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VSV-G Pseudotyped, Semi-Replication-Competent Vector for Breast Cancer Gene Therapy
Retroviral vectors have been widely used as gene delivery vehicles for cancer gene therapy. However,insufficient gene delivery with non-replicating retroviral vectors has severely limited their application for clinical protocols. A replicating vector should produce higher efficiency of gene transfer due to local spreading inside the solid tumor tissue. We recently developed novel, semi-replication-competent retroviral vector. The novelty of this vector is that: (1) We take the advantage of VSV-G (vesicular stomatitis virus G protein) to overcome interference of viral superinfection. This allows us to get multiple viral particles into the same cell to complete replication of both viruses (so called semi-replication); (2) The separation of viral elements into two vectors allows this system to have higher capacity compared to the classical single replication-competent retrovirus. (3) These semi-replication-competent vectors have anti-tumor effect in vitro and in vivo due to the high level of VSVG expression. We now hypothesize that this novel vector can be exploited to generate more potent gene therapies for breast cancer. Three specific aims are proposed : (1) We will incorporate a HSVtk mutant gene, TK30 into viral vectors. Our previous data has shown that the breast tumor cells (MCF-7 and MDA-MB231) stably transduced with non-replicating retroviral vectors carrying TK30 strongly improved GCV sensitivity (67- and 10-fold) and its bystander effect compared to cells transduced with wild-type TK. In addition, the immune stimulatory molecules will be combined with TK30 to improve therapeutic effect. (2) We will incorporate tight transcriptional regulatory elements of the HER-2/neu promoter replacing the U3 region of 3' LTR. The replication of modified vectors will therefore be restricted to breast cancer cells with overexpression of HER-2/neu gene. (3) We will use T cells, which are engineered to traffic and produce our vectors in the tumor sites by expressing chimeric receptors bearing the single chain Fv domain (scFv) derived from HER-2/neu- specific antibody linked the z-chain of TCR. The similar strategy has been successfully tested in our laboratory in a model of CEA+ liver metastatic colon cancer. Therefore, the potential of using such cell carriers to chaperone our vectors might be exploited to achieve sufficient viral delivery into tumors including metastatic breast cancer disease. Finally, this study will open new possibilities to develop retroviral vectors for breast cancer gene therapy and convert them into clinical benefit.
No satisfactory conventional treatments are available for metastatic breast cancer. Gene therapy has been recently proposed for the treatment of breast cancer. Suicide gene therapy in the context of replication-defective retroviral vectors has been extensively investigated in animals, leading to clinical trials in patients. One promising suicide gene therapy strategy is that tumor cells expressing HSVtk (herpes simplex virus thymidine kinase) can be eradicated in the presence of a nontoxic prodrug, like ganciclovir (GCV), in addition, non-transduced tumor cells can be killed due to a so-called bystander effect. However, the low transduction efficiency reported with retroviral vectors in vivo limits their application. Therefore, the development of improved retrovirus titre, transduction efficiency and safety in vivo are highly desirable. We have recently developed novel replication-competent retroviral vector, which can be produced in a 10- to 100- fold higher titer than non-replicating vector. Importantly, we have shown that the new vector is replicative and can increase transduction efficiency in solid tumors. We also observed anti-tumor effect of the vectors alone. We now hypothesize that this novel vector can be exploited to generate more effective therapies for breast cancer. It has been reported that HER-2/neu is overexpressed in 25 to 30% of all breast cancers and is associated with invasive metastasis and poor overall survival. The goal of this project will focus on developing new therapies for this group of patient. We will insert a potent TK mutant gene (TK30) in the vector, and restrict vector replication and therapeutic gene expression to HER-2/neu overexpressing tumor cells through incorporating a tumor specific promoter in the vector. We will use T cells expressing the single chain Fv domain (scFv) derived from, HER-2/neu specific antibody linked to the z-chain of TCR as virus carriers. These engineered T cells can be redirected to HER-2/neu+ tumor cells so that they can chaperone and release viruses into tumor sites. The aim of this strategy is to protect viruses from dilution and exposure to viral inactivating factors such as immune effecters, resulting in high level viral titers reaching metastatic disease. All these strategies will increase the safety of retroviral vectors for clinical trials. Finally, immune therapeutic genes will be combined in the vector in order to obtain a higher anti-tumor effect. Therefore, the work proposed here will provide a new strategy for the gene therapeutic approach to HER-2/neu+ breast cancers.