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1
FROECHLICH, GUENDALINA. "DISSECTING THE STING-DEPENDENT MOLECULAR MECHANISMS IN A PRECLINICAL MODEL OF COMBINED TREATMENT WITH TUMOUR-TARGETED HERPES SIMPLEX VIRUS AND IMMUNE CHECKPOINT BLOCKADE." Doctoral thesis, Università degli Studi di Milano, 2021. http://hdl.handle.net/2434/883382.
Full textAbstract:
Oncolytic viruses promote anti-tumour immune response by direct tumour cell killing and
activation of intratumoural immune system. The role of innate antiviral immune response to
oncolytic viruses is still debated, as they counteract viral replication and trigger adaptive antitumor
immunity. The DNA sensing-mediated cGAS/STING axis may act as a key balancer between lytic and
immunotherapeutic activity of oncolytic viruses. Indeed, upon infection, viral DNA is sensed by
cGAS/STING axis that, in turn, induces type-I interferon cascade counteracting viral replication and
spread. For this reason, STING represents a hurdle for classical lytic-centric function of oncolytic
viruses. On the other side, the immunological role of STING should also be considered, as it is
emerging as a key bridge between innate and adaptive immunity. To evaluate the role of STING
expression in tumour cells in response to onco-virotherapy, we generated murine STING KO tumour
cell lines through CRISPR/Cas9 genome editing. Preclinical studies in syngeneic immunocompetent
tumour-bearing mice showed that the inactivation of STING in tumour cells, while favouring
oncolytic viral replication, impaired the immunotherapeutic effects of combination therapy based
on herpetic oncolytic virus and PD1 blockade. Molecular characterization of tumours revealed that
loss of STING prevents antitumour immune activation inducing a tolerogenic cell death and
immunosuppressive tumour microenvironment. Accordingly, I propose that antiviral, tumourresident
STING provides fundamental contributions to heat-up the TME eliciting
immunotherapeutic efficacy of oncolytic viruses.
2
Komar, Monica. "Potentiating the Oncolytic Efficacy of Poxviruses." Thèse, Université d'Ottawa / University of Ottawa, 2012. http://hdl.handle.net/10393/23114.
Full textAbstract:
Several wild-type poxviruses have emerged as potential oncolytic viruses (OVs), including orf virus (OrfV), and vaccinia virus (VV). Oncolytic VVs have been modified to include attenuating mutations that enhance their tumour selective nature, but these mutations also reduce overall viral fitness in cancer cells. Previous studies have shown that a VV (Western Reserve) with its E3L gene replaced with the E3L homologue from, OrfV (designated VV-E3LOrfV), maintained its ability to infect cells in vitro, but was attenuated compared to its parental VV in vivo. Our goal was to determine the safety and oncolytic potential VV-E3LOrfV, compared to wild type VV and other attenuated recombinants. VV-E3LOrfV, was unable to replicate to the same titers and was sensitive to IFN compared to its parental virus and other attenuated VVs in normal human fibroblast cells. The virus was also less pathogenic when administered in vivo. Viral replication, spread and cell killing, as measures of oncolytic potential in vitro, along with in vivo efficacy, were also observed..
The Parapoxvirus, OrfV has been shown to have a unique immune-stimulation profile, inducing a number of pro-inflammatory cytokines, as well as potently recruiting and activating a number of immune cells. Despite this unique profile, OrfV is limited in its ability to replicate and spread in human cancer cells. Various strategies were employed to enhance the oncolytic efficacy of wild-type OrfV. A transient transfection/infection screen was created to determine if any of the VV host-range genes (C7L, K1L, E3L or K3L) would augment OrfV oncolysis. Combination therapy, including the use of microtubule targeting agents, Viral Sensitizer (VSe) compounds and the addition of soluble VV B18R gene product were employed to see if they also enhance OrfV efficacy. Unfortunately, none of the strategies mentioned were able to enhance OrfV.
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Heiber, Joshua F. "Characterization and Development of Vesicular Stomatitis Virus For Use as an Oncolytic Vector." Scholarly Repository, 2011. http://scholarlyrepository.miami.edu/oa_dissertations/600.
Full textAbstract:
Oncolytic virotherapy is emerging as a new treatment option for cancer patients. At present, there are relatively few oncolytic virus clinical trials that are underway or have been conducted, however one virus that shows promise in pre-clinical models is Vesicular Stomatitis Virus (VSV). VSV is a naturally occurring oncolytic rhabdovirus that has the ability to preferentially replicate in and kill malignant versus normal cells. VSV also has a low seroprevalence, minimal associated morbidity and mortality in humans, and simple non-integrating genome that can be genetically manipulated, making it an optimal oncolytic vector. Currently, many labs are using a variety of different strategies including inserting trans genes that can modulate the innate and adaptive immune response. VSV can also be retargeted by altering its surface glycoprotein (G) or be made replication incompetent by deleting the G protein. Currently, our lab has engineered a series of new recombinant VSVs, incorporating either the murine p53 (mp53), IPS-1, or TRIF transgene. mp53, IPS-1 and TRIF were incorporated into the normal VSV-XN2 genome and mp53 was also incorporated into the mutated VSV-ΔM vector generating VSV-mp53, VSV-IPS-1, VSV-TRIF and VSV-ΔM-mp53. Our data using these new viruses indicate that these viruses preferentially replicate in and kill transformed versus non-transformed cells and efficiently express the transgene. However, despite the ability for VSV-IPS-1 and VSV-TRIF to induce a robust type 1 IFN response, VSV-ΔM-mp53 was the only construct that had reduced toxicity and elicited an increased anti-tumor response against a syngeneic metastatic mammary tumor model. VSV- ΔM-mp53 treatment lead to a reduction in IL-6 and IP-10 production, an increase in tumor specific CD8+ T cells, and immunologic memory against the tumor. Collectively these studies highlight the necessity for additional VSV construct development and the generation of new clinically relevant treatment schema.
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Jamieson-Datzkiw, Taylor Rae. "A Tailored Viro-Immunotherapy Combination Approach for the Treatment of BRCA1/2 Mutated Breast and Ovarian Cancers." Thesis, Université d'Ottawa / University of Ottawa, 2021. http://hdl.handle.net/10393/42736.
Full textAbstract:
Hereditary breast and ovarian cancers (HBOC) represent 5-10% of breast and 10-15% of ovarian cancer cases. These cancers tend to be aggressive and curative treatment strategies are scarce. Poly(ADP-ribose) polymerase inhibitors (PARPi), a family of drugs that inhibit DNA repair, are a promising therapy for cancers harbouring mutations in their DNA repair machinery, such as HBOC. Unfortunately, nearly all patients ultimately become resistant to PARPi, leaving limited options for definitive treatment. Oncolytic or “cancer-killing” viruses are an innovative immunotherapeutic platform capable of selectively targeting cancer cells, leaving healthy tissues unharmed. Our group has demonstrated that oncolytic rhabdoviruses may be used to deliver therapeutic payloads by encoding targeting sequences to act on genes via RNA interference. In the present work, I have engineered the oncolytic virus, vesicular stomatitis virus (VSV), to express a variety of microRNA (miRNA) sequences that target genes essential for DNA repair, sensitizing resistant cancer cells to PARPi therapy. After initial experiments revealed hurdles concerning the functionality of artificial miRNAs which specifically target BRCA1 and BRCA2 I encoded the naturally occurring hsa-miR-182 into VSV to knockdown BRCA1 and additional genes essential for DNA repair. Using a 3D spheroid model, I have demonstrated sensitization of initially resistant MDA-MB-231 breast cancer cells to the PARPi, rucaparib. Complementary work exploring the shuttling of miRNAs into small extracellular vesicles, or EVs, has also shown that we can take advantage of the EV packaging facilities in infected cells, inducing the packaging of miRNAs over-expressed by VSV (EV-miRNAs) into EVs. Future work will address the functionality of these EV-miRNAs, testing their ability to knockdown targets in uninfected cancer cells.
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Selman, Mohammed. "Pharmacological Improvement of Oncolytic Virotherapy." Thesis, Université d'Ottawa / University of Ottawa, 2018. http://hdl.handle.net/10393/37626.
Full textAbstract:
Oncolytic viruses (OV) are an emerging class of anticancer bio-therapeutics that induce antitumor immunity through selective replication in cancer cells. However, the efficacy of OVs as single agents remains limited. We postulate that resistance to oncolytic virotherapy results in part from the failure of tumor cells to be sufficiently infected. In this study, we provide evidence that in the context of sarcoma, a highly heterogeneous malignancy, the infection of tumors by different oncolytic viruses varies greatly. Similarly, for a given oncolytic virus, productive infection of tumors across patient samples varies by many orders of magnitude. To overcome this issue, we hypothesize that the infection of resistant tumors can be achieved through the use of selected small molecules. Here, we have identified two novel drug classes with the ability to improve the efficacy of OV therapy: fumaric and maleic acid esters (FMAEs) and vanadium compounds. FMAEs are enhancing infection of cancer cells by several oncolytic viruses in cancer cell lines and human tumor biopsies. The ability of FMAEs to enhance viral spread is due to their ability to inhibit type I IFN production and response, which is associated with their ability to block nuclear translocation of transcription factor NF-κB. Vanadium-based phosphatase inhibitors enhance OV infection of RNA viruses in vitro and ex vivo, in resistant cancer cell lines. Mechanistically, this involves subverting the antiviral type I IFN response towards a death-inducing and proinflammatory type II IFN response, leading to improved OV spread, increased bystander killing of cancer cells, and enhanced anti-tumor immune-stimulation. Both FMAEs and vanadium compounds improve therapeutic outcomes of OV treatment in syngeneic tumor models, leading to durable responses, even in models otherwise refractory to OV and drug alone. Overall, we showcased novel avenues for the development of improved immunotherapy strategies.
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Muharemagic, Darija. "Aptamers as Enhancers of Oncolytic Virus Therapy." Thesis, Université d'Ottawa / University of Ottawa, 2015. http://hdl.handle.net/10393/32170.
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Oncolytic viruses promise to significantly improve current cancer treatments through their tumour-selective replication and multimodal attack against cancer cells. However, one of the biggest setbacks for oncolytic virus therapies is the intravenous delivery of the virus, as it can be cleared by neutralizing antibodies (nAbs) from the bloodstream before it reaches the tumour cells. In our group, we have succeeded in developing aptamers to vesicular stomatitis virus (VSV), as well as to rabbit anti-VSV polyclonal neutralizing antibodies (nAbs). We tested these aptamers’ biological activity with a cell-based plaque forming assay and found that the aptamers prevented in vitro neutralization of VSV by nAbs and increased the virus infection rate of transformed cells up to 77%.
In line with this approach, we enhanced the delivery of oncolytic viruses by selecting aptamers to the CT26 colon carcinoma cell line. The binding of aptamer pools has been tested on flow cytometry and the best pools were subjected to high throughput sequencing. Selected aptamers were linked to anti-VSV aptamers and applied for target delivery of the virus to cancer cells. Development of this aptamer-based technology aims to improve viral anti-cancer therapies, with a potential to be applied as treatment for patients affected with cancer.
Finally, in collaboration with a group from Erlangen University, we performed an aptamer selection using capillary electrophoresis and cell-SELEX. The target, the extracellular domain of human CD83, is a maturation marker for dendritic cells and is involved in the regulation of the immune system. Selected aptamer sequences bound selectively to mature dendritic cells, in comparison to immature dendritic cells, and thus hold promise to be applied for further studies leading to a better understanding of CD83’s mechanism of action.
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Lemay, Chantal. "Harnessing Oncolytic Virus-mediated Anti-tumour Immunity." Thèse, Université d'Ottawa / University of Ottawa, 2012. http://hdl.handle.net/10393/23318.
Full textAbstract:
Treatment of permissive tumours with the oncolytic virus (OV) VSV-Δ51 leads to a robust anti-tumour T cell response, which contributes to efficacy; however, many tumours are not permissive to in vivo treatment with VSV-Δ51. In an attempt to channel the immune stimulatory properties of VSV-Δ51 and broaden the scope of tumours that can be treated by an OV, a potent oncolytic vaccine platform was developed, consisting of tumour cells infected with VSV-Δ51. I demonstrate that prophylactic immunization with this infected cell vaccine (ICV) protected mice from subsequent tumour challenge, and expression of GM-CSF by the virus (VSVgm-ICV) increased efficacy. Immunization with VSVgm-ICV in the VSV-resistant B16-F10 model induced maturation of dendritic cells, natural killer (NK) cells, and T cells. I demonstrate that this approach is robust enough to control the growth of established and spontaneous tumours. This strategy is broadly applicable because of VSV’s extremely broad tropism, allowing nearly all cell types to be infected at high MOIs in vitro, where the virus replication kinetics outpace the cellular IFN response. It is also personalized to the unique tumour antigen(s) displayed by the cancer cell. Histone deacetylase inhibitors (HDIs) can augment viral replication, making them particularly interesting complements to OV therapy. However, the impact of HDIs on the generation and re-stimulation of immune responses remains to be clearly elucidated. Along with my collaborators at McMaster University, I demonstrate that MS-275, but not SAHA, selectively depletes naïve and regulatory lymphocytes. Memory lymphocytes that are being boosted remain unscathed and even have enhanced cytokine production, potentially as a consequence of the depleted lymphocyte compartment. This leads to a delay in anti-VSV neutralizing antibodies and T cell responses. Interestingly, HDI treatment of B16-F10 cells appears to inhibit VSV replication but allows for a longer persistence within the tumour. When used in an oncolytic prime/boost vaccination model, MS-275 potently enhanced survival. Though the anti-tumour immune response is enhanced, a near complete reduction in autoimmune vitiligo is observed with MS-275 administration. Therefore, this HDI uniquely modulates the immune response to enhance anti-tumour immunity and decrease the anti-viral response, while also decreasing autoimmune sequelae.
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Zeicher, Marc. "Oncolytic viruses cancer therapy." Doctoral thesis, Universite Libre de Bruxelles, 2008. http://hdl.handle.net/2013/ULB-DIPOT:oai:dipot.ulb.ac.be:2013/210439.
Full textAbstract:
Wild-type viruses with intrinsic oncolytic capacity in human includes DNA viruses like some autonomous parvoviruses and many RNA viruses. Recent advances in molecular biology have allowed the design of several genetically modified viruses, such as adenovirus and herpes simplex virus that specifically replicate in, and kill tumor cells. However, still several hurdles regarding clinical limitations and safety issues should be overcome before this mode of therapy can become of clinical relevance. It includes limited virus spread in tumor masses, stability of virus in the blood, trapping within the liver sinusoids, transendothelial transfer, and/or vector diffusion of viral particles to tumor cells, limited tumor transduction, immune-mediated inactivation or destruction of the virus. For replication-competent vectors without approved antiviral agents, suicide genes might be used as fail-safe mechanism. Cancer stem cells are a minor population of tumor cells that possess the stem cell property of self-renewal. Therefore, viruses that target the defective self-renewal pathways in cancer cells might lead to improved outcomes.In this thesis, data we generated in the field of oncolytic autonomous parvoviruses are presented.
We replaced capsid genes by reporter genes and assessed expression in different types of human cancer cells and their normal counterparts, either at the level of whole cell population, (CAT ELISA) or at the single cell level, (FACS analysis of Green Fluorescent Protein). Cat expression was substantial (up to 10000 times background) in all infected tumor cells, despite variations according to the cell types. In contrast, no gene expression was detected in similarly infected normal cells, (with the exception of an expression slightly above background in fibroblasts.). FACS analysis of GFP expression revealed that most tumor cells expressed high level of GFP while no GFP positive normal cells could be detected with the exception of very few (less than 0.1%) human fibroblast cells expressing high level of GFP. We also replace capsid genes by genes coding for the costimulatory molecules B7-1 and B7-2 and show that, upon infection with B7 recombinant virions, only tumor cells display the costimulatory molecules and their immunogenicity was increased without any effect on normal cells. Using a recombinant MVM containig the Herpes Simplex thymidine kinase gene, we could get efficient killing of most tumor cell types in the presence of ganciclovir, whithout affecting normal proliferating cells. We also produced tetracycline inducible packaging cell lines in order to improve recombinant vectors yields. The prospects and limitations of these different strategies will be discussed.
An overview is given of the general mechanisms and genetic modifications by which oncolytic viruses achieve tumor cell-specific replication and antitumor efficacy. However, as their therapeutic efficacy in clinical trials is still not optimal, strategies are evaluated that could further enhance the oncolytic potential of conditionally replicating viruses in conjunction with other standard therapies.
Another exciting new area of research has been the harnessing of naturally tumor-homing cells as carrier cells to deliver oncolytic viruses to tumors. The trafficking of these tumor-homing cells (stem cells, immune cells and cancer cells), which support proliferation of the viruses, is mediated by specific chemokines and cell adhesion molecules and we are just beginning to understand the roles of these molecules. Finally, we will explore some ways deserving further study in order to be able to utilize various oncolytic viruses for effective cancer treatment.
Doctorat en sciences, Spécialisation biologie moléculaire
info:eu-repo/semantics/nonPublished
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Evgin, Laura. "Enhancing the Delivery of Oncolytic Vaccinia Virus to the Tumors of Hosts with Pre-Existing Immunity." Thesis, Université d'Ottawa / University of Ottawa, 2015. http://hdl.handle.net/10393/32423.
Full textAbstract:
Oncolytic viruses (OVs) have begun to show their promise in the clinical setting, however these results have been predominantly associated with loco-regional administration of virus. The treatment of metastatic disease necessitates a systemic approach to virus delivery. The circulatory system, though, is a hostile environment for viruses and the advantages associated with intravenous (IV) delivery come at a heavy cost that must be understood and brokered. Pre-existing immunity, specifically through the function of antibody and complement, poses a significant hurdle to the IV delivery of infectious virus to dispersed tumor beds. This is of particular importance for therapeutic vaccinia viruses as a majority of today’s cancer patients were vaccinated during the smallpox eradication campaign. In vitro neutralization assays of oncolytic vaccinia virus demonstrated that the antibodies elicited from smallpox vaccination, and also the anamnestic response in patients undergoing Pexa-Vec treatment, was minimally neutralizing in the absence of functional complement. Accordingly, in a Fischer rat model, complement depletion stabilized virus in the blood of pre-immunized hosts and correlated with improved delivery to mammary adenocarcinoma tumors. Complement depletion additionally enhanced infection of tumors following direct intratumoral injection of virus. The feasibility and safety of using a complement inhibitor, CP40, was tested in a cynomolgus macaque model. Immune animals saw an average 10-fold increase in infectious virus titer at an early point after the infusion, and a prolongation of the time during which infectious virus was still detectable in the blood. We have also demonstrated that vaccinia virus engages in promiscuous interactions with cells in the blood and that these interactions may be partially complement-dependent. Additionally, we have translated this complement inhibition approach to other OV candidates and found that reovirus, measles virus and a virus pseudo typed with the LCMV glycoprotein all elicit antibodies, that to some degree, are dependent on complement activation to neutralize their target viruses. We show here that capitalizing on the complement dependence of anti-viral antibody with adjunct complement inhibitors may increase the effective dose to enable successful delivery of multiple rounds of OV in immune hosts.
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Pelin, Adrian. "Bio-Engineering Vaccinia Viruses for Increased Oncolytic Potential." Thesis, Université d'Ottawa / University of Ottawa, 2019. http://hdl.handle.net/10393/39909.
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Vaccinia virus has a large and still incompletely understood genome although several strains of this virus are already in clinical development. For the most part, clinical candidates have been attenuated from their wild type vaccine strains through deletion of metabolic genes like the viral thymidine kinase gene. In the present work, we thoroughly examined the genetic elements of vaccinia which could be modulated to improve tailor the virus as a cancer therapeutic. Using a variety of cancer cell lines and primary tumor explants, we performed a fitness assay that directly compares multiple wild-type Vaccinia strains to identify the genetic elements that together create an optimal “oncolytic engine”. Using a transposon insertion strategy and deep sequencing of viral populations we systematically examined Vaccinia genes that do or do not play a role in the therapeutic activity of the virus. Our studies allowed us to identify a variety of genes in the vaccinia genome that when deleted, augment the oncolytic activity of a newly engineered Vaccinia virus. In the context of this thesis, I define enhanced oncolytic activity as superior therapeutic activity, increased immunogenicity and an improved safety profile, all aspects which we used to compare this novel virus to Vaccinia viruses currently in the clinic.
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Hassanzadeh, Golnoush. "Characterizing Cellular Responses During Oncolytic Maraba Virus Infection." Thesis, Université d'Ottawa / University of Ottawa, 2017. http://hdl.handle.net/10393/35871.
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The rising demand for powerful oncolytic virotherapy agents has led to the identification of Maraba virus, one of the most potent oncolytic viruses from Rhabdoviridae family which displays high selectivity for killing malignant cells and low cytotoxicity in normal cells. Although the virus is readied to be used for clinical trials, the interactions between the virus and the host cells is still unclear. Using a newly developed interferon-sensitive mutant Maraba virus (MG1), we have identified two key regulators of global translation (4E-BP1 and eIF2α) responsible for the inhibition of protein synthesis in the infected cells. Despite the translational arrest upon viral stress, we showed an up-regulation of anti-apoptotic Bcl-xL protein that provides a survival benefit for the host cell, yet facilitates effective viral propagation. Given the fact that eIF5B canonically regulates 60S ribosome subunit end joining, and is able to replace the role of eIF2 in delivering initiator tRNA to the 40S ribosome subunit upon the phosphorylation of eIF2α, we have tested whether eIF5B mediates the translation of target mRNAs during MG1 infection. Our results show that the inhibition of eIF5B significantly down-regulates the level of Bcl-xL steady-state mRNA, thus indirectly attenuates viral propagation.
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Hiley, Crispin. "Arming vaccinia virus for pancreatic cancer oncolytic virotherapy." Thesis, Queen Mary, University of London, 2011. http://qmro.qmul.ac.uk/xmlui/handle/123456789/2344.
Full textAbstract:
Vaccinia virus is a 250-300nm enveloped DNA virus from the poxvirus family and is used as a vector for oncolytic viral gene therapy. No unique cell surface receptor has been identified for Vaccinia virus and the reasons for its tropism for cancer cells are unclear. Pancreatic adenocarcinoma (PDAC) is resistant to conventional chemotherapy and typically contains areas that are profoundly hypoxic. We have investigated the utility of Vaccinia virus as a vector for targeting hypoxic regions in pancreatic adenocarcinoma, as other viral vectors have been found to replicate poorly in hypoxia. We found that cytotoxicity was equivalent in normoxia and hypoxia in some PDAC cell lines but in others cytotoxicity was enhanced in hypoxia. This increase in cytotoxicity was only seen in cell lines where there was hypoxic induction of vascular endothelial growth factor (VEGF). Functional studies using over-expression and knockdown of VEGF in pancreatic cancer cell models showed that VEGF can augment viral transgene expression, cytotoxicity and replication in vitro and in vivo. We found that VEGF facilitates the internalisation of Vaccinia virus. These results show that VEGF is an additional factor involved in the tropism and pathogenesis of Vaccinia virus. We then constructed an oncolytic Vaccinia virus to target hypoxic cancer cells using the HIF-1α oxygen degradation domain, encephalomyocarditis virus internal ribosomal entry site and the VEGF 3‟ un-translated region to regulate luciferase expression in hypoxia. We have shown a dose-, time- and oxygen-dependent effect using this construct and propose this may be adapted to regulate therapeutic genes, or produce a conditionally replicating Vaccinia virus, in hypoxic conditions.
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Kueberuwa, Gray L. B. "Development of sindbis virus as an oncolytic agent." Thesis, University of Oxford, 2012. http://ora.ox.ac.uk/objects/uuid:d9737357-ab86-4baf-92a9-b4a21b615fd7.
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The poor stability of most therapeutic viruses in the human bloodstream is a major obstacle in the field of cancer virotherapy, preventing systemic intravenous delivery to treat tumour metastases. Delivery is typically limited by inactivation of virus particles by blood components and rapid scavenging by hepatic phagocytes. Members of the Alphavirus family are exposed to blood during natural infections; as such, we hypothesised that evolutionary pressure may have led to blood stability and clearance kinetics superior to those of other viruses currently in development for use as oncolytic agents. Sindbis virus is a member of the Alphavirus family that has shown promising anti-cancer activity in pre-clinical models. A concern for the clinical use of Sindbis virus as an anticancer agent is its pathology in humans, known as Pogosta disease. The symptoms of Pogosta disease may be a result of Sindbis virus replication in neuronal, muscle or haematopoietic tissues. Inhibiting virus replication in these tissues could, therefore, alleviate such potential side effects of virotherapy treatment. Introduction of microRNA response elements, perfectly complementary to microRNAs specifically expressed in liver (miR122), neuronal (miR124), muscle (miR133 and miR206) and haematopoietic (miR142-3p) cells, successfully attenuated SV replication in these tissues. In contrast to all other viruses studied, data presented in this thesis show that Sindbis virus infectivity in vitro is not significantly inhibited by incubation with neat, whole naïve human blood. Despite full infectivity in naïve mouse blood, virus particles were rapidly cleared from the circulation of mice in vivo by the liver. An attempt to decrease the clearance rate by depletion of Kupffer cells through pre-treatment of mice with clodronate liposomes was ineffective. We also explored the use of Sindbis virus packaged in mosquito cells to more closely mimic virus particles exposed to blood in the wild during mosquito mediated transmission, but this also failed to improve virus circulation kinetics in mice. Despite rapid clearance from the circulation, intravenous administration of Sindbis virus had significant anti-cancer efficacy in C57BL/6 mice bearing syngeneic B16F10 metastatic melanomas. Overall, data presented support our proposed use of Sindbis virus as a systemically delivered oncolytic agent and suggest decreasing the rate of clearance by the liver could dramatically enhance therapeutic outcomes. In addition it is shown that microRNA targeting of Sindbis virus provides a means of alleviating potential side effects of the administration of large virus doses.
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Laporte, Aimée N. "Enhancing the Oncolytic Efficacy of Vaccinia Virus by Mutagenic Augmentation of EEV Production." Thèse, Université d'Ottawa / University of Ottawa, 2012. http://hdl.handle.net/10393/23348.
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Oncolytic viruses are currently under investigation as anti - cancer therapies due to their innate ability to selectively infect and destroy cancer cells. Major barriers to this anti - tumour effect include inefficient viral spread and immune - mediated neutralization. This study aims to overcome these limitations by taking advantage of the life cycle of the oncolytic clinical candidate known as vaccinia virus (VACV). Naturally, a small proportion (<1%) of VACV progeny are released from infected cells with a cell - derived membrane and become known as extra - cellular enveloped virus (EEV). Due to this additional membrane, EEV can be shielded from many anti -viral immune factors , allowing it to travel further and largely avoid host - mediated neutralization. This form of VACV is important for long range virus dissemination as well as sustained infection. Though the exact mechanism remains to be elucidated, it has been demonstrated that EEV release can be influenced by Abl tyrosine kinase (Abl TK) function. Specific point mutations in viral envelope proteins are known to bring about enhanced viral release, resulting in an elevated proportion of produced EEV. In this study, we investigate the effect of EEV enhancing modifications within various oncolytic VACV strains. Our data reveals that this augmentation of EEV production through the A34R L151E mutation within the Copenhagen (Cop) backbone can enhance the oncolytic potential of VACV in vivo through enhanced spread and immunoevasion.
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Becker, Michelle Caitlin. "The Combination of Carboxylesterase-Expressing Oncolytic Vaccinia Virus and Irinotecan." Thèse, Université d'Ottawa / University of Ottawa, 2013. http://hdl.handle.net/10393/23653.
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This project combines oncolytic Vaccinia virus (VV) with irinotecan (CPT-11) for the treatment of cancer. VV can infect, replicate in and destroy cancer cells, yet leave healthy cells relatively unaffected. CPT-11 is a chemotherapeutic of which ~5% is converted to the more active chemotherapeutic SN-38 by endogenous carboxylesterase (CE) enzymes. SN-38 is a topoisomerase I inhibitor that induces DNA double strand breaks, leading to growth arrest and apoptosis. Consequently, VV has been engineered to express a more effective isoform of the CE enzyme. The virus’ tumour tropism should restrict enhanced conversion of CPT-11 to the tumour.
Neither CPT-11 nor SN-38 interfered with VV replication or spread. Engineered recombinants expressed CE enzyme which, when combined with CPT-11, produced DNA double strand breaks and cancer cell death. In vitro, the combination of CE-virus and CPT-11 killed more K-562 cancer cells than its non-CE counterpart and CPT-11.
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Roy, Dominic. "Improving the Delivery and Replication of Oncolytic Viruses." Thesis, Université d'Ottawa / University of Ottawa, 2016. http://hdl.handle.net/10393/35227.
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The optimal route for clinical delivery of oncolytic viruses (OVs) is thought to be intravenous (IV) injection; however, the immune system is armed with several highly efficient mechanisms to remove pathogens from the circulatory system. To overcome the challenges in trying to deliver OVs IV, cell carriers have been investigated to determine their suitability as delivery vehicles for systemic administration of OVs. Here we demonstrate the utility of a Drosophila melanogaster cell platform for the production and in vivo delivery of multi-gene biotherapeutic systems. We show that cultured Drosophila S2 cell carriers can stably propagate OV therapeutics that are highly cytotoxic for mammalian cancer cells without adverse effects on insect cell viability or cellular gene expression. Drosophila cell carriers administered systemically to immunocompetent animals trafficked to tumours to deliver multiple biotherapeutics with little apparent off-target tissue homing or toxicity, resulting in a therapeutic effect. S2 cells provide a genetically tractable platform supporting the integration of complex, multi-gene biotherapies while avoiding many of the barriers to systemic administration of mammalian cell carriers.
Once OVs are delivered to tumour beds, they initiate replication in tumour cells, which often possess defects in antiviral pathways and are thus susceptible to infection. However, not all tumours have defects in their antiviral defenses and thus virus replication in these tumours is rather limited. Identifying and modulating host factors that regulate virus replication in OV-resistant cancer cells, but not normal cells, could lead to increased replication in these tumours and potentially improve therapeutic outcomes. We therefore conducted an RNA interference screen using Sindbis virus (SINV) in order to identify host factors that modulate OV replication in tumour cells. Specifically, serial passage of a SINV- artificial microRNA (amiRNA) library in a tumour cell line followed by deep sequencing of
ii
the selected virus populations led to the identification of several amiRNA sequences that were enriched. Furthermore, the identified amiRNA sequences increased the replication of various OVs both in vitro and in vivo, ultimately resulting in an enhanced therapeutic effect.
Overall, the work presented here highlights strategies in which both the systemic delivery and tumour-specific replication of OVs can be improved.
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Marguerie, Monique. "Combining the Immunogenic Cancer Mutanome with Oncolytic Virus Therapy." Thesis, Université d'Ottawa / University of Ottawa, 2014. http://hdl.handle.net/10393/31409.
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Oncolytic viruses (OVs) are effective anti-cancer agents, however their abilities to induce anti-tumor immunity are not yet optimal. Mutanome epitopes are a novel source of tumor antigen formed as a result of mutations within the tumor genome. Within this project we attempted to combine B16F10 mutanome vaccination with OV therapy. We confirmed previous findings that significant immune responses to these epitopes can be generated. Furthermore, we designed and cloned a multi-epitope mutanome construct into MG1 Maraba virus and E1-/E3- deleted type 5 Adenovirus to use for heterologous prime-boost vaccination. While we demonstrated that these viruses induced T-cell responses to one mutanome epitope, we failed to detect responses to the other epitopes. Furthermore there was no effect seen on overall survival. This approach warrants further investigation because coupling mutanome vaccination with OV therapy has the potential to exploit the therapeutic effects of the OV while inducing anti-tumor immunity to tumor-unique antigens.
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Aitken, Amelia. "Blocking the RNA Interference Pathway Improves Oncolytic Virus Therapy." Thesis, Université d'Ottawa / University of Ottawa, 2017. http://hdl.handle.net/10393/36821.
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Oncolytic viruses are novel candidates for cancer therapy and their efficacy relies on their capacity to overcome the host’s anti-viral barriers. In mammalian cells, the anti-viral response involves a protein-signaling cascade known as the interferon pathway, which alerts the immune system and limits the propagation of infection. Given that most cancer cells have defects in this pathway, they are susceptible to viral infection and responsive to oncolytic virotherapy. For reasons that remain unknown, many cancers are still refractory to oncolytic viruses, which suggests the existence of additional antiviral mechanisms. In this study, we investigate the potential involvement of an alternative antiviral pathway in cancer cells. Given that insects and plants rely on the RNA silencing pathway for their anti-viral protection, we investigated the presence of a similar mechanism in cancer cells. We found viral genome-derived small RNAs in various cancer cell lines upon infection, which is indicative of an RNA-mediated antiviral response. Also, various viruses encode suppressors of the RNA interference pathway. To determine if an oncolytic virus could benefit from such a factor, we engineered an oncolytic virus variant to encode the Nodamura virus B2 protein, a known inhibitor of RNA silencing-mediated immune responses. Using this virus, we observed enhanced cytotoxicity in 33 out of the 38 human cancer cell lines tested. Furthermore, our results show inhibition of viral genome cleavage and altered microRNA processing by our B2-expressing oncolytic virus. Taken together, our data suggests the blockade of RNA silencing antiviral pathways and/or antiviral microRNA processing improves the efficacy of our B2-encoding virus in a cell-line specific manner. Overall, our results establish the improved potential of our novel virus therapy and demonstrate for the first time the involvement of RNA pathways in the antiviral defense of cancer cells.
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Whilding, Lynsey May. "Activity of oncolytic vaccinia virus vectors in ovarian cancer." Thesis, Queen Mary, University of London, 2012. http://qmro.qmul.ac.uk/xmlui/handle/123456789/8553.
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Oncolytic vaccinia virus has great potential in the treatment of cancer and two engineered strains have entered clinical trials. As the advent for oncolytic vaccinia virus as an approved therapy beckons, it is critical to consider some of the barriers that may hinder this progress. These include suboptimal delivery of the virus to tumour sites, incomplete destruction of the tumour mass, and a lack of full understanding of the way in which oncolytic vaccinia kills its target cells. This thesis attempts to address these issues, with a particular focus on ovarian cancer. As ovarian cancer is generally restricted to the peritoneal cavity, intraperitoneal delivery may be preferable over intravenous delivery. Here, it is shown that Lister-dTK, an engineered vaccinia strain, is able to selectively replicate in ovarian tumours, including metastases to the liver following intraperitoneal delivery. To determine whether Lister-dTK could potentially be used in combination with current therapies for ovarian cancer, the effect of cisplatin and Lister-dTK together was assessed in vitro but showed no improvement in overall cell death. In an attempt to further improve the anti-tumour efficacy of Lister-dTK, the extracellular matrix protein (ECM) decorin was expressed from the virus. Decorin interacts with various signalling pathways and is proposed to enhance virus spread. However, abrogation of EGFR and TGFβ signalling could not be demonstrated in vitro, nor could improved virus spread. In an intraperitoneal model of ovarian cancer, Lister-mDCN did not demonstrate enhanced efficacy over a control virus. To determine the mechanisms of ovarian cancer cell death induced by Lister-dTK, the roles of apoptosis, autophagy and necrosis were investigated. Whilst some features of both apoptosis and autophagy were observed, inhibition of these pathways did not attenuate Lister-dTK. It is proposed that necrosis is the primary cause of cell death but that this process may occur in a regulated manner.
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Ishino, Ryo. "Oncolytic virus therapy with HSV-1 for hematologic malignancies." Doctoral thesis, Kyoto University, 2021. http://hdl.handle.net/2433/263570.
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Russell, Luke Russell. "Oncolytic Virus Expression of PTENα Directs Antitumor Immune Response." The Ohio State University, 2017. http://rave.ohiolink.edu/etdc/view?acc_num=osu1511979972361293.
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Dave, Rajiv Vipool. "Oncolytic vaccinia virus for the treatment of liver cancer." Thesis, University of Leeds, 2014. http://etheses.whiterose.ac.uk/6887/.
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Aims: Current treatment of colorectal cancer (CRC) liver metastases has a success rate of 50% 5-year survival, and recurrence rates of 50%. There is therefore still a need for a novel treatment modality. We aimed to examine the ability of JX-594 to preferentially replicate in and kill CRLM in vitro and ex vivo, and induce immunemediated tumour cytotoxicity by activation of natural killer (NK) cells. Methods: The Wyeth strain of vaccinia virus has been genetically manipulated to encode for granulocyte macrophage colony stimulating factor (JX-594-GM-CSFfLuc) and green fluorescent protein (JX-594-GM-CSF-GFP) in the disrupted thymidine kinase locus. Viability assays and Enzyme Linked Immunoabsorbent Assay (ELISA) was used to confirm tumour cell killing and production of inflammatory cytokines when CRC cell lines were infected with JX-594. Viral replication in vitro was investigated by plaque assay and using an ex vivo ‘tissue core’ method. Induction of the innate immune response was measured by upregulation of activatory markers on virus-treated-NK cells and monocytes by flow cytometry and anti-tumour cytotoxicity by chromium release. Results: JX-594 can directly lyse CRC cell lines, with greater lysis and replication (up to 250-fold) in cells with upregulated surface EGFR. JX-594 treatment resulted in substantial expression of GM-CSF and induction of inflammatory cytokines within the tumour microenvironment, and inhibition of anti-inflammatory and proangiogenic cytokines. Ex vivo infection of CRLM with JX-594-GFP-GM-CSF resulted in tumour-specific GFP and GM-CSF expression. Treatment of NK cells with JX-594-GM-CSF led to activation, degranulation and increased cytotoxicity against CRC cell targets. This was dependent on the presence of CD14+ve monocytes, which acquired an antigen-presenting phenotype (CD86+veCD11c+veClassIIDR+ve). Conclusions: JX-594 holds promise as a novel treatment modality for disseminated CRC. Direct tumour-specific lysis and transgene expression and the induction of tumour-specific innate immunity means that it may provide a twopronged attack against tumour cells whilst sparing normal tissue.
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Scut, Elena. "Characterization of the Immune Response Induced by Rhabdovirus-Infected Leukemia Cell Vaccines." Thesis, Université d'Ottawa / University of Ottawa, 2020. http://hdl.handle.net/10393/40944.
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Acute lymphoblastic leukemia (ALL) and acute myeloid leukemia (AML) are blood cancers that are often treated with stem cell transplantation (SCT). Since SCT treatments have variable success, especially in adults with AML whose disease frequently relapses, novel and more effective solutions must be considered. In this thesis, I will explore one type of immunotherapy in murine models for ALL (L1210) and AML (C1498) using in vitro and in vivo techniques such as flow cytometry and transcriptomics. In my approach, I am attempting to enhance the immunogenicity of whole cell vaccines by pre-infecting the leukemia cells with oncolytic virus (OV) and thus producing leukemia infected cell vaccines (ICVs). While it has been previously shown that L1210-ICV pre-treatment works well in protecting mice from ALL challenge, I have found that pre-immunization with C1498-ICV has a limited efficacy in protecting animals from AML progression. By investigating the downstream effects of ICV, I was able to show that unlike C1498 cells, L1210 cells produce previously unknown immunogenic factors following OV infection.
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Watson, Margaret. "Characterizing a Novel Viral Sensitizer BI-D1870." Thesis, Université d'Ottawa / University of Ottawa, 2019. http://hdl.handle.net/10393/39364.
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Oncolytic viruses (OVs) are an emerging cancer therapy that use an oncotropic virus to selectively infect and kill cancer cells, as well as stimulate long-lasting anti-tumor immune responses. In order to achieve high therapeutic efficacy, OVs need sufficient replication within the tumor tissue to mediate these effects. However, OV’s infectivity varies between different tumors and the host’s immune system can rapidly clear the virus, hampering treatment efficiency. Oncolytic virus sensitizers are chemical compounds that specifically enhance OV’s infectivity and efficacy. In our lab, I found that treatment of various cancer cell lines with BI-D1870, a pan-RSK (ribosomal S6 kinase) inhibitor, resulted in augmented Herpes Simplex Virus-1 (HSV1) and Vesicular Stomatitis Virus (VSVΔ51) infectivity. I also demonstrated that the effects of BI-D1870 on viral infection are virus-specific, and that RSK inhibition is not the primary target causing the enhancement of HSV1 and VSVΔ51 infection. Finally, BI-D1870 structural analogs were generated in an attempt to enhance the efficacy and selectivity of BI-D1870-based OV sensitizers. One of the analogs synthesized, KA-019, showed an improvement in the augmentation of OV infection over BI-D1870. As a genetically engineered strain of HSV1 has been approved by FDA for treatment of melanoma, the results of my project propose a novel viral sensitizer to improve viral replication within tumour cells with the hope of improving therapeutic efficacy.
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Ramachandran, Mohanraj. "Cancer Immunotherapy : Evolving Oncolytic viruses and CAR T-cells." Doctoral thesis, Uppsala universitet, Science for Life Laboratory, SciLifeLab, 2016. http://urn.kb.se/resolve?urn=urn:nbn:se:uu:diva-302891.
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In the last decade cancer immunotherapy has taken huge strides forward from bench to bedside and being approved as drugs. Cancer immunotherapy harnesses the power of patient’s own immune system to fight cancer. Approaches are diverse and include antibodies, therapeutic vaccines, adoptively transferred T-cells, immune checkpoint inhibitors, oncolytic viruses and immune cell activators such as toll-like receptor (TLR) agonists. Excellent clinical responses have been observed for certain cancers with checkpoint antibodies and chimeric antigen receptor (CAR)-engineered T-cells. It is however becoming evident that strategies need to be combined for broader effective treatment responses because cancers evolve to escape immune recognition. A conditionally replication-competent oncolytic adenovirus (Ad5PTDf35-[Δ24]) was engineered to secrete Helicobacter pylori Neutrophil Activating Protein (HP-NAP, a TLR-2 agonist) to combine viral oncolysis and immune stimulation. Treatment with Ad5PTDf35-[Δ24-sNAP] improved survival of mice bearing human neuroendocrine tumors (BON). Expression of HP-NAP in the tumor microenvironment promoted neutrophil infiltration, proinflammatory cytokine secretion and increased necrosis. We further studied the ability of HP-NAP to activate dendritic cells (DCs) a key player in priming T-cell responses. HP-NAP phenotypically matured and activated DCs to secrete the T-helper type-1 (Th-1) polarizing cytokine IL-12. HP-NAP-matured DCs were functional; able to migrate to draining lymph nodes and prime antigen-specific T-cell proliferation. CAR T-cells were engineered to secrete HP-NAP upon T-cell activation. Secreted HP-NAP was able to mature DCs, leading to a reciprocal effect on the CAR T-cells with improved cytotoxicity in vitro. Semliki Forest virus (SFV), an oncolytic virus with natural neuro-tropism was tagged with central nervous system (CNS)-specific microRNA target sequences for miR124, miR125 and miR134 to selectively attenuate virus replication in healthy CNS cells. Systemic infection of mice with the SFV4miRT did not cause encephalitis, while it retained its ability to replicate in tumor cells and cure a big proportion of mice bearing syngeneic neuroblastoma and gliomas. Therapeutic efficacy of SFV4miRT inversely correlated with type-I antiviral interferon response (IFN-β) mounted by tumor cells. In summary, combining immunotherapeutic strategies with HP-NAP is a promising approach to combat cancers and SFV4miRT is an excellent candidate for treatment of neuroblastomas and gliomas.
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Šilanskas, Mantas. "Oncolytic viruses armed with immunostimulatory genes for cancer treatment." Thesis, Uppsala universitet, Institutionen för immunologi, genetik och patologi, 2018. http://urn.kb.se/resolve?urn=urn:nbn:se:uu:diva-353153.
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Cancer is a major health burden in modern society, costing millions of lives worldwide and negatively impacting many more. With increasing rates of cancer, there is a need for new approaches to its treatment. This is where immunotherapies step in, this a relatively new approach to cancer treatment which caught public’s attention only in recent years. The main goal of these therapies is to enhance and help immune cells to identify and kill tumor cells, thereby initiating the cycle of cancer immunity. In this project LOAd platform viruses were evaluated and compared for their ability to induce oncolysis in cancer cells and ability to produce immunostimulatory molecules. Established LOAd703 virus armed with CD40L and 4-1BBL transgenes was compared to new constructs LOAd732, LOAd780 and LOAd786. All three new viruses are armed with CD40L and 4-1BBL, but also have additional transgenes X, Y and Z, respectively. Specific molecules coded by these transgenes cannot be disclosed at this moment. All viruses demonstrated high competence in oncolysis of A549-lung, T24-bladder and 526-mel melanoma cancer cell lines and were able to express transgenes coding for CD40L and 4-1BBL in all cell lines. New viruses were able to induce expression of new transgenes in infected cells, except for LOAd780 infected cell which had low concentration of protein Y in their supernatants. Also dendritic cells matured using LOAd viruses were able to induce expansion of CMV-specific T cells and a major expansion of natural killer cells.
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Bolyard, Chelsea M. "Oncolytic Virus Therapy in Combination with Chemotherapy for Ovarian Cancer." The Ohio State University, 2013. http://rave.ohiolink.edu/etdc/view?acc_num=osu1386007453.
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Ferguson, Mark Simon. "Enhancement of systemic delivery of oncolytic Vaccinia virus for cancer treatment." Thesis, Queen Mary, University of London, 2014. http://qmro.qmul.ac.uk/xmlui/handle/123456789/11668.
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Survival for patients with advanced cancer has remained dismal, and there is a need for new treatments. In this context viral immune therapy is a promising novel strategy. Intravenous delivery confers advantages as it enables simultaneous treatment of primary tumour and any metastatic deposits but host defences limit Vaccinia virus's (VV) ability to infect tumour after systemic administration. Although Vaccinia virus can potentially be delivered systemically as it can evade both complement and neutralising antibodies, our investigations have revealed that VV cannot effectively infect tumour cells in immunocompetent mice after systemic delivery. Strikingly, we observed that if macrophages were depleted in the mice using clodronate liposomes, VV infection of tumours was dramatically enhanced. However, clodronate liposomes non-selectively deplete macrophages and potentially diminish any beneficial macrocytic activity in the tumour microenvironment unrelated to viral clearance. Consequently, a more clinically appropriate agent is needed. Macrophages recognise and ingest pathogenic microorganisms through phagocytosis, a process for which several lines of evidence have highlighted an important role for phosphatidylinositol 3-kinases. Accordingly, in these investigations I have evaluated the effect of selective PI3K inhibitors on macrophage phagocytosis in vitro and demonstrated that IC87114 (a PI3 kinase delta inhibitor) is effective at reducing uptake of VV by macrophages, confirming this finding in transgenic macrophages with a mutant of the PI3 kinase delta isoform knocked in. Subsequently, it was confirmed that IC87114 affects attachment of the virus to macrophages but plays no role in internalisation of the virus. In cancer cells cultured in isolation, the inhibitor has no direct cytotoxic effect and when combined with VV, in the same in vitro system, there is no change in the amount of cell death compared to VV alone treated controls. Biodistribution studies have established that IC87114 combined with VV results in statistically significantly higher levels of virus detected in tumours compared to the groups treated with VV alone, with similarly limited off-target effects. Finally, three different efficacy studies have demonstrated statistically significantly superior tumour responses in the VV+IC87114 group. In conclusion, PI3k delta blockade is an effective strategy for enhancing systemic delivery of VV in a preclinical model and could be a useful adjuvant in VV clinical trials.
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Pizzuto, Matteo Samuele. "Avian influenza virus as an oncolytic therapy for pancreatic cancer." Thesis, Imperial College London, 2015. http://hdl.handle.net/10044/1/38476.
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Pancreatic ductal adenocarcinoma (PDA) is one of the leading causes of cancer-related deaths worldwide and the development of new treatment strategies for patients suffering from PDA is of crucial importance. Virotherapy uses natural or engineered oncolytic viruses (OVs) to selectively kill tumour cells. Although various OVs are being investigated as agents for pancreatic cancer treatment, due to the genetic heterogeneity of PDA cells and their consequent mixed permissiveness to viruses, virotherapy should not rely on a short list of possible candidates. As such, preliminary data from our group demonstrating the potent pro-apoptotic effect of the low pathogenicity avian influenza virus (LPAIV) H7N3 A/turkey/Italy/2962/03 in PDA cells, previously established to be resistant to other OVs, suggested that this virus might be effective against specific sub-classes of pancreatic cancer. Therefore, in the present studies, the avian isolate was selected for further development. Preferential replication of the H7N3 virus in IFN-deficient cells, a trait of the majority of PDA cell lines, was improved by the truncation of the viral NS1 protein (NS1-77), which compromised the virus' ability to counteract the IFN-mediated antiviral response and, as bystander effect, enhanced the killing of uninfected cancer cells by stimulating IFN expression from healthy cells. Introduction of L75H mutation within the mitochondrial targeting sequence (MTS) of the viral protein PB1-F2 increased virus replication in permissive IFN deficient cells and also lessened PB1-F2's ability to counter the IFN response induced by overexpression of mitochondrial antiviral signalling protein (MAVS), further enhancing selectivity for PDA cells. Substitution of the genes expressing avian surface antigens haemagglutinin (HA) and neuraminidase (NA) with those of the well characterized human H1N1 A/Puerto Rico/8/1934 virus eliminated the potential risk of reassortment of the H7N3 virus with circulating human influenza strains that might result in a new pandemic virus, nonetheless, preserving the oncolytic properties of the parental isolate. Finally, to promote anti-tumour immunity, the newly generated PR8 H1/N1-2962 NS1-77 virus was 'armed' to express the human granulocyte-macrophage colony-stimulating factor (GM-CSF) gene during infection. Taken together our data demonstrate the possibility to select an IAV from the avian reservoir on the basis of its strain-specific oncolytic skills in PDA cells and, through engineering, improve its selectivity, safety and tumour debulking activity.
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Raghunath, Shobana. "Targeted Oncolytic Virotherapy Using Newcastle Disease Virus Against Prostate Cancer." Diss., Virginia Tech, 2012. http://hdl.handle.net/10919/77985.
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Prostate cancer (CaP) is the second leading cause of cancer related deaths in men in the United States. Currently, androgen depletion is an essential strategy for CaP combined with surgery, chemotherapy and radiation. Hormone independent cancer stem cells escaping conventional therapy present a major therapeutic challenge. The available treatment regimens for hormone resistant CaP are only palliative and marginally increase survival. Therefore, novel strategies to eradicate CaP including stem cells are imperative. Oncolytic virus (OV) therapy is a novel approach that overcomes the limitations posed by radiation and chemotherapy. Oncolytic virotherapy of cancer is based on the use of replication competent, tumor selective viruses with limited toxicity. Newcastle Disease Virus (NDV), an avian paramyxovirus, is a safe and promising OV successfully used in many clinical trials. NDV is inherently tumor selective and cytotoxic but replication restricted in normal cells. But, systemically delivered NDV fails to reach solid tumors in therapeutic concentrations and also spreads poorly within the tumors due to barriers including complement, innate immunity and extracellular matrix. Overcoming these hurdles is paramount to realize the exceptional oncolytic efficacy of NDV. Therefore, we engineered the fusion (F) glycoprotein of NDV and generated a recombinant NDV (rNDV) cleavable exclusively by prostate specific antigen (PSA). The rNDV replicated efficiently and specifically only in prostate cancer (CaP) cells but failed to replicate in the absence of PSA. Further, PSA-cleavable rNDV caused specific lysis of androgen independent and dependent/responsive CaP cells with a mean effective concentration (EC50) ranging from 0.01 to 0.1 multiplicity of infection (MOI). PSA retargeted rNDV efficiently lysed three-dimensional prostaspheres, suggesting efficacy in vivo. Also, PSA-cleavable NDV failed to replicate in chicken embryos, indicating absence of pathogenicity to its natural host, chickens. Prostaspheres generated from DU-145 CaP cell line derived xenografts showed self-renewal, proliferative and clonogenic potential in vitro, and exhibited increased tumorigenicity in vivo. Embryonic stem and progenitor cell markers like Nanog, Nestin and CD44 were overexpressed in spheres as compared to the cell line suggesting prostaspheres comprise tumor-initiating cells from CaP. Xenograft and cell line derived prostaspheres were permissive for rNDV replication, when the fusion protein was activated by exogenous PSA. The EC50 against tumor initiating cells was 0.11-0.14 MOI, suggesting an excellent therapeutic margin for in vivo studies. PSA retargeting is likely to enhance the therapeutic index of rNDV owing to tumor restricted replication and enhanced fusogenicity. Our results suggest PSA retargeted rNDV selectively replicates and lyse PSA producing CaP cells including tumor-initiating cells and is a promising candidate for immediate Phase I/II clinical trials.Ph. D.
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Rintoul, Julia. "ORFV: A Novel Oncolytic and Immune Stimulating Parapoxvirus Therapeutic." Thèse, Université d'Ottawa / University of Ottawa, 2012. http://hdl.handle.net/10393/22925.
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Replicating viruses for the treatment of cancer have a number of advantages over
traditional therapeutic modalities. They are highly targeted, self-amplifying, and have the
added potential to act as both gene-therapy delivery vehicles and oncolytic agents. ORFV,
(Parapoxvirus ovis, or Orf virus) is the prototypic species of the Parapoxvirus genus,
causing a benign disease in its natural ungulate host. ORFV possesses a number of unique
properties that make it an ideal viral backbone for the development of a cancer therapeutic: it
is safe in humans, has the ability to cause repeat infections even in the presence of antibody,
and it induces a potent Th-1 dominated immune response. Here I show for the first time that
live replicating ORFV induces an anti-tumour immune response in multiple syngeneic mouse
models of cancer that is mediated largely by the potent activation of both cytokine-secreting,
and tumouricidal natural killer (NK) cells. I have also highlighted the clinical potential of the
virus by demonstration of human cancer cell oncolysis including efficacy in an A549
xenograft model of cancer. The mechanism of ORFV-mediated activation of NK cells has
been explored, where I have demonstrated activation via direct ex vivo infection of NK cells.
I have also highlighted ORFV-mediated activation of dendritic cells (DCs), both in vivo and
by direct infection ex vivo. An in vivo DC depletion study demonstrated an indirect
mechanism for ORFV NK cell activation, where in the absence of DCs, NK cell activation
was diminished, as was the ability of ORFV to clear lung metastases. The ORFV innate
immune stimulatory profile has been harnessed for therapeutic application in an experimental
surgery model of cancer, where ORFV therapy at the time of surgery reduces the number of
cancer metastases. These data highlight the clinical potential of a live, immune stimulating
Parapoxvirus therapeutic.
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Rowe, Katelynn. "Validating Transgenic Farmington Viruses for the Treatment of Glioblastoma Multiforme." Thesis, Université d'Ottawa / University of Ottawa, 2015. http://hdl.handle.net/10393/33354.
Full textAbstract:
Glioblastoma is the most common primary brain tumour in adults. Despite the aggressive standard of care currently used, median patient survival following treatment is only 14 months. Innovative treatment options are needed for these patients. Recently, oncolytic viruses have emerged as promising immunotherapies for the treatment of solid tumours. Preliminary work in our lab has demonstrated that Farmington virus, a novel brain-safe oncolytic rhabdovirus, can be engineered to encode a tumour-associated antigen (TAA) to prime and boost antigen-specific adaptive immune responses. Since other rhabdoviruses share this boosting capacity, a heterologous rhabdovirus prime/boost regimen can be designed to combine two powerful oncolytics and a robust anti-TAA adaptive immune response. We evaluated Farmington’s ability to vaccinate against a self- glioblastoma antigen and two foreign glioblastoma-associated antigens. Farmington was able to vaccinate against the foreign antigens, leading to efficacy in prophylactic and therapeutic glioblastoma models. Additionally, treatment with heterologous rhabdoviruses demonstrated efficacy in an aggressive murine mammary carcinoma model. Herein, we demonstrate promising preliminary results for a novel glioblastoma therapeutic approach.
Le glioblastome est la tumeur primaire la plus fréquente chez l’adulte. La survie moyenne des patients n’excède pas 14 mois malgré une prise en charge thérapeutique agressive. Par conséquent, la mise au point de traitements innovants et efficaces est une nécessité pour ces patients. Des avancées récentes ont mise en évidence l’intérêt des virus oncolytiques dans le traitement des tumeurs solides. Des travaux préliminaires réalisés au sein de notre laboratoire ont, en effet, démontré que le virus Farmington pouvait être modifié afin d’exprimer un antigène associé aux tumeurs (AAT), pour initier et potentialiser une réponse immunitaire adaptative spécifique. D’autres rhabdovirus possèdent des capacités de potentialisation immunitaire similaires et peuvent être utilisés en association avec le virus Farmington modifié pour amorcer et amplifier la réponse immunitaire oncolytique de l’hôte. Le but de ce projet était d’évaluer le potentiel du virus Farmington comme vaccin contre des antigènes tumoraux d’origine endogène ou exogène associés au glioblastome. Nos résultats ont montré que le virus Farmington a la capacité d’induire une réponse immunitaire prophylactique et thérapeutique contre les antigènes tumoraux exogènes dans des modèles de glioblastome. De plus, l’utilisation de rhabdovirus hétérologues s’est aussi révélée efficace pour le traitement de carcinome mammaire agressif chez la souris. Cette étude préliminaire apporte des résultats prometteurs pour le développement de nouvelles approches thérapeutiques efficaces dans le traitement du glioblastome.
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Calderon, Hugo. "Investigating the oncolytic properties of a group B adenovirus on cancer cells and its effects on the local immune response." Thesis, University of Oxford, 2017. http://ora.ox.ac.uk/objects/uuid:da89b317-5f76-4447-bbb1-26740db3b3ef.
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Oncolytic viruses are characterised by their ability to selectively infect and kill tumour cells. Recently it has emerged that they can exert an additional anticancer mechanism stimulating adaptive immune-mediated cancer cell killing. Enadenotucirev (EnAd, formerly known as ColoAd1), is a chimeric Ad11p/Ad3 virus group B oncolytic adenovirus that binds CD46 and is under development for the systemic treatment of metastatic carcinomas. The central aim of this thesis was to to assess whether EnAd provides an adjuvant effect on tumour-associated antigen presenting cells (APCs) that could drive TH1 polarisation for an effective anti-tumour immune response. This thesis describes the potent oncolytic properties, fast replication and high numbers of virus progeny production by EnAd in cancer cells. Recombinant EnAd variants were engineered to investigate the roles of the mutant regions in the genome of EnAd, and how these influence the modified phenotype. A chemical drug panel was used to identify pathways and cellular factors involved in cellular production of EnAd, finding that several mTOR inhibitors and microtubule inhibitors could improve virus replication. An in vitro system using partially matured human monocyte-derived dendritic cells (DCs), which displayed a similar phenotype to tumour-infiltrating DCs, was used to explore the effect of EnAd on APC responses. EnAd induced a strong adjuvant effect on these cells by up-regulating surface markers and secretion of pro-inflammatory factors. Further mechanistic experiments, alongside a CAR-binding group C adenovirus 5, indicated these adjuvant effects were virus particle-mediated and dependent on CD46 binding. To understand the functional implications downstream of these interactions, T cell activation and phenotype was assessed using a mixed lymphocyte reaction approach. The data indicated EnAd was a good candidate compared to other adenoviruses, that may steer the response of activated T-cells towards a TH1 phenotype, for an effective immune response. In conclusion, the potent oncolytic properties of EnAd virus may provide an adjuvant effect on tumour-associated APCs, helping to harness an adaptive immune response.
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McKenzie, Christopher David. "Activation of oncogenic signalling pathways by vaccinia virus." Thesis, The University of Sydney, 2016. http://hdl.handle.net/2123/15789.
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Vaccinia virus (VACV) is a dsDNA virus and a member of the Poxviridae family. VACV induces a wide range of morphological changes to cells during infection, collectively known as cytopathic effect (CPE). VACV-induced CPE includes loss of cell–cell contacts between neighbouring cells, induction of cell migration and extensive remodelling of the actin cytoskeleton in order to aid viral egress. These characteristic changes during VACV infection closely resemble the changes associated with epithelial-mesenchymal transition (EMT) — an important process in development, wound healing and cancer progression. EMT is controlled by multiple signalling pathways, with three of the most well-characterised pathways being the Wnt, TGFβ and Notch. We hypothesised that these same signalling pathways may also be involved in inducing the EMT-like changes observed during VACV infection. Using a luciferase reporter assay, we show that VACV activates TGFβ/Smad signalling but not Wnt/β-catenin signalling. Using both an inhibitor of the type-I TGFβ receptor and a cell line that lacks expression of the TGFβ receptor we found that VACV activates Smad signalling independent of receptor activity. We found that activation of Smad signalling is common to multiple strains of VACV but not the highly attenuated MVA strain or the related poxvirus, ectromelia virus. We identify and test several candidate VACV genes for their role in activation of Smad signalling and subsequently describe efforts to map candidate Smad-activating genes by partial rescue of the MVA genome. Finally, we aim to examine the contribution of Smad signalling activation to plaque phenotype in VACV. By adding TGFβ to exogenously activate Smad signalling in ECTV we found that activation of TGFβ signalling increases plaque clearance and size in ECTV plaques, as well as further increasing the size of VACV plaques, suggesting a role for activation of TGFβ/Smad signalling in the spread of infected cells within plaques.
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Tehranipour, Pegah. "Treatment outcomes on malignant gliomas using oncolytic viruses." Thesis, Uppsala universitet, Institutionen för farmaceutisk biovetenskap, 2020. http://urn.kb.se/resolve?urn=urn:nbn:se:uu:diva-412395.
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Purpose: The objective of this thesis is to evaluate clinical studies that have used oncolytic viruses as treatment and to compare their treatment-outcomes on patients with malignant glioma. Method: This thesis is a systematic literature review where PubMed has been used as the database for data collection. Two searches were done using the search phrases oncolytic virus AND Glioma and oncolytic virus AND brain tumor. Several of the articles showed up multiple times in different searches. After having applied the inclusion criteria, ten of the seventeen articles were removed. Remaining were seven articles used for the thesis. Results: The study conducted by Forsythe et al., using reovirus showed the median overall survival (OS) to be 21 weeks and the median time to progression (TTP) was 4.3 weeks. The study conducted by Kicielinski et al., using REOLYSIN showed the median OS to be 140 days. Median TTP was 61 days. The study conducted by Geletneky et al., 2017 was the first dose-escalating clinical trial for the use of H-1 parvovirus. The median TTP was 111 days and the median OS was 464 days. The study conducted by Lang et al., DNX-2401 was used and in group A the median OS time was 9.5 months. In group B the median OS in the group was 13 months. In another example of an oncolytic adenovirus is ONYX-015, the median TTP after treatment for all patients was 46 days. The median OS for patients diagnosed with glioblastoma multiforme was 4.9 months and for patients with anaplastic astrocytoma and anaplastic oligodendroglioma was 11.3 months across. In a study conducted by Freeman et al. using newcastle disease virus, the OS ranged from 3-66 weeks from the start of treatment and TTP ranged from 2-53 weeks. The study conducted by Markert et al., the median OS from treatment with G207 was 7.5 months. The median TTP was around 2.5 months. Conclusion: Oncolytic viruses are promising agents for treatment against malignant gliomas. No definite outcomes of the treatment could be concluded, however, the median survival was extended in certain cases. The patients tolerated the oncolytic viruses well with no adverse effects correlated with the treatments. There are currently more virus vectors being tested as new developments are needed in this field.
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Leja, Justyna. "Oncolytic Adenovirus Therapy of Neuroendocrine Tumors." Doctoral thesis, Uppsala universitet, Institutionen för immunologi, genetik och patologi, 2011. http://urn.kb.se/resolve?urn=urn:nbn:se:uu:diva-146966.
Full textAbstract:
Neuroendocrine tumors (NETs), originally described as carcinoids, represent a rare and heterogeneous group of neoplasms associated with intensive secretion of hormones, bioactive peptides and amines. Most of the patients are diagnosed at a late stage of disease, often with liver metastases. Surgery remains the main treatment to control metastatic disease, but is not curative. Oncolytic virotherapy represents a promising approach to treat cancer and different strategies have been exploited to restrict viral replication to tumor cells. We developed an oncolytic adenovirus based on serotype 5, Ad5[CgA-E1A], where the chromogranin A (CgA) promoter controls expression of the E1A gene and thereby virus replication. We found that Ad5[CgA-E1A], selectively replicates in NET cells and it is able to suppress fast-growing human BON carcinoid tumors in nude mice. The activity of Ad5[CgA-E1A] was not completely blocked in liver cells. We further repressed virus replication in hepatocytes by targeting E1A with miR122, an miRNA specifically expressed in the liver. miRNAs bind to mRNA and induce its cleavage or translational blockage. By insertion of tandem repeats of miR122 target sequences in 3’UTR of E1A gene, we observed reduced E1A protein expression and replication arrest in miR122 expressing liver cells. The oncolytic potency of the miR122-targeted virus was not affected in NET cells. Since some NET and neuroblastoma cells express high levels of somatostatin receptors (SSTRs), we introduced in the virus fiber knob cyclic peptides, which contain four amino acids (FWKT) and mimic the binding site of somatostatin for SSTRs. The FWKT-modified Ad5 transduces midgut carcinoid cells from liver metastases about 3-4 times better than non-modified Ad5. Moreover, FWKT-modified Ad5 overcomes neutralization in an ex vivo human blood loop model to a greater extent than Ad5, indicating that the fiber knob modification may prolong the systemic circulation time. NETs represent a huge therapeutic challenge and novel diagnostic markers are needed for early detection and effective treatment of NETs. We have profiled primary tumors and liver metastases of ileocaceal NETs, using Affymetrix microarrays and advanced bioinformatics. We have identified six novel marker genes and show high similarity between primary lesions and liver metastases transcriptome by hierarchical clustering analysis.
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Kus, Agnieszka. "Neoadjuvant Oncolytic Virus Therapy in a Murine Model of Cancer Surgery." Thesis, University of Ottawa (Canada), 2010. http://hdl.handle.net/10393/28847.
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Surgery is the primary treatment modality for most solid tumours. Despite complete resection, the development of metastatic disease limits it curative potential and provides the rationale for neoadjuvant (preoperative) therapies. Oncolytic Viruses (OVs) are replicating therapeutics that are selected or engineered to grow in malignant cell types and are capable of killing the infected target cell, while leaving normal, adjacent cells unharmed. OVs may be an ideal candidate for generating a potent anti-tumour immune response due to effective recruitment of immune cells into the tumour microenvironment, proper activation of immune cells, and generation of tumour antigen-specific T cells. Preoperative OV therapy may serve as a neoadjuvant immunotherapy capable of preventing the development of metastatic disease by generating an effective immune response. Although VSVDelta51GM-CSF treatment of CT26lacZ tumours is able to generate an anti-tumour immune response capable of preventing the growth of a subsequent CT26lacZ tumour, preoperative VSVDelta51GM-CSF treatment of primary tumours in a CT26lacZ surgery model was unable to generate an immune response capable of rejecting a secondary tumour. This abrogation of protection against the secondary CT26lacZ tumour was observed to be a result of the surgical intervention. Preoperative VSVDelta51GM-CSF and VSVDelta51 treatment IV of primary tumours in a B16-F10 surgery model was unable to generate an immune response capable of reducing the number of secondary surface lung metastases. JXS94mGMCSF treatment IT of a primary tumour in a B16-F10 surgery model was able to reduce the number of secondary surface lung metastases, while JXS94mGMCSF injected IV was not. This suggests that one of the factors in the induction of an anti-tumour immune response by neoadjuvant therapy in a surgical model may be the route of administration of ov.
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Leddon, Jennifer. "Oncolytic Herpes Simplex Virus Therapy for the Treatment of Pediatric Rhabdomyosarcoma." University of Cincinnati / OhioLINK, 2015. http://rave.ohiolink.edu/etdc/view?acc_num=ucin1427980753.
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Hoang, Huy-Dung. "Translation Control to Improve Oncolytic Virus Efficacy and Regulate Inflammatory Diseases." Thesis, Université d'Ottawa / University of Ottawa, 2021. http://hdl.handle.net/10393/42410.
Full textAbstract:
Translation control is crucial during virus-host interaction, in which the host relies on the translation machinery to mount an antiviral response or induce the inflammation response to reduce virus spread, while the virus aims to take control of this system to thwart the host defense while producing viral progeny. The field of oncolytic virus (OV) therapy relies on replicating, engineered viruses that preferentially infect tumor cells to induce direct oncolysis or promote an antitumor immune response. Despite the importance of translation control in virus-host interaction, not much has been described on the interaction at the translation level between OV and cancer cells. I propose that this knowledge gap could reveal significant improvements in OV efficacy in treating cancer. In my first study, I set out to characterize the translatome of an infection-resistant breast cancer cell line infected by three clinically advanced OVs to identify residual antiviral activity in cancer cells regulated by translation control. I found the inositol phosphatase Inpp5e to be a novel antiviral gene that is translationally induced during infection via a transcript variant shift. Mechanistically, I showed that the majority of Inpp5e transcripts in uninfected cells contain a long 5’ UTR that harbor four translationally inhibitory upstream reading frames (uORF). Yet, OV infection induced the expression of a shorter 5’ UTR with a spliced intron that removes three uORFs, derepressing the translation of Inpp5e mRNA. CRISPR-Cas9 knockout of Inpp5e also enhanced the infectivity of oncolytic HSV1 and VSV. My study suggests the existence of a class of translationally regulated antiviral genes in cancer cells. In my second study, I sought to adapt the translation of transgenes to the unique translation condition imposed by the infecting virus via the incorporation of a viral 5’UTR. I identified HSV1 5’UTRs by locating the transcription start site of most HSV1 genes using RNA-seq data, then determined the 5’UTR of US11 as a potent translation enhancer during HSV1 infection. Incorporation of this 5’UTR into the transgene expression cassette inserted into the HSV1 genome enhanced transgene expression significantly at the translation level. In my third study, I set out to explore the mechanism of miR-223 mediated inflammation inhibition. miR-223 is a protective miRNA in the context of atherogenesis via suppressing inflammatory signaling. Using transcriptome and translatome profiling (RNA-seq and Ribo-seq), I found that the inhibitory effect of miR-223 on inflammation occurs primarily at the translation level. Overall, my work highlights the importance of translation control in OV-cancer cells interaction, as well as in inflammation-related diseases.
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Krishnan, Ramya. "Characterization of Novel Small Molecule Potentiators of Oncolytic Virotherapy." Thesis, Université d'Ottawa / University of Ottawa, 2018. http://hdl.handle.net/10393/37551.
Full textAbstract:
The use of oncolytic viruses (OVs) to selectively destroy cancer cells is poised to make a major impact in the clinic and potentially revolutionize cancer therapy. Pre-clinical and clinical studies have shown that OV therapy is safe, well-tolerated and effective in a broad range of cancers. Still, resistance due to tumour heterogeneity highlights areas for improvement in OV based therapeutics. Combining OVs and small molecules is a promising strategy to selectively enhance OV-mediated anti-tumour effects. To this end, we have previously identified the synthetic compound Viral Sensitizer 1 (VSe1) that enhances the spread of oncolytic vesicular stomatitis virus (VSVΔ51) in resistant cancer cell lines up to 1000-fold, resulting in synergistic cell killing and improved efficacy in vitro and in vivo. The electrophilic nature of VSe1 prompted us to investigate the scaffold to identify active analogs with more favourable physiochemical properties and explore structure-activity relationships (SAR). In vitro assays and a rational approach in the design of VSe1 analogs allowed us to identify functional groups that can be modified without hampering activity. Lead compounds created in this study based on a pyrrole scaffold increase OV growth up to 2000-fold in vitro and demonstrate remarkable selectivity for cancer cells over normal tissue ex vivo and in vivo. Compared to the parental VSe1, these small molecules also possess enhanced stability with reduced electrophilicity and are well-tolerated in animals, leading to reduced tumour burden and prolonged survival in vivo when used in combination with VSVΔ51.
It was known from previous studies that VSe1 suppresses the type I interferon response generated by cancer cells to defend against viral infection. In this study, further investigation revealed that VSe1 and its analogs inhibit the nuclear translocation of nuclear factor kappa-light-chain-enhancer of activated B cells (NFκB), resulting in dampened transcriptional expression and secretion of IFN-β and interferon stimulated genes, thereby increasing viral replication and spread. While these findings further elucidated the effect these compounds have on the innate antiviral response, the molecular mechanisms leading to NFκB inhibition remained unclear. We used the newly generated VSe1 analogs to perform ligand-based affinity capture studies leading to the identification of glutathione-s-transferases as interacting proteins, catalytically inhibited by VSe1 and to a lesser extent by its pyrrole analogs. Further inquiry revealed that VSe1 and its analogs cause an imbalance in cellular glutathione homeostasis and increase oxidative stress, which is associated with inhibition of the nuclear translocation of NFκB. However, further studies are required to assess whether these phenomena are directly or indirectly linked.
Overall, this study highlights a novel approach to improving OV therapy by using a previously uncharacterized class of compounds that ultimately alter the innate cellular antiviral response through inhibition of NFκB.
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Wedge, Marie-Ève. "Tailoring Oncolytic Viruses for the Treatment of Pancreatic Cancer." Thesis, Université d'Ottawa / University of Ottawa, 2020. http://hdl.handle.net/10393/40384.
Full textAbstract:
Pancreatic cancer (PC) is a highly aggressive disease with unmet therapeutic needs. Recent advances in the use of oncolytic viruses (OVs) as cancer therapeutic agents bring new hope to fight the notorious disease that is PC. Although OVs have shown promising results in certain cancers, some tumors remain resistant to OV therapy due to their inherent residual antiviral mechanisms. We hypothesized that the use of OV-encoded artificial microRNAs (amiRNAs) could help target the cellular antiviral components associated with the observed OV resistance and could also sensitize neighboring tumor cells to OV therapy and small molecule inhibitors through the secretion of amiRNA-containing extracellular vesicles (EVs) from infected cells. To find such amiRNAs, a viral surrogate library encoding ~16,000 unique amiRNAs was passaged in pancreatic cancer cell lines to enrich for sequences that could enhance OV replication. An amiRNA that improves PC cell killing when expressed from an OV was identified. Target identification of this amiRNA (amiR-4) revealed ARID1A as a key player in resistance to OV therapy in pancreatic cancers. This target is of particular interest, since its downregulation acts in a synthetic lethal fashion with inhibition of the EZH2 methyltransferase. Combining VSV51-amiR-4 with a small molecule inhibitor of EZH2 enhances PC cell death. Moreover, amiR-4 is packaged in cancer cell-secreted EVs which can reach neighboring naïve cells to sensitize them to EZH2 inhibition-mediated cell death and to spread the OV-mediated tumor killing effect throughout the tumor. This data translates into tumor debulking and survival in animal models of highly aggressive PC. This work not only broadens our knowledge on the resistance of select tumors to oncolytic virotherapy and the EV-mediated bystander killing effect in OV-infected tumors, but it also establishes OVs as a novel tool to produce anti-cancer therapeutic EVs in situ to improve therapeutic gain. Ultimately, our work provides new hope for a cure to the grim disease that is PC.
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Pinkl, Joseph T. "Characterizing Osteologic Effects of Cholesteatoma and Oncolytic Virotherapy." University of Cincinnati / OhioLINK, 2021. http://rave.ohiolink.edu/etdc/view?acc_num=ucin1627663262820533.
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Barkley, Russell. "Investigation of an Oncolytic MeV Cell-Cell Fusion Phenomenon Induced by an siRNA." Thesis, Université d'Ottawa / University of Ottawa, 2020. http://hdl.handle.net/10393/41531.
Full textAbstract:
Oncolytic measles virus is a promising cancer therapeutic in clinical trials which
possesses multiple characteristics that are advantageous over traditional therapies.
Currently, clinical oncolytic measles virus vectors are unmodified or express reporter
transgenes that benefit its therapeutic efficacy. The next phase in its development will
see genetically engineered vectors encoding transgenes that enhance its antineoplastic effects. To this end, preclinical research has focused on studying novel transgenes which favour viral replication, cytotoxicity, and the anti-cancer immune response. We sought to encode artificial micoRNAs targeting RIG-I as a strategy to interfere with innate immunity. Silencing RIG-I with multiple siRNAs yielded one which promotes measles virus syncytia formation through a mechanism that appears to be independent of RIG-I. The mechanism caused by the siRNA leads to enhanced measles virus cell-cell fusion and has peculiar characteristics which are not fully understood.
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Alkayyal, Almohanad. "Exploiting the Antitumor Immune Response Using IL-12 Armed Oncolytic MG1 Virus In An Infected Cell Vaccine." Thesis, Université d'Ottawa / University of Ottawa, 2016. http://hdl.handle.net/10393/35599.
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Despite improvements in chemotherapy and radical surgical debulking, peritoneal carcinomatosis (PC) remains among the most common causes of death for abdominal cancers. Immunotherapies have demonstrated efficacy in selected solid malignancies but their potential in PC is poorly explored. Here I report that intraperitoneal injection of an infected cell vaccine (ICV), consisting of autologous tumor cells infected ex-vivo with an oncolytic Maraba MG1 virus expressing interleukin-12 (IL-12), promotes the migration of activated natural killer (NK) cells to the peritoneal cavity in response to the secretion of interferon gamma-induced protein-10 (IP-10) from dendritic cells. This recruitment of cytotoxic, IFNγ-secreting NK cells is associated with a dramatic reduction in tumor burden and improved survival in a colon cancer model of PC. Even in mice with bulky PC (tumors >8 mm), a complete radiological response was demonstrated within 8-14 weeks, associated with 100% long-term survival. Importantly, these results were recapitulated in human lymphocytes exposed to human tumor cell lines infected with MG1-IL12. Finally, I demonstrate that MG1-IL12-ICV generates an effective CD4 and CD8 T cell response in mice following prophylactic immunization associated with the maturation of peritoneal dendritic cells and enrichment of tumor-specific peritoneal T cells. The research presented in this thesis suggests that an MG1-IL12-ICV is a promising therapy that could provide benefit to the thousands of patients diagnosed with PC each year.
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Jilesen, Zachary Keavin. "Discovery and Application of Neoepitopes in an Oncolytic Rhabdovirus Vaccine Approach to Treat Glioblastoma Multiforme." Thesis, Université d'Ottawa / University of Ottawa, 2019. http://hdl.handle.net/10393/39688.
Full textAbstract:
Glioblastoma multiforme is the most common and lethal primary brain tumour in adults. Its aggressive and invasive phenotype makes it resistant to current standards of care, with a patient median survival following treatment of only 14 months. Potent and safe therapeutics are necessary to improve patient prognosis. Globally, efforts are being made in immunotherapies to combat such deleterious tumours. Preliminary work in the Stojdl lab has developed a novel oncolytic virus platform for brain cancer therapy that is non-toxic and exhibits potent anti-tumour efficacy. This platform is based on the rhabdovirus Farmington, identified for its potent oncolytic properties and engineering malleability. Herein, we begin to show our capability to discover and vaccinate against immunogenic neoepitopes derived from a mouse cancer mutanome. Engineering Farmington virus to express neoepitopes, allows for robust tumour specific immune proliferation following a prime vaccination. Overcoming problems of targeting self-antigen and antigen loss variants, a multi-neoepitope vaccine, presented here, is one of many alternative approaches to help combat cancer resistance. Despite achieving robust anti-tumour immunity by vaccination, selectivity of the tumour microenvironment remains an enormous challenge. Cumulative efforts in immunotherapy research will help drive novel therapeutics, like Farmington, into clinic and, ultimately, improve patient’s prognosis and quality of life.
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El-Sayes, Nader. "Small Molecule Potentiators of Oncolytic Virus Therapy Suppress the Innate Antiviral Response." Thesis, Université d'Ottawa / University of Ottawa, 2018. http://hdl.handle.net/10393/37115.
Full textAbstract:
Oncolytic Viruses (OVs) are often attenuated to increase their safety profile, however this can lead to reduced efficacy in heterogeneous malignancies and result in resistance to OV therapy. Our group utilizes small molecule enhancers of OV therapy termed viral sensitizers. These small molecules have been shown to enhance the replication and spread of oncolytic rhabdovirus VSVΔ51 in vitro and prolong survival in tumour-bearing mice. In this study, we evaluate the ef-fect of these viral sensitizers on the innate antiviral response in order to identify the mechanism of action responsible for their viral-sensitizing properties. Our previous data suggest that VSe1 and its structural analogues affect the type I IFN antiviral response and have the potential to af-fect cellular redox homeostasis. We hypothesized that VSe1 and its structural analogues potenti-ate VSV∆51 activity by inhibiting the type I IFN response via redox-mediated dysregulation. In this study, we demonstrate that the viral sensitizers inhibit the nuclear translocation and transcrip-tional activity of NFκB, which in turn dampens the expression of antiviral cytokines IFN-, TNFα and IL-6. We also provide evidence supporting the possibility that the NFκB inhibition may be a result of the formation of ROS intermediates by the viral sensitizers, which leads to re-duced nuclear translocation of NFκB subunits, thereby preventing NFκB-mediated cytokine production. Overall, this work contributes to the identification of the mechanism of action of our viral sensitizers and highlights the finding that oncolytic VSV infection can be enhanced through redox-mediated modulation of the innate antiviral response.
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Haseley, Amy M. "The Effect of the Tumor Microenvironment on Oncolytic Virus Therapy for Glioblastoma." The Ohio State University, 2012. http://rave.ohiolink.edu/etdc/view?acc_num=osu1350413344.
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Sprague, Leslee. ""Please Stand By: Investigating the bystander effects of the oncolytic virus HSV1716"." The Ohio State University, 2018. http://rave.ohiolink.edu/etdc/view?acc_num=osu1523960439218798.
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Ghobadloo, Shahrokh. "From Virus Protection to Cell Isolation and Biomarker Discovery with Aptamers." Thesis, Université d'Ottawa / University of Ottawa, 2017. http://hdl.handle.net/10393/36615.
Full textAbstract:
New affinity molecules such as nucleic acid aptamers are in demand in the science and medical fields. Current aptamer selection technologies can generate unique aptamers with desired properties to targets of interest. My thesis describes a series of investigations on the protection of an oncolytic virus, the isolation of target cells from biological fluids, and aptamer-facilitated biomarker discovery.
We tested individual aptamers and constructed a tetramer aptamer structure (quadramer) to increase virus infectivity. The quadramer protects vesicular stomatitis virus (VSV) during freeze–thaw cycles, shields the virus from neutralizing antibodies and increases viral active units. In addition to aptamers, we screened carbohydrate-based ice recrystallization inhibitors for the possible elimination of the cold chain of Vaccinia virus, VSV, and Herpes virus-1. N-octyl-gluconamide provides the longest shelf life for Vaccinia virus and Herpes virus-1 as tested according to the World Health Organization’s requirements for viral vaccines efficiency during transportation and distribution.
We generated switchable aptamers capable of isolating cells expressing LIFR, NRP1, DLL4, uPAR, or PTCH1. These aptamers bind to the receptor positive cells in the presence of Mg2+ and Ca2+, and release the pure cells upon addition of EDTA. The aptamers were applied for a sequential positive immunomagnetic isolation of cells from mice bone marrow. We also utilized fluorescence-activated cell sorting (FACS) in our aptamer selections to develop switchable aptamers to positive isolation of monocytes from human blood. Moreover, we have selected non-switchable aptamers as an affinity probe to the cells expressing Axl receptor for immunofluorescent analysis and cell sorting.
We determined aptamers to CD107a and applied them for biomarker discovery with mass spectrometry and found that CD107a was co-expressing with PD-1. Furthermore, we identified CD91 as binding partners to our aptamers in human monocytes using FACS and orbitrap mass spectrometry.
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Hardcastle, Jayson James. "Vstat120 modulates inhibits oncolytic viral therapy induced angiogenesis and innate pro-inflamatory response, augmenting oncolytic viral thereapy of glioblastom multiforme." The Ohio State University, 2011. http://rave.ohiolink.edu/etdc/view?acc_num=osu1305920551.
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