Stromal cells play a critical role in the outcome of the oncolytic efficacy of conditional replicative adenovirus.

M. VERÓNICA LOPEZ; DIEGO VIALE; EDUARDO G. CAFFERATA; OSVALDO L. PODHAJCER

Jerusalem, Israel

Congreso; Translational Cancer Medicine.; 2008

AACR

Treatment of advanced cancer remains a formidable task. Oncolytic adenoviruses appeared recently as a strategy to directly and specifically target cancer cells. Conditional Replicative Adenoviruses (CRAds) replication is driven by promoters corresponding to genes differentially expressed in the tumor tissue. These CRAds have demonstrated marked safety in clinical trials although their efficacy has been limited. In order to establish if tumor associated-stromal cells might affect CRAd oncolytic efficacy, we established a model based on the administration in nude mice of human malignant cells in combination with transformed endothelial cells or fetal fibroblast as a strategy to establish the role stromal cells may play on CRAd activity. In this novel CRAd (AdF512-TK), E1A expression is driven by a 0.5 Kb fragment of the SPARC promoter, since this gene is overexpressed in malignant and cancer associated-stromal cells in different human cancers, including melanoma. AdF512-TK is a type 5 adenovirus armed with the Herpes Simplex thymidine kinase gene. AdF512-TK(+GCV) was lytic on human melanoma cells expressing high levels of SPARC. In addition, AdF512-TK(+GCV) was cytotoxic at high MOI on transformed human microendothelial cells (HMEC-1) and WI-38 fetal fibroblasts but its activity was completely attenuated in normal melanocytes, keratinocytes, mammary and colonic cells and adult fibroblasts. On the contrary, adenovirus wild type (Ad-WT) lysed every cell type. AdF512-TK(+GCV) administration o nude mice carrying established tumors made of SB2 human melanoma cells alone induced the elimination of 80% of tumor (8/10). Contrary to that, AdF512-TK +GCV induced only a delay in tumor growth when established tumors were composed of SB2 cells mixed either with HMEC-1 or WI-38 demonstrating that the presence of stromal cells hampered viral oncolytic efficacy. We next asked whether this CRAd could be active in human cancer cells in which SPARC expression is silenced due to promoter hypermethylation such as in pancreatic cancer cells. Treatment of mice carrying established MIA PaCa-2 pancreatic tumors made of malignant cells alone led to growth inhibition in 2/7 cases. Surprisingly 5/6 mice showed completed arrest of tumor growth when tumors were composed of malignant and HMEC-1 cells, indicating that the presence of microendothelial cells enhanced CRAd activity. Indeed, conditioned media (CM) produced either HMEC-1 or WI-38 cells enhanced SPARC promoter and CRAd oncolytic activity in MIA PaCa-2 cells but not in SB2 melanoma cells. Extended analysis of this effect demonstrated that CM produced by stroma cells had not effect of was inhibitory of CRAd activity on MEL- and A375N melanoma cells while CM obtained from WI-38 but not HMEC-1 enhanced CRAd activity on T47D mammary tumor cells. We are currently identifying potential soluble factors produced by stroma cells that might modulate CRAd activity. We conclude that the presence of stroma cells that in some human tumors account for 90% of the tumor, have a critical role in defining the therapeutic efficacy of an oncolytic virus.