Research

NPA: Dr. David Marchant

We are designing and synthesizing an antiviral product called RespVirex that can be used to treat acute respiratory viral infections. Our target consumers are high-risk populations such as seniors and the immunocompromised can greatly benefit from a small molecule, non-toxic therapeutic option. This compound is unique as it binds to a new druggable pocket we have discovered through our computational investigations of RNA-dependent RNA polymerases (RdRp) of several respiratory viruses, including RSV and SARS-CoV-2. Our ultimate goal with this work is to further develop our hit broad spectrum, non-toxic, non-nucleoside antiviral drug to be used as a single therapeutic option or in combination with existing approved antivirals as adjunct/combination therapy.

NPA: Dr. James Nieman

Although vaccines and treatments are available for COVID-19, the virus (SARS-CoV-2) continues to mutate and better antivirals are needed for future variants and outbreaks of other coronavirus. Current antivirals all suffer from serious shortcomings and are not ideal for combination therapy. We have discovery compounds, such AVI-8106, that have shown impressive activity, improved metabolic stability and reasonable pharmacokinetics (PK). Further exploration of the structure to optimize the properties (such as improved permeability, solubility, metabolic stability and volume of distribution to improve PK) will provide compounds that have high bioavailability, longer half-life and thus the prospects of a reduced pill burden and no need for a PK booster, such as ritonavir. If successful, further improvement of properties will generate a compound that achieves the target product profile to move towards testing as a second generation protease drug to treat COVID-19.

NPA: Dr. Tom Hobman

The endemic pathogen Respiratory Syncytial Virus (RSV) poses an enormous burden on healthcare systems. Moreover, targeted antiviral options are limited and therefore, effective therapeutics against this pathogen are urgently needed. To this end, we recently discovered that pharmacological upregulation of peroxisomes, metabolic organelles that also function in the interferon (IFN) response, have broad-spectrum antiviral activity against pathogenic RNA viruses. Here, we will test a panel of 40-50 drugs that upregulate peroxisomes (via different mechanisms) for antiviral activity against RSV. In addition, we will create reporter cell lines that can be used to screen for other compounds that induce peroxisomes. These hits will then be tested against RSV, influenza virus, flavi- and alphaviruses. At the end of this project, we expect to identify 5 or more compounds that can be tested against RSV and other endemic RNA viruses in small animal models.

In Alberta, >8,000 people are hospitalized each year because of RSV. There is but one licensed drug (Palivizumab) for RSV, an expensive antibody-based therapeutic that can only be used preventatively. As such, additional antiviral therapies are urgently needed to mitigate RSV infection. In the present proposal, we propose to screen a panel of IFN-inducing drugs for antiviral activity against RSV. As IFN treatment has been shown to reduce severity of RSV infection, we expect that pharmacologically enhancing the IFN response will be an effective strategy for mitigating the effects of RSV infection.

NPA: Dr. Joanne Lemieux

Coronavirus’ RNA genome encodes for two polyproteins that are autocatalytically processed by viral proteases – PLpro and Mpro into 16 non-structural proteins (nsp1–16). Mpro cleaves two polypeptides at 11 sites to produce 13 mature proteins, and is therefore indispensable for virus replication (ref). Since Mpro is also very well conserved it is considered an attractive drug target. Nimatrelvir (PaxlovidTM), developed by Pfizer, a peptidomimetic inhibitor of Mpro, was approved by FDA and released on the market to treat SARS-CoV-2 in December 2021. However, nirmatrelvir suffers from rapid oxidative metabolism and requires co-dosing with ritonavir as part of PaxlovidTM3. We are developing new generation of oral antivirals which initially demonstrated some broad specificity against main strains of coronaviruses.

We aim to identify broadly specific inhibitors and concurrently develop a commercialized multiplex assay, which will include isolated Mpro enzymes from HCoV-229E, -OC43, -NL63, and -HKU1 strains, and MERS, SARS-CoV and SARS-CoV-2 for compound screening. The ability of screening inhibitors against multiple analytes in high-throughput manner will extremely benefit the program in terms of decreasing the cost and time towards our identification of an antiviral inhibitor with broad specificity

NPA: Dr. John Vederas

The overall goal is to construct more metabolically robust inhibitors of the main protease (3CL) of SARS-CoV-2 that causes COVID-19. These are based on the structures of nirmatrelvir (protease inhibitor in Paxlovid™) and of GC376 that cures feline infectious peritonitis (FIP) coronavirus in cats. Such compounds will also be tested with the main protease (3C) of human rhinovirus (a picornavirus) as its active site, substrate and mechanism are have some similarity. Successful analogs will provide leads for single agent antiviral drugs to treat acute COVID-19 and potentially other respiratory viruses.

GC376 is an injectable drug shown to cure usually fatal FIP coronaviral infections in cats. It is structurally related to nirmatrelvir and potently inhibits the SARS2 3CL protease and viral replication. Currently, it is not orally bioavailable and is administered to animals by injection. Specific objectives of this project include:

1) Chemical synthesis of analogs of nirmatrelvir and GC376 with sites of oxidation by CYP3A4 having hydrogen substituted by deuterium to slow metabolism by isotope effect;

2) Synthesis of SARS2 3CL protease inhibitors with the cyclic glutamine analog having methylene (CH2) next to nitrogen replaced by oxygen to block oxidation;

3) Testing of all analogs for rate of oxidative transformation by CYP3A4 and for inhibition of SARS2 3CL protease, FIP coronaviral 3CL protease and human rhinovirus 3C protease.

NPA: Dr. Stephanie Yanow

Over 13 million pregnant women are exposed to malaria annually and there is currently no vaccine available to protect this vulnerable group. During infection in pregnancy, parasites accumulate in the placenta, leading to harmful outcomes for both the mother and the fetus. Our goal is to develop a vaccine that targets the protein used by the parasite to bind in the placenta. In this project, we will produce specific segments of this protein as synthetic molecules and encapsulate them into particles that will be released during vaccination. We will focus on two synthetic molecules and compare different strategies to encapsulate them and stimulate immune responses in mice. We anticipate that these vaccines will produce strong antibodies that will prevent the binding of parasites to the placenta. We expect to identify one vaccine formulation that shows promise and can progress toward human clinical trials.

NPA: Dr. Michael Meanwell

The continued discovery and development of small molecule pharmaceuticals is critical to treating patients and managing infection rates during the next pandemic. The COVID-19 pandemic highlighted the importance of having libraries of broad-spectrum antivirals readily available for antiviral screening. These libraries enable scientists to work quickly in developing urgently needed medicines against rapidly evolving viral diseases. Nucleoside analogues represent the largest class of antivirals on the market today, however, despite their success, there remains several long-standing challenges associated with nucleoside analogue syntheses that if addressed would inspire new opportunities in antiviral drug development. This project aims to develop a new process for making nucleoside analogues that serves to address many of these challenges – this will provide access to novel molecules that have otherwise been inaccessible and thus are unexplored as antivirals. We will then utilize this process for making a library of novel nucleoside analogues (~ 100) and from there proceed to evaluate these molecules as potential antiviral drugs. Given the success of nucleoside antivirals to date, we believe that further innovations in this area should lead to important discoveries in the near future for improving human health.

NPA: Dr. Holger Wille

Some neurodegenerative diseases are caused by the misfolding of specific, regular proteins that are found in healthy brain cells and tissues. This misfolding turns the normal proteins into disease-causing forms of the same proteins. We developed a new technique to create vaccine candidates that specifically target only the disease-causing proteins without affecting the regular, healthy forms of the proteins. In our current project we want to develop improved versions for our vaccine candidates that target Parkinson’s disease and the prion diseases. We will engineer improved vaccine candidates, produce them in bacteria, purify the resulting antigens, conduct multiple quality control experiments, and test the resulting immune responses in vaccinated rodent models. If the newly improved vaccine candidates satisfy these quality control criteria, we will prepare to test the prophylactic effectiveness of the vaccine candidates in animal models of Parkinson’s disease and the prion diseases. However, these effectiveness tests will take several years and are therefore not part of the current project.

NPA: Dr. Troy Baldwin

A key element of a strong public health system is the availability of vaccines that stop people from getting sick or dying. As highlighted by the recent pandemic, vaccination reduced hospitalizations and deaths due to COVID-19 and sped up the return to a more normal way of life. Developing improved ways to generate vaccines and understanding the strengths and limitation of new vaccines is critical to prepare us for a future pandemic. We will study the capability of a new vaccine platform we recently created and determine whether it can be used as a universal vaccine platform.

NPA: Dr. Vanessa Meier-Stephenson

Influenza viruses remain one of the leading causes of worldwide pandemics and one of the several respiratory viruses deemed to have a high likelihood of precipitating the next global pandemic. While vaccination will certainly have a strong role in pandemic preparedness and control, so too will the development of directly targeting therapies that can be taken at the first sign of infection. Given influenza’s diverse host reservoirs (i.e., birds and swine), a therapeutics approach will need to derive from a strategy that is both robust and adaptable. One protein complex that is common to all influenza viruses is its multipurpose polymerase. This protein is the virus’ Swiss army knife and plays many roles in its replication process from beginning to end. Interfering with the ‘knife’s’ functioning at even one of its steps could have an important impact on the virus’ ability to replicate. We propose the use of a diverse molecular probe strategy to screen the nooks and crannies of influenza’s polymerase to determine the best way to jam its pocketknife. This process uses a library of millions of molecular probes and sequentially works to find a shortlist of ones that can bind the strongest. We can then use that shortlist of probes to determine how and where they have bound and whether any modifications will be needed before testing in different culture models. This molecular probe approach is not only a valuable tool against influenza virus, but for other potential emerging pathogens, giving us the ability to pivot to whichever key target is of greatest interest.

Developing this platform on-site will have significant implications for turn-around time to probing function and acquiring first drug options.

NPA: Dr. John Klassen

The goal of this project is to develop new treatments for a wide range of viral infections, including COVID19 and the flu. We will do this by creating molecules that make it harder for viruses to enter our cells. Many human viruses use sugars (glycans) on the surface of our cells to help them gain entry. We will make drug molecules that disrupt the production of the sugars that viruses use for cell entry.  We will use advanced analytical methods to identify the molecules that bind strongly and selectively to specific enzymes that produce a specific class of cell surface sugars. We will then test the ability of these molecules to block cell infection by SARS-CoV-2, the virus responsible for COVID19, to demonstrate proof-of-concept.

NPA: Dr. Sue Tsai

Obesity is a leading risk factor for severe respiratory infections (e.g., H1N1 and SARS-CoV2), yet the same population of individuals is also afflicted with increased risks of vaccine failure. Evidence shows that excessive inflammation leads to disrupted insulin signaling and metabolic homeostasis, and our own work indicates that insulin resistance intrinsic to adaptive immune cells impairs immunological memory. Here, we propose a proof-of-concept study, where we will test the effectiveness of administering anti-diabetes agents that ameliorate systemic metabolic homeostasis and insulin sensitivity, in boosting immunological memory. We will carry out a series of experiments where preclinical models of diet induced obesity will be subjected to insulin sensitizing therapies involving adiponectin receptor agonism, followed by survival and immune function analysis. We will then investigate the specific effects of the therapies on memory B cells and T cells via multi-omics approaches, to gain insights into the metabolic pathways that can be leveraged to perpetuate long lasting protection.

NPA: Dr. Wael Elhenawy

The antimicrobial resistance (AMR) crisis has reached alarming levels worldwide. Our arsenal of antibiotics is failing in the face of bacterial infections due to the rise in AMR and the simultaneous decline in drug discovery. While the COVID-19 pandemic is viral, it was associated with a surge in AMR due to secondary infections with multi-drug resistant bacteria. This poses a threat on vulnerable patients in hospitals and nursing homes in Alberta and the rest of the world. Therefore, there is a pressing need to develop new antimicrobials to face the burgeoning AMR crisis. In the proposed work, we combined computational and experimental approaches to expedite the discovery of antimicrobials at a low cost. We developed a screening pipeline that identified new anti bacterial agents.  We were able to show that these drugs have a broad spectrum against several pathogens like Escherichia coli, Salmonella enterica, and Vibrio  cholera. Future work includes testing the efficacy of these drugs in our established infection models. Together, our program provides a framework for the preclinical development of antimicrobial therapeutics.  

NPA: Dr. Matthew Macauley

Vaccines are one of the greatest feats in human history and have been the solution to many medical crises, including the recent SARS-CoV-2 pandemic. In the past, vaccines using inactive versions of the virus have been widely successful. While this has proven effective, it is challenging to make inactivated viral vaccines on scales large enough for the public during a pandemic. Moreover, vaccine quality control on large scales is another challenge. Modern vaccines use smaller pieces of the virus rather than the entire inactivated virus to generate an immune response. These vaccines that use a piece of the virus, rather than the whole virus, are referred to as subunit vaccines. The major advantage of subunit vaccines is their modular construction, enabling rapid scaling and greater quality control. Indeed, it is these advantages that led to lipid nanoparticles (LNPs) being the vaccine of choice during the COVID-19 pandemic. However, a huge open question is whether these types of vaccine elicit the long-term protection characterized by inactivated viruses. A fundamental difference between whole viruses (inactivated or otherwise) and LNPs, is how they initiate immune responses. In this project, we will develop liposomal nanoparticles that exploit the natural features of how viruses first stimulate immune response, to examine if this can produce a more robust immune response. In summary, will marry the advantages of subunit vaccines (modularity and scalability) with how our immune system captures and responds to viruses naturally.

NPA: Dr. David Evans

The CODEX BioXpTM 3250 gene synthesizer offers stand-alone technology that, among other features, can be used to construct duplex DNAs up to 7.2 kbp in length in <16 hr. The instrument is a modified robot which is preloaded with oligonucleotide fragments and other custom reagents (e.g., enzymes, buffers) ordered from CODEX.
 
Some important features include:

• Assemble genes, genomes, clones, DNA variant libraries, and mRNA using the same platform
• Generate up to 32 different DNA fragments in <24 hr
• Generate up to 10 μg of DNA from cloned de novo synthesized fragments
• Synthesize up to 10 μg of mRNA per reaction in ~20 hours, option to include modified nucleosides - Clone into off-the-shelf or user-supplied custom vectors
• Low error rates (1:10,000 to 1:30,000)
• Built-in thermocycler and chiller

The primary intent of the gene synthesizer is to ensure that SPP-ARC researchers can acquire the recombinant DNAs needed for research requiring novel antigens, enzymes, and other proteins. Most importantly, the BioXP system ensures that researchers can rapidly respond to new demands in time-sensitive circumstances. Emerging pathogens represent exactly the kind of challenge requiring a rapid response.

NPA: Dr. Olivier Julien

We are acquiring an infrared imager with chemiluminescence that will be used to identify and validate interactions between viral and host proteins, such as SARS-CoV-2 and human cells. With high precision and sensitivity, the Odyssey M Imager is everything we need to quantify viral and host proteins on the same blot. The loading control can be a single host or human protein, or the new gold standard for normalization, full protein stain quantification, leaving the other channels free for viral and host protein identification and quantification. With the highest sensitivity and multiplexing capabilities currently available, the Li-COR Odyssey offers the best solution to quantify multiple proteins within the same sample. With such accuracy we will be able to provide robust data for our current projects on SARS-CoV-2, Mayaro, Chikungunya, and monkeypox viruses. It will be used extensively for all our current and future projects.

NPA: Dr. Matthew Macauley

Preparing for the next pandemic means having the infrastructure in place to development, test, and manufacture vaccines. Developing and testing new vaccines requires innovative approaches but also a firm understanding of the materials being used. In most cases, these materials are in the nanometer range. For examples, some of the most successful long-lasting vaccines are attenuated viruses, which members of the SPPARC team are aiming at advancing (e.g. Reovirus and Vaccinia virus). Newer generations of vaccines use nanoparticles loaded with mRNA. In both cases, particles are typically in a 50-200 nm range. Such particles require careful characterization to measure their size and heterogeneity (uniformity). While there are instruments that can measure the average size of particles in solution, there is only one reliable instrument, the Malvern Nanosight Pro, that allows for both parameters to be easily quantified. Quantifying heterogeneity is particularly important because it addresses the quality of the preparation - specifically the lack of clumping. Indeed, aggregates can give adverse effects in vivo and is a parameter that is important to understand. Accordingly, this new infrastructure will enable all members of

NPA: Dr. Howard Young

Cryogenic electron microscopy (CryoEM) is an advanced imaging technique used to determine a three-dimensional picture of biological molecules and complexes at near-atomic resolution. In this technique, the sample is rapidly frozen at very low temperatures (below -180°C), trapping it in vitreous ice (a glass-like, frozen state of water). The sample is then imaged from different angles using an electron microscope, where a beam of electrons passes through the sample. The recorded images of the sample can be used to determine atomic-level pictures of biological molecules and complexes. The requested equipment will aid in the preparation of frozen samples for cryoEM.