Abstract LB-135: New cancer prevention model using imaging for early detection of mesothelin-expressing cancers
Authors: Sharma Khushboo , Yin Catherine , Kamat Kalika , Repellin Claire , Sambucetti Lidia C. , Scholler Nathalie 
Published: July 2017

Abstract
A major challenge to the discovery and development of novel cancer prevention strategies is the availability of reliable animal models. Key elements for chemoprevention testing in animal include relevance to human cancers, with tumors of similar pathology and genetic abnormalities, the presence of intermediate lesions that resemble human cancer development both histologically and molecularly; and consistent tumor incidence of &gt;60% within less than 6 months. To our knowledge no established animal model responds to these requirements for ovarian cancer prevention. We sought to adapt the Dicerflox/flox-Ptenflox/flox double knockout (DKO) ovarian cancer mouse model (Kim, Matzuk PNAS 2012) for cancer prevention studies, by synchronized induction of the disease with intra-ovary injection of an adenovirus encoding CRE protein (Ad-mCherry-Cre). Premalignant lesions in DKO AdCRE mice were visible by H&E staining after 12 weeks, and evolved into typical serous ovarian cancer by 24 weeks. Histology demonstrated that tumors overexpressed cytokeratin 8, cyclin D, CA125, and mesothelin. In addition, CA125, the main biomarker for ovarian cancer in woman, was consistently elevated in the serum of DKO AdCRE mice 15 weeks after CRE induction, but not in age-matched DKO mice. To assess whether mesothelin-targeted imaging could help monitor cancer development in DKO AdCRE mice, we used an anti-mesothelin recombinant antibody derived from llama (G3a nanobody) that we previously identified and in vitro characterized (Prantner, Scholler, JBN 2015). G3a nanobody is conveniently secreted by yeast with an N-terminal biotinylation (Scholler, JIM 2006) for streptavidin coupling. We addressed here whether G3a nanobody could detect mesothelin-expressing tumors using in vivo imaging (IVIS). Ni-purified G3a nanobody was incubated with fluorescently-labelled streptavidin (IRB680W) for 30 min and injected retro orbitally just before imaging. We found that G3a nanobody generated a robust and mesothelin-specific signal that peeked 4 hours after injection, and detected DKO AdCRE tumors as early as 8 weeks after CRE induction. G3a nanobody specific detection of mesothelin was confirmed in other mouse models of cancer, including in syngeneic models of ovarian cancer induced by orthotopic or intraperitoneal injection of mesothelin+ Luc-ID8 cells, and in xenograft models of human lung cancer obtained by IV injection of mesothelin high EKVX, mesothelin int H460, and mesothelin lo A549 cells. In summary, our results indicate that G3a nanobody is a valuable reagent for early detection of mesothelin-expressing cancer, and that DKO mice induced for Pten and Dicer double knockout by intra ovarian injection of CRE adenovirus develop serous ovarian cancer that express CA125 and mesothelin. We propose this model for in vivo monitoring of cancer development, as a novel system model for ovarian cancer prevention studies.
Citation Format: Khushboo Sharma, Catherine Yin, Kalika Kamat, Claire Repellin, Lidia C. Sambucetti, Nathalie Scholler. New cancer prevention model using imaging for early detection of mesothelin-expressing cancers [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr LB-135. doi:10.1158/1538-7445.AM2017-LB-135