TARGETING MALIGNANT AND TUMOR ASSOCIATED-STROMAL CELLS AS A NOVEL STRATEGY FOR CANCER GENE THERAPY

MARIA VERONICA LOPEZ; DIEGO VIALE; EDUARDO G. CAFFERATA; HONGJU WU; ANGEL A. RIVERA; DAVID GOULD; YUTI CHERNAJOVSKY; DAVID T. CURIEL; OSVALDO L. PODHAJCER

Cold Spring Harbor New York

Congreso; In vivo barriers to gene delivery; 2007

Cold Spring Harbor Laboratory

SPARC is a matricellular protein that is overexpressed in malignant and tumor associated-stromal cells in human melanomas and other cancers as well. By using luciferase expression as a reporter gene, we performed a detailed analysis of the activity and specificity of different fragments of the SPARC promoter. A promoter sequence extending from –513 bp to +35 bp named F512 showed the best ratio of activity vs. specificity in human melanoma cells compared to non-melanoma malignant and normal cells. We constructed a conditional replicative adenoviral vector (CRAd) in which the E1A gene was driven by F512 (Ad-F512) and a second virus Ad(I)-F512-TK in which an insulator sequence was placed between the ITR and F512 and also contained the thymidine kinase gene. Both CRAds were cytotoxic on a panel of melanoma cells but were unable to replicate in normal cells from different origins including melanocytes, keratinocytes, mammary and colon cells. Ad-F512 treatment of mice carrying established melanomas inhibited tumor growth in 7 out of 9 mice compared to none in the control group treated with Ad-âgal. Ad(I)-F512-TK combined with GCV significantly enhanced the efficacy of the CRAd both in melanoma and in HMEC-1 human transformed microendothelial cells expressing SPARC. Therefore, we sought to establish whether treatment with Ad(I)-F512-TK + GCV can be effective on established tumors composed of malignant and endothelial cells in which only the latter expressed SPARC. Indeed, Ad(I)-F512-TK +GCV treatment of mixed tumors made of MiaPaca human pancreatic cancer cells and HMEC-1 inhibited tumor growth in 5 out of 6 mice. Since adenovirus type 5 efficacies for therapeutic gene transfer to human melanoma cells can be limited by the low expression of the coxsackie and adenovirus receptor, CAR, several strategies have been proposed to ablate Ad5 native tropism. Therefore, we modified the tropism of Ad-F512 and Ad(I)-F512-TK by 5/3, RGD or pK7 fiber modifications. These new CRAds were tested in a panel of cancer, stromal and normal cells. We found that capsid modified CRAds possess enhanced ability for transduction of melanoma, transformed fibroblasts and endothelial cells. Normal cells are being tested. The whole data indicate that oncolytic adenoviruses whose replication is driven by a specific SPARC promoter fragment might be considered as a potential therapeutic tool for different cancer types.