Abstract 100: Establishment and characterization of allografts derived from a genetically engineered mouse model of non-small cell lung cancer (NSCLC)

Abstract

Abstract Introduction: Genetically engineered mouse models (GEMM) represent an attractive system for preclinical research since GEMM tumors develop in the presence of a competent immune system, thereby closely resembling the tumor microenvironment in patients. One example is the KP model (DuPage et al., 2009), which reflects the tumorigenesis of NSCLC in humans. KP tumors carry Kras and Trp53 mutations, comparable to subtypes of human NSCLC. Typical drawbacks of GEMM tumors include differences in their genetic make-up to human tumors and the very slow and heterogeneous tumor development, making the use of these models challenging for routine in vivo efficacy studies. The latter can be improved by grafting primary tumors on a genetically dissimilar member of the same strain, so-called allografting. Thus, the advantages of xenograft and GEMM models can be combined. Here, we described the establishment and characterization of allografts from the KP GEMM. Material & Methods: KP tumors were excised from the lungs of KP mice and implanted subcutaneously (s.c.) into C57BL/6N female mice. When tumor growth was detected, passaging steps from animal to animal were performed. Furthermore, two cell lines derived from KP tumors, KP1 and KP4 (provided by Müller et al.), were injected s.c., the developing tumors excised and directly re-implanted as described above. The passages were analyzed molecularly, histologically, and via IHC (e.g. Ki67 proliferation marker) , and compared to the original in-situ tumors. A therapeutic study with Erlotinib and BEZ235 was performed on KP1 allografted mice. Results: All investigated allografts showed similar histological patterns. Ki67 scores reflected the average time from implantation to passaging. The average passage times ranged from 19±3 days (KP4 cell line) to 26±6 days (KP1 cell line). Allografts from the original KP model showed a time to passaging of 23±5 days. The mRNA expression of EpCAM (CD326) was on average 10-fold higher in KP and KP1 allografted tumors compared to normal C57BL/6 lung tissue, which could be confirmed by flow cytometry and cell-binding assay, determining >99% EPCAM positive cells in KP1 tumors. All allografts carried the Kras mutation and loss of function point mutations in Trp53. The therapy study revealed no effect of Erlotinib on tumor growth, but a significant (p=0.0003) tumor load reduction in mice treated with BEZ235 (519 ± 244 mm³, n=5) compared to the control group (1678 ± 275 mm³, n=4) on day 28, respectively. Conclusion: The generation of s.c. transplantable allografts from the KP GEMM was successfully conducted. The allograft models enable the use of immunocompetent mice in a feasible time frame and still resemble key aspects of human disease. Further studies will elucidate the interaction of the allografts with the host and compare sensitivity characteristics between GEMM and allograft of the same model. Citation Format: Damaris Kukuk, Philipp Müller, Kerstin Klingner, Anne-Lise Peille, Alfred Zipelius, Julia B. Schüler. Establishment and characterization of allografts derived from a genetically engineered mouse model of non-small cell lung cancer (NSCLC). [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 100. doi:10.1158/1538-7445.AM2014-100