Development of a mouse model for sporadic and metastatic colon tumors and its use in assessing drug treatment

Abstract

Most genetically engineered mouse (GEM) models for colon cancer are based on tissuewide or germline gene modification, resulting in tumors predominantly of the small intestine. Several of these models involve modification of the adenomatous polyposis coli (Apc) gene and are excellent models for familial cancer predisposition syndromes. We have developed a stochastic somatic mutation model for sporadic colon cancer that presents with isolated primary tumors in the distal colon and recapitulates the entire adenoma-carcinoma-metastasis axis seen in human colon cancer. Using this model, we have analyzed tumors that are either solely mutant in the Apc gene or in combination with another colon cancer-associated mutant gene, the Kras G12D allele. Because of the restricted location in the distal colon, the natural history of the tumors can be analyzed by serial colonoscopy. As the mammalian target of rapamycin (mTOR) pathway is a critical component of the complex signaling network in colon cancer, we used this model to assess the efficacy of mTOR blockade through rapamycin treatment of mice with established tumors. After treatment, Apc mutant tumors were more than 80% smaller than control tumors. However, tumors that possessed both Apc and Kras mutations did not respond to rapamycin treatment. These studies suggest that mTOR inhibitors should be further explored as potential colorectal cancer therapies in patients whose tumors do not have activating mutations in KRAS.

Fig. 1.

Adeno-cre treatment of floxed Apc mice results in distal colonic tumors. (A) Gross picture of solitary tumor (red arrow) after adeno-cre infection of floxed Apc mice in distal colon. Resulting primary tumors range from (B) adenomas to (C) carcinomas. (D) Immunohistochemistry nuclear β-catenin staining (red arrows) in tumors indicating activation of Wnt pathway.

Fig. 2.

Adeno-cre treatment of Apc CKO/LSL-Kras mice accelerates tumor progression. Adeno-cre administered to the distal colon of Apc CKO/LSL-Kras mice results in primary tumors that present as (A) adenomas, (B) carcinomas, and (C) invasive carcinomas. Furthermore, in a subset of animals (D) gross (white arrows) and (E) histological liver metastases (red arrows) are seen. Nuclear staining for cdx-2 demonstrates (F) primary and (G) metastatic tumors are of intestinal origin. (H) Nuclear β-catenin staining (red arrows) demonstrates Wnt pathway is activated in metastatic tumors.

Fig. 3.

Endoscopic assessment of mouse sporadic colon cancers. (A) Representative serial endoscopic images after adeno-cre administration to the distal colon of Apc CKO mice. (B) Tumor growth curves of adeno-cre-induced Apc and Apc/Kras mutant mice do not show a significant difference (P = 0.406). (C) Early endoscopic determination in Apc CKO and Apc CKO/LSL-Kras animals after adeno-cre induction shows an increase in tumor multiplicity (P = 0.02). Endoscopic imaging of mouse colon tumors (red arrows) after injection with Prosense 680: (D) white light imaging and (E) near-infrared imaging.

Fig. 4.

Rapamycin treatment blocks the mTOR pathway in Apc and Apc-Kras mutant tumors. Phospho-S6 levels are decreased after rapamycin treatment in both Apc and Apc-Kras mutant tumors by (A) Western blot analysis and (B) immunohistochemistry.

Fig. 5.

mTOR blockade results in tumor regression that is dependent upon Kras mutational status. Endoscopic growth curves demonstrate decrease in tumor size with rapamycin treatment in (A) Apc mutant tumors (P = 0.04) but not (B) Apc-Kras (P = 0.26).

Fig. 6.

Apc-Kras tumors show increased levels of MAPK activation. Phospho-ERK levels are increased in Apc-Kras tumors by (A) Western blot analysis and (B) immunohistochemistry.