Abstract A02: A novel orthotopic ovarian patient derived xenograft model platform to investigate novel therapies for BRCA deficient ovarian cancers
Authors: George Erin , Kim Hyoung , Tanyi Janos , Ragland Ryan , Brown Eric , Zhang Rugang , Brafford Patricia , Sproesser Katrin , Beqiri Marilda , Vultur Adina , Krepler Clemens , Weis Brandon , Nathanson Kate , Lu Yuling , Mills Gordon , Makvandi Mehran , Mach Robert , Morgan Mark , Simpkins Fiona 
Published: March 2017

Abstract
Introduction: To create a personalized, targeted approach to high grade serous ovarian cancers (HGSOC), reliable preclinical models are essential. About ~50% of HGSOC have defects in genes involved in homologous recombination (HR) such as BRCA. PARP inhibitors (PARPi) capitalize on synthetic lethality in HR-deficient tumors, however, clinical efficacy is limited (response rate only ~40%). Patient derived xenografts (PDXs) are emerging as a reliable preclinical model that recapitulates principal characteristics of the patients' tumor while remaining biologically stable while passaged in mice. We developed a BRCA1/2 orthotopic PDX experimental platform to study alternative strategies for synthetic lethality. We hypothesized that targeting the ATR/CHK1 axis is synthetically lethal in BRCA mutant HGSOC models.
Experimental Procedures: Fresh HGSOC tumor was transplanted orthotopically to the fallopian tube/ovary of NSG 5-8 wk mice. Tumor growth was followed. Tumors were evaluated by IHC, genomic and proteomic analysis. Alu II probe staining was used to evaluate human stroma content. DNA sequencing was performed using a 153 OVCA gene panel. Reverse Phase Protein Array Analysis (RPPA) was evaluated for signaling pathway activation. Primary ovarian tumor cultures were developed from patients' tumor for mechanistic studies. To study the ATR/CHK1 axis in HR-deficient HGSOC, PARPi (Olaparib), CHK1 inhibitor (CHK1i, MK8776), and ATR inhibitor (ATRi, AZD6738) were evaluated. PEO1 (BRCA2 mutant), PEO4 (BRCA wildtype) and JHOS4 (BRCA1 mutant) HGSOC cells were evaluated for cell proliferation, survival, and genome stability before and after treatment. BRCA2 mutant (8945delAA) PDX (WO-2-1) was expanded in 70 mice. Mice were randomized into 5 gps: untreated, carboplatin, PARPi, CHK1i, and ATRi. Treatment was initiated when tumors were 70-100mm3 and volume was assessed weekly with ultrasound. PARP tumor activity and response to PARPi was assessed with a PET PARP1 radiotracer [18F]FTT (fluorthanatrace).
Results: We developed a pipeline to study HR deficient HGSOC. We created an orthotopic PDX platform from 15 BRCA mutant patients in order to accurately study OVCA tumorigenicity and metastasis in the native environment with a 90% take rate in generating tumors in mouse passage 1 (MP1), and 100% take rate for MP2 and MP3. The PDX model (WO-2-1) was similarly platinum sensitive as the patient after platinum treatment. Tumors were evaluated by genomic and proteomic analysis to identify a target population and streamline therapeutic approaches. Pathogenic mutation profiles from the original patient tumor were preserved in PDXs serially passaged (MP1-3). High pCHK1 (s345) was used as a marker for investigation of ATR/CHK1 inhibition in BRCA mutant PDX models. We showed that ATRi and CHK1i are similarly effective to PARPi in a BRCA2 mutant PDX. A novel PET PARP1 radiotracer [18F]FTT was used and demonstrated co-localization of signal in a BRCA2 mutant PDX, which was diminished with olaparib treatment.
Conclusions: Although technically more challenging, the orthotopic transplantation technique is feasible in generating HGSOC PDX models with a high success rate that more closely resembles the natural environment for HGSOC progression. Evaluation of genomic and proteomic profiles of a tumor allows one to streamline targeted therapies for testing in PDX preclinical trials that may in the future be translated back to the patient.
Citation Format: Erin George, Hyoung Kim, Janos Tanyi, Ryan Ragland, Eric Brown, Rugang Zhang, Patricia Brafford, Katrin Sproesser, Marilda Beqiri, Adina Vultur, Clemens Krepler, Brandon Weis, Kate Nathanson, Yuling Lu, Gordon Mills, Mehran Makvandi, Robert Mach, Mark Morgan, Fiona Simpkins. A novel orthotopic ovarian patient derived xenograft model platform to investigate novel therapies for BRCA deficient ovarian cancers. [abstract]. In: Proceedings of the AACR Special Conference: Patient-Derived Cancer Models: Present and Future Applications from Basic Science to the Clinic; Feb 11-14, 2016; New Orleans, LA. Philadelphia (PA): AACR; Clin Cancer Res 2016;22(16_Suppl):Abstract nr A02.