Preclinical Breast Cancer Models to Investigate Metabolic Priming by Methionine Restriction

  • Elena Strekalova
  • Dmitry Malin
  • Harisha Rajanala
  • Vincent L. CrynsEmail author
Part of the Methods in Molecular Biology book series (MIMB, volume 1866)


We have developed a novel therapeutic paradigm (“metabolic priming”) for cancer whereby restriction of the essential amino acid methionine activates a number of cell-stress-response pathways that can be selectively targeted to enhance the therapeutic impact of methionine restriction. One example of metabolic priming is the combination of methionine restriction with proapoptotic TRAIL receptor-2 (TRAIL-R2) agonists. Methionine restriction enhances the cell surface expression of TRAIL-R2 selectively in transformed breast epithelial cells and renders them more susceptible to cell death induction by TRAIL-R2 agonists in cellular and murine models of breast cancer. This methods review focuses on preclinical models of breast cancer to investigate metabolic priming by methionine restriction. Multiple cell-based methods are detailed to measure cell viability, cell survival, caspase activity, apoptosis, and matrix detachment-induced cell death (anoikis). In addition, we describe an orthotopic model of metastatic breast cancer that utilizes mCherry-fluorescently-labeled human breast cancer cells. This model captures the entire metastatic cascade from the mammary gland to the lung and mimics key features of the human disease. These breast-cancer models can be readily adapted to other tumor types. Overall, we provide a stepwise, translationally-relevant approach to study metabolic priming in the context of cancer.

Key words

Methionine Metabolism TRAIL Breast cancer Metastasis Apoptosis Therapeutics Murine models 



We are indebted to Robin Humphreys for providing agonistic TRAIL receptor mAbs. This work was supported by grants from the V Foundation for Cancer Research, Breast Cancer Research Foundation, Avon Breast Cancer Crusade, UW Carbone Cancer Center pilot funding, and the Wisconsin Partnership Program.


  1. 1.
    Guo HY, Herrera H, Groce A, Hoffman RM (1993) Expression of the biochemical defect of methionine dependence in fresh patient tumors in primary histoculture. Cancer Res 53:2479–2483PubMedGoogle Scholar
  2. 2.
    Kreis W, Baker A, Ryan V, Bertasso A (1980) Effect of nutritional and enzymatic methionine deprivation upon human normal and malignant cells in tissue culture. Cancer Res 40:634–641PubMedGoogle Scholar
  3. 3.
    Lu S, Hoestje SM, Choo EM, Epner DE (2002) Methionine restriction induces apoptosis of prostate cancer cells via the c-Jun N-terminal kinase-mediated signaling pathway. Cancer Lett 179:51–58CrossRefGoogle Scholar
  4. 4.
    Mecham JO, Rowitch D, Wallace CD, Stern PH, Hoffman RM (1983) The metabolic defect of methionine dependence occurs frequently in human tumor cell lines. Biochem Biophys Res Commun 117:429–434CrossRefGoogle Scholar
  5. 5.
    Hoshiya Y, Guo H, Kubota T, Inada T, Asanuma F, Yamada Y et al (1995) Human tumors are methionine dependent in vivo. Anticancer Res 15:717–718PubMedGoogle Scholar
  6. 6.
    Sugimura T, Birnbaum SM, Winitz M, Greenstein JP (1959) Quantitative nutritional studies with water-soluble, chemically defined diets. VII. Nitrogen balance in normal and tumor-bearing rats following forced feeding. Arch Biochem Biophys 81:439–447CrossRefGoogle Scholar
  7. 7.
    Durando X, Thivat E, Farges MC, Cellarier E, D'Incan M, Demidem A et al (2008) Optimal methionine-free diet duration for nitrourea treatment: a phase I clinical trial. Nutr Cancer 60:23–30CrossRefGoogle Scholar
  8. 8.
    Epner DE, Morrow S, Wilcox M, Houghton JL (2002) Nutrient intake and nutritional indexes in adults with metastatic cancer on a phase I clinical trial of dietary methionine restriction. Nutr Cancer 42:158–166CrossRefGoogle Scholar
  9. 9.
    Thivat E, Farges MC, Bacin F, D'Incan M, Mouret-Reynier MA, Cellarier E et al (2009) Phase II trial of the association of a methionine-free diet with cystemustine therapy in melanoma and glioma. Anticancer Res 29:5235–5240PubMedGoogle Scholar
  10. 10.
    Strekalova E, Malin D, Good DM, Cryns VL (2015) Methionine deprivation induces a targetable vulnerability in triple-negative breast cancer cells by enhancing TRAIL receptor-2 expression. Clin Cancer Res 21:2780–2791CrossRefGoogle Scholar
  11. 11.
    Nair P, Lu M, Petersen S, Ashkenazi A (2014) Apoptosis initiation through the cell-extrinsic pathway. Methods Enzymol 544:99–128CrossRefGoogle Scholar
  12. 12.
    Camidge DR, Herbst RS, Gordon MS, Eckhardt SG, Kurzrock R, Durbin B et al (2010) A phase I safety and pharmacokinetic study of the death receptor 5 agonistic antibody PRO95780 in patients with advanced malignancies. Clin Cancer Res 16:1256–1263CrossRefGoogle Scholar
  13. 13.
    Herbst RS, Eckhardt SG, Kurzrock R, Ebbinghaus S, O'Dwyer PJ, Gordon MS et al (2010) Phase I dose-escalation study of recombinant human Apo2L/TRAIL, a dual proapoptotic receptor agonist, in patients with advanced cancer. J Clin Oncol 28:2839–2846CrossRefGoogle Scholar
  14. 14.
    Herbst RS, Kurzrock R, Hong DS, Valdivieso M, Hsu CP, Goyal L et al (2010) A first-in-human study of conatumumab in adult patients with advanced solid tumors. Clin Cancer Res 16:5883–5891CrossRefGoogle Scholar
  15. 15.
    Trarbach T, Moehler M, Heinemann V, Kohne CH, Przyborek M, Schulz C et al (2010) Phase II trial of mapatumumab, a fully human agonistic monoclonal antibody that targets and activates the tumour necrosis factor apoptosis-inducing ligand receptor-1 (TRAIL-R1), in patients with refractory colorectal cancer. Br J Cancer 102:506–512CrossRefGoogle Scholar
  16. 16.
    Toft DJ, Cryns VL (2011) Minireview: basal-like breast cancer: from molecular profiles to targeted therapies. Mol Endocrinol 25:199–211CrossRefGoogle Scholar
  17. 17.
    Lev DC, Kiriakova G, Price JE (2003) Selection of more aggressive variants of the gI101A human breast cancer cell line: a model for analyzing the metastatic phenotype of breast cancer. Clin Exp Metastasis 20:515–523CrossRefGoogle Scholar
  18. 18.
    Moyano JV, Evans JR, Chen F, Lu M, Werner ME, Yehiely F et al (2006) αB-crystallin is a novel oncoprotein that predicts poor clinical outcome in breast cancer. J Clin Invest 116:261–270CrossRefGoogle Scholar
  19. 19.
    Malin D, Chen F, Schiller C, Koblinski J, Cryns VL (2011) Enhanced metastasis suppression by targeting TRAIL receptor 2 in a murine model of triple-negative breast cancer. Clin Cancer Res 17:5005–5015CrossRefGoogle Scholar
  20. 20.
    Mentch SJ, Mehrmohamadi M, Huang L, Liu X, Gupta D, Mattocks D et al (2015) Histone methylation dynamics and gene regulation occur through the sensing of one-carbon metabolism. Cell Metab 22:861–873CrossRefGoogle Scholar
  21. 21.
    Malin D, Strekalova E, Petrovic V, Deal AM, Al Ahmad A, Adamo B et al (2014) αB-crystallin: a novel regulator of breast cancer metastasis to the brain. Clin Cancer Res 20:56–67CrossRefGoogle Scholar
  22. 22.
    Halo TL, McMahon KM, Angeloni NL, Xu Y, Wang W, Chinen AB et al (2014) NanoFlares for the detection, isolation, and culture of live tumor cells from human blood. Proc Natl Acad Sci U S A 111:17104–17109CrossRefGoogle Scholar
  23. 23.
    Fu X, Le P, Hoffman RM (1993) A metastatic orthotopic-transplant nude-mouse model of human patient breast cancer. Anticancer Res 13(4):901–904PubMedGoogle Scholar
  24. 24.
    Li X, Wang J, Yang M, Baranov E, Jinag P, Sun F, Moussa AR, Hoffman RM (2002) Optically imageable metastatic model of human breast cancer. Clin Exp Metastasis 19(4):347–350CrossRefGoogle Scholar
  25. 25.
    Puchalapalli M, Zeng X, Mu L, Anderson A, Hix Glickman L, Zhang M et al (2016) NSG mice provide a better spontaneous model of breast cancer metastasis than athymic (nude) mice. PLoS One 11:e0163521CrossRefGoogle Scholar
  26. 26.
    Harrell JC, Dye WW, Allred DC, Jedlicka P, Spoelstra NS, Sartorius CA et al (2006) Estrogen receptor positive breast cancer metastasis: altered hormonal sensitivity and tumor aggressiveness in lymphatic vessels and lymph nodes. Cancer Res 66:9308–9315CrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2019

Authors and Affiliations

  • Elena Strekalova
    • 1
  • Dmitry Malin
    • 1
  • Harisha Rajanala
    • 1
  • Vincent L. Cryns
    • 1
    Email author
  1. 1.Department of Medicine, University of Wisconsin Carbone Cancer CenterUniversity of Wisconsin School of Medicine and Public HealthMadisonUSA

Personalised recommendations