A novel ex vivo tumor system identifies Src-mediated invasion and metastasis in mesenchymal tumor cells in non-small cell lung cancer

Aparna Padhye,Don L Gibbons,Jordan S Miller,Jared J Fradette,Christin Ungewiss,Jacob L Albritton,B Leticia Rodriguez

doi:10.1038/s41598-019-41301-2

Abstract

Lung cancer is the foremost cause of cancer related deaths in the U.S. It is a heterogeneous disease composed of genetically and phenotypically distinct tumor cells surrounded by heterotypic cells and extracellular matrix dynamically interacting with the tumor cells. Research in lung cancer is often restricted to patient-derived tumor specimens, in vitro cell cultures and limited animal models, which fail to capture the cellular or microenvironment heterogeneity of the tumor. Therefore, our knowledge is primarily focused on cancer-cell autonomous aberrations. For a fundamental understanding of lung cancer progression and an exploration of therapeutic options, we focused our efforts to develop an Ex Vivo Tumor platform to culture tumors in 3D matrices, which retains tumor cell heterogeneity arising due to in vivo selection pressure and environmental influences and recapitulate responses of tumor cells to external manipulations. To establish this model, implanted syngeneic murine tumors from a mutant KRAS/p53 model were harvested to yield multicellular tumor aggregates followed by culture in 3D extracellular matrices. Using this system, we identified Src signaling as an important driver of invasion and metastasis in lung cancer and demonstrate that EVTs are a robust experimental tool bridging the gap between conventional in vitro and in vivo models.

Highlights

Lung cancer has the highest mortality rate of all cancer types[1] primarily because two-thirds of the patients present at a stage when the cancer has already metastasized to distant organs
Prior to embedding in 3D matrices, Ex Vivo Tumor (EVT) were cultured in laser ablated poly(dimethylsiloxane) (PDMS) microwell inserts designed for standard tissue culture plates, which we have previously described for single-cell suspensions[13]
Tumors are not just a mass of cancer cells, but instead are composed of a heterogeneous tumor microenvironment, including changes in the extracellular matrix (ECM) and heterotypic cell types, that dynamically interacts with tumor cells

Summary

Introduction

Lung cancer has the highest mortality rate of all cancer types[1] primarily because two-thirds of the patients present at a stage when the cancer has already metastasized to distant organs. In order to address these deficiencies in current lung cancer models, we established an Ex Vivo Tumor (EVT) platform to culture lung tumors in 3D matrices This system has specific advantages over the more commonly used in vitro and in vivo systems. Using syngeneic models derived from these GEMMs, we have previously demonstrated that upon loss of the microRNA-200 family, the mesenchymal tumor cells are dependent on the interaction of the cell adhesion molecule integrin β1 and the extracellular matrix component collagen type I. This interaction drives the formation of the focal adhesion complex through recruitment of the adaptor molecule CRKL, which is a direct miR-200 target[10]. The system presented here can be extended to identify and understand other novel signaling pathways that regulate malignant progression or define therapeutic sensitivities in lung cancer

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