Abstract 321: In vitro recapitulation of 3D tumor microenvironment with defined oxygen and pH levels through a novel scalable bioreactor-based strategy
Authors: Santo Vitor E. , Estrada Marta , Rebelo Sofia , Anderson Elizabeth , Alves Paula M. , Brito Catarina 
Published: August 2015

Abstract
Drug discovery for cancer therapy has faced tremendous inefficacy at advanced stages of clinical trials. The scientific community has been trying to tackle this issue by improving preclinical models for target validation, in which the complexity and heterogeneity found in human tumours can be recapitulated. In particular, the tumour microenvironment is known to influence tumour progression and drug resistance through tumour-stroma crosstalk. Three dimensional (3D) cell models allow maintenance of cell-cell and cell-extracellular matrix (ECM) interactions, mimicking in vivo tissue organization. However, current 3D cell models are typically generated in non-scalable culture systems, with poor robustness and use undefined, biologically active matrices. In this work, we present a novel strategy for in vitro recapitulation of solid tumour microenvironments.
H1650 (human NSCLC lung adenocarcinoma) and MCF7 (human ER+ breast cancer) cells were selected for the establishment of lung and breast cancer models respectively. An alginate microencapsulation strategy for co-culture of either lung or breast tumour cells with fibroblasts was developed. The constructs were then cultured in stirred-tank bioreactors, with precise definition and control of oxygen and pH levels for several weeks, and characterized by analysis of cell viability, proliferation and apoptosis along culture time, as well as metabolic profiling, phenotypic characterization and analysis of ECM deposition
The strategy developed allowed the formation of stable hydrogel microcapsules, with tumour aggregates presenting an epithelial phenotype with partial polarization, surrounded by fibroblasts, recapitulating tumour-stroma organization in vivo. Co-cultures of lung or breast cancer cell aggregates with fibroblasts led to the deposition and accumulation of collagen I type inside the capsules, along with altered tumour cell phenotype, with loss of epithelial character. Perturbation studies with chemotherapeutic agents revealed differences between models established with mono-cultures of tumour cells and co-cultures of tumour cells and fibroblasts.
In conclusion, we have developed a scalable, robust and versatile strategy for establishment of in vitro long-term recapitulation of tumour-stroma crosstalk and hypoxic microenvironment, providing tools for characterization of disease progression mechanisms, target validation and drug response in different cancer types.
Acknowledgements: The research leading to these results has received support from the Innovative Medicines Initiative Joint Undertaking (grant agreement n° 115188), resources composed of financial contribution from EU- FP7and EFPIA companies in kind contribution. M.Estrada acknowledges support from FCT, Portugal (SFRH/BD/52208/2013).The authors are also thankful to Cathrin Brisken, Heiko van der Kuip, Moshe Oren and Erwin Boghaert.
Citation Format: Vitor E. Santo, Marta Estrada, Sofia Rebelo, Elizabeth Anderson, Paula M. Alves, Catarina Brito. In vitro recapitulation of 3D tumor microenvironment with defined oxygen and pH levels through a novel scalable bioreactor-based strategy. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 321. doi:10.1158/1538-7445.AM2015-321