Luciferase Expressing Preclinical Model Systems Representing the Different Molecular Subtypes of Colorectal Cancer.

Abstract

Colorectal cancer (CRC) is a heterogeneous disease. More insight into the biological diversity of CRC is needed to improve therapeutic outcomes. Established CRC cell lines are frequently used and were shown to be representative models of the main subtypes of CRC at the genomic and transcriptomic level. In the present work, we established stable, luciferase expressing derivatives from 10 well-established CRC cell lines, generated spheroids and subcutaneous xenograft tumors in nude mice, and performed comparative characterization of these model systems. Transcriptomic analyses revealed the close relation of cell lines with their derived spheroids and xenograft tumors. The preclinical model systems clustered with patient tumor samples when compared to normal tissue thereby confirming that cell-line-based tumor models retain specific characteristics of primary tumors. Xenografts showed different differentiation patterns and bioluminescence imaging revealed metastatic spread to the lungs. In addition, the models were classified according to the CMS classification system, with further sub-classification according to the recently identified two intrinsic epithelial tumor cell states of CRC, iCMS2 and iCMS3. The combined data showed that regarding primary tumor characteristics, 3D-spheroid cultures resemble xenografts more closely than 2D-cultured cells do. Furthermore, we set up a bioluminescence-based spheroid cytotoxicity assay in order to be able to perform dose-response relationship studies in analogy to typical monolayer assays. Applying the established assay, we studied the efficacy of oxaliplatin. Seven of the ten used cell lines showed a significant reduction in the response to oxaliplatin in the 3D-spheroid model compared to the 2D-monolayer model. Therapy studies in selected xenograft models confirmed the response or lack of response to oxaliplatin treatment. Analyses of differentially expressed genes in these models identified CAV1 as a possible marker of oxaliplatin resistance. In conclusion, we established a combined 2D/3D, in vitro/in vivo model system representing the heterogeneity of CRC, which can be used in preclinical research applications.