629. Preclinical Evaluation of Prodrug Activator Gene Therapy by Retroviral Replicating Vectors (RRV) in Experimental Models of Breast Cancer CNS Metastasis

Abstract

Retroviral replicating vectors (RRV) are capable of highly efficient replication in cancer cells in vitro and in vivo. In a variety of cancer models this is associated with high levels of tumor-selective gene transfer and significantly enhanced survival benefit when employed for prodrug activator gene therapy. An RRV encoding a modified yeast cytosine deaminase (CD) prodrug activator gene (Toca 511) in combination with extended-release flucytosine (Toca FC) is under investigation in multi-center ascending-dose trials in patients with recurrent high grade glioma. In the present study, our goal is to establish the feasibility of also applying this novel gene therapy strategy to secondary brain tumors arising from metastasis of systemic cancers. In particular, brain metastases of breast cancer frequently arise from highly aggressive, treatment-refractory, “triple-negative” (ER(-), PR(-), HER2(-)) cells, and are associated with a dismal prognosis of 4-6 months survival. First, RRV encoding GFP (AC3-emd, RRV-GFP) showed robust replication activity in the human breast cancer tumor cell lines MDA-MB-231-BR and JC over time at both MOI = 0.01 and 0.1, resulting in high levels of transduction within 1-2 weeks. Next, we tested in vitro cytotoxicity after 5-FC treatment of MDA-MB-231BR and JC cells infected with an RRV expressing CD (AC3-yCD2, Toca 511). In both of the AC3-yCD2-transduced cell lines, cell viability was reduced approximately 70-85% after exposure to 12.9 mg/mL 5-FC and complete cell killing was observed with 129 mg/mL 5-FC (6 days 5-FC treatment for 231-BR; 4 days for JC). In a 231-BR intracranial xenograft model, intratumoral injection of 10^6 Transducing Units of RRV-GFP resulted in >80% transduction efficiency on Day 10 post-virus injection. In survival studies, animals treated with Toca 511 and 5-FC showed statistically significant (p<0.0001) survival benefit. These preclinical data support the advancement of RRV gene therapy towards clinical investigation in patients with CNS-metastatic breast cancer.