Abstract 150: A 515 tumor specimens collection from surgery to benchtop: the WFORCE organoid experience in personalized research
Authors: Forsythe Steven Donald , Erali Richard A. , Laney Preston , Meeker William , Abdulkhuseynova Salomat , Wajih Nadeem , Edenhoffer Nicholas , Schaaf Cecilia R. , Sivakumar Hemamyammal , Skardal Aleksander , Strowd Roy E. , Tatter Stephen B. , Clark Clancy J. , Randle Reese W. , Shen Perry , Levine Edward A. , D'Agostino Ralph B. , Soker Shay , Votanopoulos Konstantinos I. 
Published: April 2023

Abstract
Introduction: Patient tumor organoids (PTOs) are a new and developing preclinical model in cancer research. However, there is variation in protocols between centers and lack of consistency in important variables, impacting PTO culture success. In this study, we analyzed factors in clinical data and tumor tissue processing to determine optimal conditions for cell isolation and PTO success.
Methods: Under IRB approval, tumor tissues from primary and metastatic sites were procured in clinically indicated surgeries then dissociated using a standardized protocol. PTOs were fabricated using collagen-hyaluronic acid ECM and underwent drug screening at 10 days post fabrication, where viability was assessed. Clinical and tissue processing inputs such as type of primary, peritoneal carcinomatosis index (PCI), prior surgery or neoadjuvant chemotherapy, presence of mucin, tissue digestion time, specimen mass, digestion time, and the diameter of isolated cells were analyzed. These factors were assessed by calculating the Pearson Correlation Coefficient (r) and best fit modeling to determine the effect on outputs including tissue viability as measured by % of viable cells using an automated cellular counter, viable cells per mg tissue, and PTO log10 viability by Promega 3D Cell Titer Glo assay.
Results: From December 2018-July 2022, 515 tumors from 286 patients with 25 primary types were processed for cellular isolation and PTO fabrication. 407 (79%) provided PTOs for therapeutic testing, with 354 (69%) providing sufficiently viable PTOs for therapeutic interrogation. The most common primary tumor type accrued was appendiceal cancer (n=223, 43%). Tissues processed at 24 hours did not differ in viability outputs when compared to tissues processed immediately (p&gt;0.05). Tissue % viability was better maintained in larger specimens (r=.18, p&lt;0.001), resulting in more viable cells per mg of tissue (r=.12, p=0.009) while it was negatively related to larger cell diameter (r=-.32, p&lt;0.001). The number of viable cells per mg of tumor correlated to higher post-processing cell counts (r=.49, p&lt;0.001) while it was decreased by longer digestion times (r=-.11, p=0.017). Finally, PTO log10 viability, correlated with higher burden of disease as measured by PCI scores (r=.35, p&lt;0.001), larger specimens (r=.19, p&lt;0.001), mucinous tumors (PTO log10 viability 5.46 vs 5.17, p=0.01), and tissues with higher cellular viability (r=.31, p&lt;0.001), while it was negatively impacted by neoadjuvant chemotherapy (5.06 vs 5.41, p&lt;0.001).
Conclusions: Higher rates of successful PTO fabrication are obtained in patients with untreated tumors, more aggressive biologic behavior, and increased tumor burden. Creating a unified protocol for tissue processing, agnostic to tumor type, will facilitate incorporation of tissue from the operating room to research protocols with wider translational implications.
Citation Format: Steven Donald Forsythe, Richard A. Erali, Preston Laney, William Meeker, Salomat Abdulkhuseynova, Nadeem Wajih, Nicholas Edenhoffer, Cecilia R. Schaaf, Hemamyammal Sivakumar, Aleksander Skardal, Roy E. Strowd, Stephen B. Tatter, Clancy J. Clark, Reese W. Randle, Perry Shen, Edward A. Levine, Ralph B. D'Agostino, Shay Soker, Konstantinos I. Votanopoulos. A 515 tumor specimens collection from surgery to benchtop: the WFORCE organoid experience in personalized research [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 150.