furthermore, studies with several chemotherapeutics utilizing different mechanisms of action displayed differences in efficacy in comparing 3D and 2D cultures. Finally, patient colorectal tumor tissue was acquired and employed to reconstruct micro-tumor constructs, providing a system for the testing of novel chemotherapeutics against tumors in a patient-specific manner. Collectively, the results describe a system capable of high throughput testing while maintaining important characteristics of the parent tissue.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Figure 1
Figure 2
Figure 3
Figure 4
Figure 5

Similar content being viewed by others

Abbreviations

2D:

Two dimensional

3D:

Three dimensional

5-FU:

5-Fluorouracil

CK18:

Cytokeratin 18

CRC:

Colorectal cancer

DMEM:

Dulbecco’s minimum essential medium

DMSO:

Dimethyl sulfoxide

ECM:

Extracellular matrix

EGFR:

Epidermal growth factor receptor

FBS:

Fetal bovine serum

HA:

Hyaluronic acid

IHC:

Immunohistochemistry

IRB:

Institutional Review Board

Ki67:

Antigen Ki-67, a biomarker for proliferation

µTCs:

Micro-tumor constructs

PDMS:

Polydimethylsiloxane

PEGDA:

Polyethylene glycol diacrylate

RNA:

Ribonucleic acid

References

  1. Ahmed, D., P. W. Eide, I. A. Eilertsen, S. A. Danielsen, M. Eknaes, M. Hektoen, G. E. Lind, and R. A. Lothe. Epigenetic and genetic features of 24 colon cancer cell lines. Oncogenesis 2:e71, 2013.

    Article  CAS  Google Scholar 

  2. Ahronian, L. G., E. M. Sennott, E. M. Van Allen, N. Wagle, E. L. Kwak, J. E. Faris, J. T. Godfrey, K. Nishimura, K. D. Lynch, C. H. Mermel, E. L. Lockerman, A. Kalsy, J. M. Gurski, Jr, S. Bahl, K. Anderka, L. M. Green, N. J. Lennon, T. G. Huynh, M. Mino-Kenudson, G. Getz, D. Dias-Santagata, A. J. Iafrate, J. A. Engelman, L. A. Garraway, and R. B. Corcoran. Clinical acquired resistance to RAF inhibitor combinations in BRAF-mutant colorectal cancer through MAPK pathway alterations. Cancer Discov. 5:358–367, 2015.

    Article  CAS  Google Scholar 

  3. Armaghany, T., J. D. Wilson, Q. Chu, and G. Mills. Genetic alterations in colorectal cancer. Gastrointest. Cancer Res. 5:19–27, 2012.

    PubMed  PubMed Central  Google Scholar 

  4. Berg, K. C. G., P. W. Eide, I. A. Eilertsen, B. Johannessen, J. Bruun, S. A. Danielsen, M. Bjornslett, L. A. Meza-Zepeda, M. Eknaes, G. E. Lind, O. Myklebost, R. I. Skotheim, A. Sveen, and R. A. Lothe. Multi-omics of 34 colorectal cancer cell lines—a resource for biomedical studies. Mol. Cancer 16:116, 2017.

    Article  Google Scholar 

  5. Breslin, S., and L. O’Driscoll. The relevance of using 3D cell cultures, in addition to 2D monolayer cultures, when evaluating breast cancer drug sensitivity and resistance. Oncotarget 7:45745–45756, 2016.

    Article  Google Scholar 

  6. Chin, L., J. N. Andersen, and P. A. Futreal. Cancer genomics: from discovery science to personalized medicine. Nat. Med. 17:297–303, 2011.

    Article  CAS  Google Scholar 

  7. de Gramont, A. F. A., M. Seymour, M. Homerin, A. Hmissi, J. Cassidy, C. Boni, H. Cortes-Funes, A. Cervantes, G. Freyer, D. Papamichael, N. Le Bail, C. Louvet, D. Hendler, F. de Braud, C. Wilson, F. Morvan, and A. Bonetti. Leucovorin and fluorouracil with or without oxaliplatin as first-line treatment in advanced colorectal cancer. J. Clin. Oncol. 18:2938–2947, 2000.

    Article  Google Scholar 

  8. De Rosa, M., U. Pace, D. Rega, V. Costabile, F. Duraturo, P. Izzo, and P. Delrio. Genetics, diagnosis and management of colorectal cancer (Review). Oncol. Rep. 34:1087–1096, 2015.

    Article  Google Scholar 

  9. Edmondson, R., J. J. Broglie, A. F. Adcock, and L. Yang. Three-dimensional cell culture systems and their applications in drug discovery and cell-based biosensors. Assay Drug Dev. Technol. 12:207–218, 2014.

    Article  CAS  Google Scholar 

  10. Fleming, M., S. Ravula, S. F. Tatishchev, and H. L. Wang. Colorectal carcinoma: pathologic aspects. J. Gastrointest. Oncol. 3:153–173, 2012.

    PubMed  PubMed Central  Google Scholar 

  11. Gilbert, C. A., and A. H. Ross. Cancer stem cells: cell culture, markers, and targets for new therapies. J. Cell. Biochem. 108:1031–1038, 2009.

    Article  CAS  Google Scholar 

  12. Gmeiner, W. H., W. Debinski, C. Milligan, D. Caudell, and T. S. Pardee. The applications of the novel polymeric fluoropyrimidine F10 in cancer treatment: current evidence. Future Oncol. 12:2009–2020, 2016.

    Article  CAS  Google Scholar 

  13. Greystoke, A., M. Ayub, D. G. Rothwell, D. Morris, D. Burt, C. L. Hodgkinson, C. J. Morrow, N. Smith, K. Aung, J. Valle, L. Carter, F. Blackhall, C. Dive, and G. Brady. Development of a circulating miRNA assay to monitor tumor burden: from mouse to man. Mol. Oncol. 10:282–291, 2016.

    Article  CAS  Google Scholar 

  14. Hamburg, M. A., and F. S. Collins. The path to personalized medicine. N. Engl. J. Med. 363:301–304, 2010.

    Article  CAS  Google Scholar 

  15. Hu, T., Z. Li, C. Y. Gao, and C. H. Cho. Mechanisms of drug resistance in colon cancer and its therapeutic strategies. World J. Gastroenterol. 22:6876–6889, 2016.

    Article  CAS  Google Scholar 

  16. Institute N. S. Cancer Stat Facts: Colon and Rectum Cancer. Bethesda: NIH, 2017.

    Google Scholar 

  17. Kapalczynska, M., T. Kolenda, W. Przybyla, M. Zajaczkowska, A. Teresiak, V. Filas, M. Ibbs, R. Blizniak, L. Luczewski, and K. Lamperska. 2D and 3D cell cultures—a comparison of different types of cancer cell cultures. Arch. Med. Sci. 14:910–919, 2018.

    PubMed  Google Scholar 

  18. Karlsson, H., M. Fryknas, R. Larsson, and P. Nygren. Loss of cancer drug activity in colon cancer HCT-116 cells during spheroid formation in a new 3-D spheroid cell culture system. Exp. Cell Res. 318:1577–1585, 2012.

    Article  CAS  Google Scholar 

  19. Katt, M. E., A. L. Placone, A. D. Wong, Z. S. Xu, and P. C. Searson. In Vitro tumor models: advantages, disadvantages, variables, and selecting the right platform. Front. Bioeng. Biotechnol. 4:12, 2016.

    Article  Google Scholar 

  20. Langhans, S. A. Three-dimensional in vitro cell culture models in drug discovery and drug repositioning. Front. Pharmacol. 9:6, 2018.

    Article  Google Scholar 

  21. Loessner, D., K. S. Stok, M. P. Lutolf, D. W. Hutmacher, J. A. Clements, and S. C. Rizzi. Bioengineered 3D platform to explore cell-ECM interactions and drug resistance of epithelial ovarian cancer cells. Biomaterials 31:8494–8506, 2010.

    Article  CAS  Google Scholar 

  22. Luca, A. C., S. Mersch, R. Deenen, S. Schmidt, I. Messner, K. L. Schafer, S. E. Baldus, W. Huckenbeck, R. P. Piekorz, W. T. Knoefel, A. Krieg, and N. H. Stoecklein. Impact of the 3D microenvironment on phenotype, gene expression, and EGFR inhibition of colorectal cancer cell lines. PLoS ONE 8:e59689, 2013.

    Article  CAS  Google Scholar 

  23. Mazzocchi, A. R., S. A. P. Rajan, K. I. Votanopoulos, A. R. Hall, and A. Skardal. In vitro patient-derived 3D mesothelioma tumor organoids facilitate patient-centric therapeutic screening. Sci. Rep. 8:2886, 2018.

    Article  Google Scholar 

  24. Mazzocchi, A. R., S. Soker, and A. Skardal. Biofabrication technologies for developing in vitro tumor models. In: Tumor Organoids, edited by S. Soker, and A. Skardal. Berlin, Germany: Springer Nature, 2017, pp. 51–70.

    Google Scholar 

  25. Miserocchi, G., L. Mercatali, C. Liverani, A. De Vita, C. Spadazzi, F. Pieri, A. Bongiovanni, F. Recine, D. Amadori, and T. Ibrahim. Management and potentialities of primary cancer cultures in preclinical and translational studies. J. Transl. Med. 15:229, 2017.

    Article  Google Scholar 

  26. Parasuraman, S. Toxicological screening. J. Pharmacol. Pharmacother. 2:74–79, 2011.

    Article  CAS  Google Scholar 

  27. Riedl, A., M. Schlederer, K. Pudelko, M. Stadler, S. Walter, D. Unterleuthner, C. Unger, N. Kramer, M. Hengstschlager, L. Kenner, D. Pfeiffer, G. Krupitza, and H. Dolznig. Comparison of cancer cells in 2D vs 3D culture reveals differences in AKT-mTOR-S6K signaling and drug responses. J. Cell Sci. 130:203–218, 2017.

    Article  CAS  Google Scholar 

  28. Sameer, A. S. Colorectal cancer: molecular mutations and polymorphisms. Front. Oncol. 3:114, 2013.

    Article  Google Scholar 

  29. Scholzen, T., and J. Gerdes. The Ki-67 protein: from the known and the unknown. J. Cell. Physiol. 182:311–322, 2000.

    Article  CAS  Google Scholar 

  30. Siegel, R. L., K. D. Miller, S. A. Fedewa, D. J. Ahnen, R. G. S. Meester, A. Barzi, and A. Jemal. Colorectal cancer statistics, 2017. CA Cancer J. Clin. 67:177–193, 2017.

    Article  Google Scholar 

  31. Skardal, A. Biopolymers for in vitro tissue model biofabrication. In: Biopolymers for Medical Applications, edited by J. M. Ruso, and P. V. Messina. Boca Raton, FL: CRC Press, 2016.

    Google Scholar 

  32. Skardal, A., M. Devarasetty, S. Forsythe, A. Atala, and S. Soker. A reductionist metastasis-on-a-chip platform for in vitro tumor progression modeling and drug screening. Biotechnol. Bioeng. 113:2020–2032, 2016.

    Article  CAS  Google Scholar 

  33. Skardal, A., M. Devarasetty, C. Rodman, A. Atala, and S. Soker. Liver-tumor hybrid organoids for modeling tumor growth and drug response in vitro. Ann. Biomed. Eng. 43:2361–2373, 2015.

    Article  Google Scholar 

  34. Skardal, A., L. Smith, S. Bharadwaj, A. Atala, S. Soker, and Y. Zhang. Tissue specific synthetic ECM hydrogels for 3-D in vitro maintenance of hepatocyte function. Biomaterials 33:4565–4575, 2012.

    Article  CAS  Google Scholar 

  35. Votanopoulos, K. I., A. Mazzocchi, H. Sivakumar, S. Forsythe, J. Aleman, E. A. Levine, and A. Skardal. Appendiceal cancer patient-specific tumor organoid model for predicting chemotherapy efficacy prior to initiation of treatment: a feasibility study. Ann. Surg. Oncol. 26:139–147, 2019.

    Article  Google Scholar 

Download references

Acknowledgments

We wish to thank Libby McWilliams (Procurement Manager), Kathleen Cummings (Protocol and Data Manager) and the Wake Forest Advanced Tumor Bank Shared Resource. AS acknowledges funding through the Wake Forest Clinical and Translational Science Institute Open Pilot Program, supported by the National Center for Advancing Translational Sciences (NCATS), National Institutes of Health, through Grant Award Number UL1TR001420. AS and KV acknowledge funding through the Comprehensive Cancer Center at Wake Forest Baptist Medical Center’s Clinical Research Associate Director Pilot Funds, and services from the Tumor Tissue and Pathology Shared Resource supported by the Comprehensive Cancer Center at Wake Forest Baptist Medical Center’s NCI Cancer Center Support Grant P30CA012197.

Conflict of interest

Dr. Skardal is an inventor on several patents associated with this work related to the generation of patient-derived tumor models for drug screening and personalized medicine.

Author information

Authors and Affiliations

  1. Department of Cancer Biology, Wake Forest School of Medicine, Medical Center Boulevard, Winston-Salem, NC, 27157, USA

    Steven Forsythe, William Gmeiner, Shay Soker & Aleksander Skardal

  2. Wake Forest Institute for Regenerative Medicine, Wake Forest School of Medicine, 391 Technology Way, Winston-Salem, NC, 27101, USA

    Steven Forsythe, Naren Mehta, Mahesh Devarasetty, Hemamylammal Sivakumar, Shay Soker & Aleksander Skardal

  3. Comprehensive Cancer Center at Wake Forest Baptist Medical, Medical Center Boulevard, Winston-Salem, NC, 27157, USA

    William Gmeiner, Shay Soker, Konstantinos Votanopoulos & Aleksander Skardal

  4. Department of Surgery – Oncology, Wake Forest Baptist Medical Center, Medical Center Boulevard, Winston-Salem, NC, 27157, USA

    Konstantinos Votanopoulos

  5. Virginia Tech-Wake Forest, School of Biomedical Engineering and Sciences, Wake Forest School of Medicine, Medical Center Boulevard, Winston-Salem, NC, 27157, USA

    Shay Soker & Aleksander Skardal

  6. Department of Molecular Medicine and Translational Science, Wake Forest School of Medicine, Medical Center Boulevard, Winston-Salem, NC, 27157, USA

    Shay Soker & Aleksander Skardal

Authors
  1. Steven Forsythe
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Naren Mehta
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Mahesh Devarasetty
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Hemamylammal Sivakumar
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. William Gmeiner
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Shay Soker
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Konstantinos Votanopoulos
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Aleksander Skardal
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Aleksander Skardal.

Additional information

Associate Editor Jane Grande-Allen oversaw the review of this article.

Publisher's Note

Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Electronic supplementary material

Below is the link to the electronic supplementary material.

Supplementary material 1 (DOCX 3826 kb)

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Forsythe, S., Mehta, N., Devarasetty, M. et al. Development of a Colorectal Cancer 3D Micro-tumor Construct Platform From Cell Lines and Patient Tumor Biospecimens for Standard-of-Care and Experimental Drug Screening. Ann Biomed Eng 48, 940–952 (2020). https://doi.org/10.1007/s10439-019-02269-2

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s10439-019-02269-2

Keywords

Search

Navigation