Abstract C236: The inhibitory effect of tumor microenvironment on oncolytic virotherapy in lung cancer.

Introduction: Oncolytic virotherapy can have a potentially high impact in the treatment of non-small cell lung cancer (NSCLC), as there are documented preclinical activities and target specificity. Oncolytic viruses have unique advantages over small molecular drugs, such as their ability to replicate, spread, and carry anti-tumoral therapeutic genes; however, their challenges include concerns over systemic toxicity, requiring intratumoral administration and restricting its applicability. This highlights the clinical significance of tumor microenvironment in oncolytic virotherapy. Although studies have shown significant stromal effects on carcinogenesis, the effects of the tumor microenvironment on viral susceptibility have been largely unstudied. Microscale 3D co-culture assays were recently developed and validated by our laboratory for studies on tumor invasion, and we have used them here to manipulate, control, and examine cell-cell interactions. Here we developed an in vitro microfluidic model to examine tumor-stromal interaction on oncolytic virotherapy. Methods: To determine the effect of tumor microenvironment on oncolytic virotherapy, we examined VSV infection in 3D microfluidic cultures of representative NSCLC cell lines (A549 lung adenocarcinoma, H226 squamous cell carcinoma) in vitro, either as a monoculture, a co-culture in combination with normal lung fibroblasts WI38, or a monoculture in WI38-conditioned media. The infection of VSV-GFP1, a recombinant virus that expresses green fluorescent protein as a byproduct of infection, in lung cancer cells was investigated using a novel microfluidic-based method. As GFP is only produced during viral replication, GFP fluorescence serves as a surrogate for viral infection. Time-lapse fluorescence microscopy of a single VSV-GFP plaque on a lung cancer cell monolayer in 3D microfluidic cultures was conducted over a 30-hour period post infection. Viral production was quantified using conventional plaque assays. Statistical analysis was based on the two-sample Student9s t-test at a two-tailed level of 0.05. Results: Up to 37% infection in 3D microfluidic monocultures of A549 lung adenocarcinoma cells compared to 12% infection in co-cultures with WI38 fibroblasts was observed over 30 hours post infection. The rate of VSV infection in A549 decreased significantly in the presence of WI38 lung fibroblasts (2% vs. 0.6% cells/hour; P=0.01). Up to 12% infection in 3D microfluidic monocultures of A549 in WI38-conditioned media was observed over 30 hours post infection; the rate of its VSV infection was equivalent to that in co-cultures with WI38 fibroblasts (0.6% cells/hour). VSV production decreased by 100 folds in A549 monocultures in WI38-conditioned media compared to that in conventional media. Conclusions: VSV spread and production in A549 lung adenocarcinoma cells decreased significantly in the presence of WI38 lung fibroblasts or WI38-conditioned media. As WI38-conditioned media have the same inhibitory effects on viral infection as do WI38 fibroblasts, secreted soluble factors appear to be more important than direct cell-cell contact in causing this phenomenon. Future studies on stroma-targeted therapy may alleviate the inhibitory effects of tumor microenvironment on virotherapy and increase the bioavailability of oncolytic viruses to the tumors systemically. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2011 Nov 12-16; San Francisco, CA. Philadelphia (PA): AACR; Mol Cancer Ther 2011;10(11 Suppl):Abstract nr C236.