Abstract
Patients with KRAS mutated colorectal cancer (CRC) represent a cohort with unmet medical needs, with limited options of FDA-approved therapies. Representing 40–45% of all CRC patients, they are considered ineligible to receive anti-EGFR monoclonal antibodies that have added a significant therapeutic benefit for KRAS wild type CRC patients. Although several mouse models of CRC have been developed during the past decade, one genetically resembling the KRAS mutated CRC is yet to be established. In this study C57 BL/6 mice with truncated adenomatous polyposis coli (APC) floxed allele was crossed with heterozygous KRAS floxed outbred mice to generate an APCf/f KRAS+/f mouse colony. In another set of breeding, APC floxed mice were crossed with CDX2-Cre-ERT2 mice and selected for APCf/f CDX2-Cre-ERT2 after the second round of inbreeding. The final model of the disease was generated by the cross of the two parental colonies and viable APC f/f KRAS +/f CDX2-Cre-ERT2 (KPC: APC) were genotyped and characterized. The model animals were tamoxifen (TAM) induced to generate tumors. Micro-positron emission tomography (PET) scan was used to detect and measure tumor volume and standard uptake value (SUV). Hematoxylin and eosin (H&E) staining was performed to establish neoplasm and immunohistochemistry (IHC) was performed to determine histological similarities with human FFPE biopsies. The MSI/microsatellite stable (MSS) status was determined. Finally, the tumors were extensively characterized at the molecular level to establish similarities with human CRC tumors. The model KPC: APC animals are conditional mutants that developed colonic tumors upon induction with tamoxifen in a dose-dependent manner. The tumors were confirmed to be malignant within four weeks of induction by H&E staining and higher radioactive [18F] fluoro-2-deoxyglucose (FDG) uptake (SUV) in micro-PET scan. Furthermore, the tumors histologically and molecularly resembled human colorectal carcinoma. Post tumor generation, the KPC: APC animals died of cachexia and rectal bleeding. Implications: This model is an excellent preclinical platform to molecularly characterize the KRAS mutated colorectal tumors and discern appropriate therapeutic strategies to improve disease management and overall survival.
Highlights
Colorectal cancer (CRC) is initiated when the colonic epithelial loses the function of the adenomatous polyposis coli (APC) pathway which is essentially a part of Wnt signaling pathway [1,2]
A current view is that most colorectal cancers arise from adenomatous precursors and accumulate gain-of-function mutations in proto-oncogenes and loss-of-function mutations in tumor suppressor genes resulting in the initiation of adenomatous lesions and progression to carcinoma [6,7]
CDX2 ERT2 Cre mice were intercrossed with mice carrying loxP-flanked Apc
Summary
Colorectal cancer (CRC) is initiated when the colonic epithelial loses the function of the adenomatous polyposis coli (APC) pathway which is essentially a part of Wnt signaling pathway [1,2]. In the current study we used simple strategies to develop conditional genetic mouse models where both the alleles, namely floxed APC and mutated KRAS, can be simultaneously expressed by tamoxifen, which activates the large intestine specific transcription factor CDX2 through CRE recombinase fused to mutated estrogen receptor ERT2 [8]. When these mice are bred with mice containing loxP-flanked sequence, tamoxifen-inducible, Cre-mediated recombination will result in deletion of the floxed sequences in the Cre-expressing cells of the offspring Following this strategy, we have successfully generated and molecularly characterized the model to establish resemblance to human KRAS mutated CRC. This model is far the most robust model to investigate the KRAS mutated CRC
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