Areas of Interest
- Gene therapy
- Vectored immunoprophylaxis (VIP)
- Oncolytic virotherapy
- Viral vectored vaccines
- Viral pathogenesis
Gene therapy for monogenetic lung diseases including Cystic Fibrosis and Alpha-1 Antitrypsin Deficiency
Cystic fibrosis (CF) is caused by inherited mutations in the gene encoding the cystic fibrosis transmembrane conductance regulator (CFTR) and is one of the most widespread inherited genetic disorders. Although the disease affects multiple organs, the primary cause of morbidity and mortality is cystic fibrosis related lung disease. Due to its recessive nature, i.e. requirement for two defective copies of the gene, and easy accessibility of the lungs to inhaled gene therapy agents; CF is an excellent candidate for gene replacement therapy. With funding from CF Canada, our lab has been developing gene therapy vectors to “functionally cure” individuals with CF. To achieve this goal, we have genetically engineered baculovirus, a harmless insect virus, to permanently insert genes of interest into a safe location within the human genome using two different cutting-edge genome-editing technologies (TALENs and CRISPR-Cas9). We have also modified the baculovirus genome so that it can “hide” from the immune system long enough to permit site-specific integration of therapeutic genes to occur. By engineering baculovirus to deliver all the “tools” required to insert a functional copy of the CFTR gene into the genome of lung cells, we hope to be able to provide a safe, economical and effective treatment for CF patients irrespective of their CFTR mutation status, gender or genetic background. Additionally, these studies would validate this platform as a general gene therapy vector for broader gene therapy applications in tissues other than the lung, as well as ex vivo correction of human stem cells.
Alpha-1-antitrypsin deficiency (AATD) is a monogenetic disorder commonly associated with adult onset lung diseases, including emphysema, chronic obstructive pulmonary disease (COPD) and airway inflammation and affects an estimated 190 million people worldwide. A promising treatment approach for AATD is gene therapy. The idea behind gene therapy is to provide an individual who has defective copies of the AAT gene, with a functional, good copy of the gene. Recently great strides have been made in gene therapy, however none of the current approaches can permanently deliver a good copy of the AAT gene into the human genome in a controlled manner. Our lab is currently employing the use of adeno-associated virus (AAV) and baculovirus gene therapy vectors and two different cutting-edge genome editing technologies (TALENs and CRISPR-Cas9) to insert a functional copy of the AAT gene, permanently, into a safe location within the human genome.
- Dr. Theo Moraes, Sick Kids
- Cystic Fibrosis Canada
- Ontario Lung Association
Antibody-based protection against Ebola virus infection by vectored immunoprophylaxis
Pathogens for which the human population has no pre-existing or vaccine-induced immunity and which pose a significant threat to public health, require rapid and cost effective countermeasures to block transmission and mitigate disease. One such method that has proven successful is the administration of neutralizing antibodies to patients as a means of pre- and post-exposure prophylaxis. While effective, and often the only treatment option available, production of clinical grade MAbs is a labor-intensive process with considerable technical challenges, and immunity does not last long. One way to circumvent the obstacles associated with passive immunization is to use the host to “manufacture” neutralizing MAbs in vivo, by employing a method called vectored immunoprophylaxis (VIP). VIP produces full-length antibodies that are identical in sequence to those produced by the immune system. VIP involves a single intramuscular injection of an adeno-associated virus (AAV) vector expressing genes encoding pathogen‑specific broadly neutralizing MAbs leading to continuous and sustained secretion of antibodies into the circulation and protection against infection. We are currently developing a vectored immunoprophylaxis approach for the protection against Ebola virus infection in humans. The long-term objective of this project is to employ the use of AAV vectors to promote prolonged in vivo expression of protective neutralizing antibodies (ZMab) against Ebola virus at therapeutic levels from a single intramuscular injection with the ultimate goal of preventing Ebola Virus Disease (EVD) in humans.
- Dr. Gary Kobinger, Laval University
- Dr. Xiangguo Qiu, Public Health Agency of Canada
Oncolytic virotherapy using Newcastle disease virus and parapox ORF virus
Newcastle disease virus (NDV) is a member of the Avulavirus genus in the Paramyxoviridae family. NDV selectively replicates in tumor cells due to defects in antiviral and apoptotic signalling. NDV has the longest history of use in clinical trials of all oncolytic viruses and has a proven safety record as a monotherapy due to its strong induction of antiviral responses in normal mammalian cells. In addition to its direct oncolytic effect, NDV also activates both innate and adaptive immune responses and therefore has strong immunotherapeutic potential. Parapoxivirus ovis (ORFV) is a poxvirus and veterinary pathogen causing transmissible pustular dermatitis in sheep. ORFV is able to infect multiple cancer cell types and has demonstrated impressive efficacy in pre-clinical models of cancer, largely due to its remarkable immunogenicity. Research in our lab focuses on the development of recombinant NDV and ORFV viruses with improved immunostimulatory and oncolytic properties for the treatment of ovarian and prostate cancer.
- Dr. Jim Petrik, University of Guelph
- Dr. Byram Bridle, University of Guelph
Development of a recombinant parapoxvirus vaccine to protect against Toxoplasma gondii infection in sheep
Live virus vaccines are excellent inducers of long-term immunity by eliciting protective humoral and cell-mediated immune responses against foreign antigens. To this end, recombinant poxviruses are one of the most versatile expression systems for foreign antigens and have been extensively developed as vaccine vectors for veterinary diseases. Recently, the type species ORFV (ORFV) of the genus Parapoxvirus in the family Poxviridae has been investigated as a novel vaccine vector. Attributes that favor the use of ORFV as a vaccine vector include: limited host range (sheep and goats), restricted tropism to the skin, lack of systemic infection, short-term vector-specific protective immunity, and unique immune-modulating properties which strongly stimulate the innate immune response at the site of infection, induce a potent Th1 immune response and rapidly generate foreign antigen-specific immune responses. Due to the large genome size of ORFV, it is possible to insert multiple antigens thereby producing multivalent vaccines.
Toxoplasmosis is a disease caused by the parasite Toxoplasma gondii (T. gondii) and is one of the most common parasitic diseases of warm-blooded animals. In humans, T.gondii infection can cause severe disease in immunocompromised individuals and pregnant women and latent Toxoplasma infections have been associated with behavioural changes and schizophrenia. T. gondii is also the most common cause of infectious abortion in sheep and goats in Ontario. Studies have indicated that there is widespread environmental contamination with T. gondii oocysts and producers in Canada have no means of controlling infection. The objective of this research project is to develop a recombinant parapoxvirus vaccine expressing three different T. gondii protective antigens (SAG1-ROP2-GRA2) and to evaluate the immunogenicity and protective efficacy of the rORFV/SAG1-ROP2-GRA2 vaccine in mice and sheep. We anticipate that sheep immunized with rORFV-SAG1-ROP2-GRA2 will develop a robust immune response against T. gondii that will lead to a reduction in tissue cyst formation and ultimately, protection against congenital infection.
- Dr. Karen Shapiro, University of California Davis, University of Guelph
- Dr. Paula Menzies, Ontario Veterinary College
- Dr. Byram Bridle, University of Guelph
Pathogenesis of ovine betaretroviruses
The study of oncogenic retroviruses has been instrumental in elucidating the functions of key cellular proteins involved in cell growth and transformation. Many of the oncogenes found to be active in human cancers were first discovered in avian, murine and feline retroviruses. Moreover, the discovery of the activation of cellular genes upon retroviral insertion into chromosomal DNA has provided a model of oncogenesis similar in mechanism to chromosome translocation and/or gene rearrangement in human tumors. Novel mechanisms of transformation by retroviruses continue to be identified, as evidenced by our discovery of the oncogenic properties of the ovine betaretroviral envelope (Env) proteins, a heretofore-unprecedented finding. Betaretroviruses, of which Jaagsiekte sheep retrovirus (JSRV), Enzootic nasal tumor virus (ENTV), mouse mammary tumor virus (MMTV) and Mason-Pfizer monkey virus (MPMV) are members, are unique in that unlike many of the oncogenic retroviruses, which cause hematopoietic malignancies, viruses in this family are often associated with the development of solid tumors, primarily carcinomas. Research in our laboratory is largely directed towards understanding the molecular pathogenesis of JSRV and ENTV, causative agents of transmissible lung and nasal cancer, respectively, in sheep and goats. As viral proteins subvert many of the same cellular pathways and checkpoints commonly mutated in tumor cells, these model systems are being used to uncover critical events in the development of lung and nasal cancer.
Lisa Santry (PhD program)
“Improving the oncolytic and immunomodulatory properties of recombinant Newcastle Disease Virus”
Van Vloten, Jacob (PhD program; co-advised)
“Role of immunogenic cell death in ORF virus-induced oncolysis and immune stimulation”
Maria Rosales Gerpe, MSc (PhD program)
“Tissue Tropism and Transformation Determinants of Jaagsiekte Sheep Retrovirus and Enzootic Nasal Tumour Virus”
Joelle Ingrao, DVM, DVSc (PhD program)
“Development of a recombinant parapoxvirus vaccine against Toxoplasma gondii”
Laura van Lieshout (PhD program)
“Development of a novel hybrid gene therapy vector using CRISPR-mediated gene targeting for the correction of alpha-1 antitrypsin deficiency”
Research and Professional Staff
Jondavid de Jong, PhD (Postdoctoral fellow)
“Nuclease-based gene therapy for permanent correction of CF lung disease”
Betty-Anne McBey, MSc
Flow cytometry, propagation of viruses, molecular biology
- MICR*4430, Medical Virology, Coordinator
- PABI*6330, Viral Diseases, Coordinator
- PABI*6960, Special Topics in Virology, Coordinator
- BIOM*4150/6702, Cancer Biology (Oncogenic Virus unit)
- PABI*6104, Mechanisms of Disease (Lecture: “Viral Pathogenesis”)
Honours and Professional Experiences
B.Sc. Biochemistry 1997 University of Guelph
Ph.D. Virology 2002 Ontario Veterinary College, University of Guelph
Postdoctoral Training 2007 Fred Hutchinson Cancer Research Center, Seattle WA
Search PubMed for additional publications by Dr. Wootton .