Abstract LB-36: Chemotherapy testing of primary human ovarian cancers in an ex vivo 3D culture platform: A novel method of phenotypic profiling for clinical trial selection and personalized medicine
Authors: Shuford Stephen , Widener Rebecca , Cheluvaraju Chaitra , Desrochers Teresa , mattingly christina , Puls Larry , gevaert matt , Orr David , Crosswell Hal E. 
Published: May 2015

Background: A goal of personalized medicine is to identify the most active and safest therapy for individual patients. One method for employing personalized medicine for cancer patients is through testing of a patient's cultured tumor tissue against a panel of predetermined drug candidates. To date, there is no standard platform by which an individual patient's tumor can be tested ex vivo in order to reliably predict if a certain drug or therapy is effective. We describe optimized methods of ex vivo 3D (EV3D™) culture and testing of primary human ovarian cancer for personalized medicine. Our hypothesis was that 3D spheroid cultures from primary human tumors would demonstrate unique growth parameters and distinct ex vivo testing responses compared to traditional 2D cultures. Materials & Methods: Optimization experiments: Methods for comparing 2D and 3D drug response, rapid spheroid formation across multiple cell types and scaffold/media conditions in both static and perfusion cultures were optimized using human cancer cell lines and reagents. Processing of Ovarian Cancers: Under informed consent, ovarian cancer samples were obtained and processed using standard mincing & digestion. Spheroids and 2D monolayers were established after viability assessment. Ex Vivo Testing & Analysis: cultured cells in 2D and 3D were exposed to a clinically relevant concentration range of cytotoxic or targeted agents. Multiple analytical techniques were applied including imaging, metabolism and dsDNA quantification; relative and absolute IC50s and total percent inhibition were used for ranking compounds. Results: Median subject age was 63(29-82), majority of which had advanced stage adenocarcinomas. Carboplatin & taxane based combination therapy was used in &gt;90% of patients. Clinical response and outcomes data collection are ongoing (median follow up 8 months). Spheroid formation was uniform across all malignant tumor types, but low grade lesions trended towards smaller, looser aggregates. All drug tested ex vivo samples were exposed to at least one agent that the subject received. Growth and response to positive control varied between 2D and 3D platforms. Doxorubicin and LY294002 showed greatest activity (median IC50 = 1, 0.8 uM, respectively) and cisplatin and topotecan the least (median IC50 16, 22). Median CA-125 fold reduction from baseline was 16 (range, 1.9-143). The subject with the greatest response by CA-125 levels (143 fold decrease) was predicted to respond to carboplatin-paclitaxel therapy in 3D culture but not in 2D (2D v 3D, p&lt;0.01).

Conclusions: EV3D allows for rapid and high throughput phenotypic profiling of novel small molecules (i.e. PI3K inhibitors) as well as conventional, FDA approved cytotoxic agents against patient-specific tumor samples in a relevant tumor microenvironment. EV3D cultures have reduced metabolism, decreased short term growth, and different drug-response profiles than 2D. Next generation exome sequencing of 39 drugable targets will allow genotypic-phenotypic correlation. Clinical data collection is ongoing to correlate ex vivo response with clinical outcomes. Current efforts are focused on developing EV3D as a novel small molecule phenotypic screen for clinical trials and as an in vitro chemo-sensitivity assay for personalized medicine.

Citation Format: Stephen Shuford, Rebecca Widener, Chaitra Cheluvaraju, Teresa Desrochers, christina mattingly, Larry Puls, matt gevaert, David Orr, Hal E. Crosswell. Chemotherapy testing of primary human ovarian cancers in an ex vivo 3D culture platform: A novel method of phenotypic profiling for clinical trial selection and personalized medicine. [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 LB-36. doi:10.1158/1538-7445.AM2014-LB-36