Abstract A15: Establishing an ex-vivo, tumor spheroid culture system to assess molecular-targeted therapies for personalized treatment of non-small cell lung cancer
Authors: Banda Malathi , McKim Karen L. , Parsons Barbara 
Published: March 2017

Molecularly-targeted therapeutics are being developed for personalized cancer treatment based on the ideas that treatments should be determined by specific characteristics of an individual patient's tumor and that treatments should target tumor cells without damaging normal cells. Accumulating evidence indicates tumor heterogeneity can lead to acquired resistance to molecularly-targeted therapies. Currently, preclinical investigations rely heavily on the use of tumor cell lines and mouse xenograft models. The high failure rate in oncology drug development suggests that such test systems do not adequately model tumor heterogeneity. Therefore, more predictive preclinical models are needed to ensure that only the most promising molecularly-targeted, combination therapies are assessed in clinical studies. Here we established a novel 3D lung adenocarcinoma tissue-originating spheroid model, which may be used to investigate the efficacy of molecularly-targeted cancer therapeutics. Fresh lung adenocarcinomas were minced and either digested with Liberase DH enzyme (5.6 units) or homogenized to generate cancer tissue-originating spheroids. Spheroids were plated in Matrigel and cultured in stem cell media, with 0.01, 0.1, 1 or 10 μM erlotinib or the DMSO vehicle. Erlotinib is an epidermal growth receptor inhibitor commonly used to treat non-small cell lung cancer (NSCLC). Using an ImageXpress Micro Widefield High Content Screening System (Molecular devices), z-stack bright field images were collected 1, 7, and 14 days after plating. Composite z-stack images were analyzed using LASX (Leica Microsystems) software. Total spheroid area (in pixels) was measured for replicate cultures exposed to different erlotinib concentrations at each time point. The total area of spheroids for 7 and 14 days were normalized to day 1 measurements and cell death was evaluated as the decrease in total area of the spheroids in culture. Our results indicate that direct homogenization of lung adenocarcinomas generated at least ten times greater yield of spheroids than was achieved using digestion with the Liberase DH. The yield and appearance of spheroids varied significantly. Spheroids prepared from some lung adenocarcinomas were more spherical and aggregated than those prepared from other tumors. There was time-dependent decrease in total area of spheroids following 7 and 14 days in culture. Also spheroids prepared from different lung adenocarcinomas responded differently to different concentrations of erlotinib. High-sensitivity quantification of specific mutations (EGFR, KRAS PIK3CA and BRAF) will be conducted before and after erlotinib treatments to determine whether this model can be used to assess the potential outgrowth of drug-resistant tumor subpopulations.

Citation Format: Malathi Banda, Karen L. McKim, Barbara Parsons. Establishing an ex-vivo, tumor spheroid culture system to assess molecular-targeted therapies for personalized treatment of non-small cell lung cancer. [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 A15.