Project
1: Vector Targeting |
My laboratory primarily uses the adenovirus (Ad) vector system to transduce cancer cells with potentially therapeutic genes. The advantages of Ad are its high efficiency of transduction, ease of propagation and purification, stability, and the wide range of cell types, both proliferating and quiescent, that can be infected by the virus. The benefit of this broad range of infectivity is that multiple cancer types can be targeted by a single vector. However, additional controls are required to prevent activity of many therapeutic genes in normal tissues. We have taken two approaches to address this problem. The first is to alter the binding interaction between the virus and the host cell (called transductional targeting) and the second approach is to regulate expression of the therapeutic gene after the virus enters the host cell (called transcriptional targeting).
Experimental Approach: Transductional Targeting
In collaboration with Dr. Frank Graham at McMaster University, we have recently generated a vector that binds to ErbB3 and ErbB4 receptors that are frequently over-expressed on tumor cells. This vector modification enhances in vitro transduction of cancer cells but not normal cells. Whether this modification results in reduced toxicity to normal tissues in vivo is currently under investigation in my lab. |
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Experimental Approach: Transcriptional Targeting
Another approach for reducing toxicity to normal cells is to target expression of the therapeutic gene using tissue-specific or tumor-specific promoters. In particular, we have focused on regulatory sequences derived from the mammaglobin gene which is expressed at high levels in mammary carcinoma cells (Shi 2005, 2006). We have recently isolated the minimal promoter and upstream enhancer that appear to be responsible for the high degree of specificity and activity of this gene. We are in the process of identifying transcription factors that regulate these sequences. In addition, we are investigating the utility of this enhancer/promoter in controlling Ad vectors designed for breast cancer gene therapy.
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Project
2: Expanding the Repertoire of Genes for Cancer Gene Therapy |
Recent additions to our repertoire of potentially toxic genes are sequences that induce RNA interference (RNAi). RNAi is a naturally occurring mechanism for gene silencing mediated by short double-stranded (ds) non-coding RNA. Endogenous ds short inhibitory RNAs (siRNAs) mediate degradation of complementary messenger RNA with nearly absolute sequence specificity. The discovery that highly specific RNAi pathways can be experimentally manipulated to silence possibly any gene of known sequence has led to an explosion of studies on gene function in the past few years. In addition, the ability to silence specific genes that contribute to the maintenance or progression of disease has enormous therapeutic potential. We are developing viral vectors to transfer short hairpin RNAs that are transcribed and processed in the cell to siRNAs that are capable of triggering RNAi. We are currently investigating whether vectors that carry part or all of the microRNA gene cluster miR17-92 can be manipulated to target genes of interest.
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Project 3. Viral Oncolysis |
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Viral oncolysis is the selective
killing of tumor cells by replicating viruses. We have recently
initiated studies using the mammaglobin promoter or other regulatory
sequences to induce adenoviral early gene expression specifically in
breast cancer or other tumor cells. An alternative mechanism for
selective killing of tumor cells by viruses is to exploit cellular
processes that are altered in both infected cells and in tumor
cells, such as stimulation of DNA synthesis, activation of signaling
pathways and blockade of the interferon response. Viruses can be
designed or selected that are unable to replicate in normal cells
due to host control of these pathways, yet can replicate efficiently
in tumor cells which have lost regulation of these pathways. In
collaboration with Dr. David Evans (Medical Microbiology and
Immunology) we are investigating myxoma virus and Shope fibroma
virus for their oncolytic potential in breast cancer and other tumor
models |
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Project 4. Viral Transformation |
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Many of the cellular processes that are subverted at early stages of virus infection are also subverted at early stages of tumorigenesis. In collaboration with Dr. Andy Shaw (Experimental Oncology), we are investigating a novel cellular protein that may play an important role in intracellular trafficking of regulatory molecules during adenoviral replication and transformation.
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