<p dir="ltr">Within the last decade, cancer immunotherapy, the therapeutic strategy of enhancing the body’s immune system to curb tumor growth, has reached the front lines in the war on cancer. Although common strategies such as adoptive cell transfer and immune checkpoint blockade have enjoyed success against some cancers, they regrettably lack durable efficacy across a broad patient population inflicted by heterogeneous and diverse cancer types. Moreover, application of these biological therapeutics is likewise limited due to various toxicities frequently encountered in the clinic. Taking these into account, the next generation of immunotherapies must exploit novel immunomodulatory targets and therapeutic strategies that can possess both enhanced efficacy compared to current options and more acceptable toxicity profiles in patients. Compared to biologics, small molecule inhibitors are desirable as they may circumvent concerns involving efficacy and toxicity, while allowing access to a broader arsenal of macromolecular targets. Recently, protein tyrosine phosphatase nonreceptor 22 (PTPN22), a key desensitization node in T cell signaling, has emerged as a systemic and translatable cancer immunotherapy target. Nonetheless, many of its precise functions in various immune cells is not fully resolved, thus there is a critical need for both novel chemical probes for biological interrogation and inhibitors with improved <i>in vivo </i>efficacy for further therapeutic development.</p><p dir="ltr">Built upon an overview of PTPN22’s structure, function, and value as an immunotherapy target, as well as a comprehensive assessment of reported inhibitors, this dissertation documents two separate medicinal chemistry campaigns on existing PTPN22 scaffolds. Herein, the structure activity relationships, design, and biological evaluation of a novel, superiorly selective and cell-active probe/ lead compound is disclosed. This dissertation also reports the design of a novel PTPN22 inhibitor with enhanced potency, selectivity, cellular efficacy, <i>in vivo </i>pharmacokinetics, and <i>in vivo </i>antitumor efficacy in mice. Our research efforts and the overall status and future directions of the field are also succinctly discussed.</p>

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The current Coronavirus disease 2019 (COVID-19) outbreak, caused by a virus called severe acute respiratory syndrome 2 (SARS-CoV-2), has become a global health emergency. Recent findings in case studies assert that the transmigration of SARS-CoV-2 to the nervous system implicates severe neurotropic pathologies, including the onset of the rare autoimmune disease called Guillain-Barré syndrome (GBS). GBS is recognized as several disorders characterized by immune-mediated polyradiculoneuropathy, which is typically preceded by an infection or other immune stimulation. The symptoms of GBS initially present as acute symmetrical ascending paresthesia, weakness, and paralysis. This meta-analysis serves to help understand the predisposing factors (such as gender, age, comorbidities) and the clinical features of COVID-19- induced GBS. Most patients affected were 40 years or older and comprised 78.2% of all the cases. Males comprised most of the cases (62.8%; n=76). The patient mortality was 9.1%, intensive care unit (ICU) admission was 46.6%, and the need for mechanical ventilation was 35.8%. It was found that concomitant GBS and COVID-19 patients most often presented with increased cerebral spinal fluid (CSF) protein levels (88%; n=106), hyporeflexia or areflexia (87.6%) (n=106), lower limb strength and sensation impairment (91.7%; n=111), upper limb strength and sensation impairment (83.5; n=101), and somatic sensation impairment (73.6%; n=89). It is postulated that COVID-19 triggers the onset of GBS through a “cytokine release storm” (CRS) that occurs in the early stages of the disease. The same cytokines and chemokines involved in this CRS caused by COVID-19 contribute to the onset of GBS. Predisposing factors which influence this concomitance include male gender and older age. Most of the reported symptoms included abnormal limb functions (including paresthesia, weakness, and paralysis) and absent or weak deep tendon reflexes. The most common variant of GBS observed was AIDP, and the most significant laboratory finding among patients was high CSF protein levels.

The COVID-19 pandemic, caused by Severe Acute Respiratory Syndrome Coronavirus 2, or SARS-CoV-2, has been ongoing for over two years. The virus spreads easily and is more unpredictable than well-known viruses like the flu, making it important to have reliable combative measures before we fully drop non-vaccine preventive actions, like mask-wearing.Therefore, we used computational protein modeling to investigate the interactions of three nonstructural proteins (abbreviated Nsp) encoded in the viral RNA genome– Nsp3, Nsp5, and Nsp6 – which are involved in the viral life cycle, with human P-type polyamine transporting ATPases ATP13A2 and ATP13A3, whose disease symptoms when mutated mimic certain COVID-19 complications. Understanding these interactions can help shed light on the mechanism of unexpected symptoms seen in COVID-19 and provide an avenue through which to treat infections. Additionally, papain-like protease (PLpro) and 3-chymotrypsin-like protease (3CLpro), which correspond to Nsp3 and Nsp5, respectively, are highly conserved between SARS-CoV and SARS-CoV-2 and thus make good potential drug targets due to their active sites and presumable lower ability to tolerate mutations (reducing the likelihood of treatments becoming ineffective), although the potential effects on the human proteasome would need to be further investigated. In addition, Nsp6 may help the virus evade host defenses by limiting the ability of autophagosomes to deliver viral particles to lysosomes, so limiting its interactions may increase the ability of the host cell to target its viral invader. One compound in particular, Haloperidol, showed promising results; predicted docking (via computational molecular docking software) to Nsp6 alone, as well as to Nsp6-heteroprotein complexes suggested strong binding, indicating a potential strong interaction that could impact the viral protein function and thus the viral life cycle.

The purpose of this study was to focus on whether solely obesity (measured by BMI) leads to an increased risk of COVID-19 mortality in terms of excess in-hospital deaths. As the grim milestone of one million deaths in the United States from COVID-19 was reached, one could assume the disease would continue to coexist with humanity in the long term. While vaccination continued to limit the spread of COVID-19, it was essential to investigate risk factors that may exacerbate the severity of the illness in humans. Obesity is already a global pandemic affecting 40% of Americans and over 650 million people worldwide. Obesity is connected to an entire range of clinical conditions including some of the leading causes of death worldwide making it a more generalizable statistic. Furthermore, the purpose of this study was to conduct a systemic review of major risk factors between obesity and COVID-19, and this analysis shall ascertain which factors have the most predictive value in determining mortality and severity of the condition. Ten research studies of 3,780,926 COVID-19 patient cases were included. Meta-analysis results indicate a pooled OR of 0.93 (0.71-1.23, p = 0.627) for in-hospital mortality of obese patients relative to non-obese patients when adjusted for confounders. All comorbidities associated with the development of severe disease were found to have an equal chance of leading to mortality. In other words, obesity did not lead to a statistically significant risk of dying from COVID-19.

I december 2019 i Wuhan, Kina, blev en grupp personer infekterade av det nya viruset SARS-CoV-2 som inducerade sjukdomen COVID-19. Sjukdomen påverkar lungfunktionen och studier har visat att långvarig påverkan på lungfunktionen kan förekomma. Syftet med den aktuella studien var att undersöka långvariga effekter av COVID-19 på diffusionskapaciteten i lungorna. Studier som undersökte diffusionskapaciteten, mätt med kolmonoxid (DLCO), hos patienter minst 30 dagar efter insjuknande i COVID-19 samlades in och analyserades. Resultatet visade att en stor del av deltagarna hade DLCO &lt;80% av det förväntade värdet. Det visade även att diffusionskapaciteten kan vara nedsatt upp till sex månader efter utskrivning från sjukhus. Då COVID-19 är en ny sjukdom var den aktuella studien begränsad av antalet studier inom området. Framtida studier bör undersöka effekterna av COVID-19 på lungfunktionen under ett längre tidsspann. Slutsatsen i den här litteraturstudien var att COVID-19 påverkar diffusionskapaciteten i lungorna. Långvariga nedsättningar i diffusionskapaciteten har, än så länge, kunnat påvisas upp till sex månader efter sjukdom. Allvarligt insjuknande i COVID-19 ökar risken för långvariga DLCO -nedsättningar.

As obligatory intracellular parasites, viruses are in a constant battle with their host to establish infection. They can facilitate their propagation by modulating host immune or metabolic pathways. This modulation involves targeting various molecular factors such as microRNAs (miRNA), enzymes, or small molecules. Understanding how viruses alter the chemical makeup of a cell is crucial to identifying what pathways are being targeted, furthering our understanding of the virus life cycle, and may aid in identifying biomarkers of disease. Here, we examine host-virus interactions in the context of two viruses, hepatitis c virus (HCV) and severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). First, the modulation of serine hydrolases by a pro-viral microRNA, miRNA-122, is investigated using activity-based protein profiling (ABPP). This study identifies a downstream target of miRNA-122 that is differentially activated during HCV infection which can be targeted pharmacologically to reduce HCV infectivity. Second, we apply similar techniques to identify serine hydrolase changes associated with SARS-CoV-2 infection. Results point towards enrichment of endocannabinoid metabolism which may offer an alternative therapeutic avenue for combating SARS-CoV-2 infection. Together, the work presented in this thesis provides avenues for further investigation into miRNA-122 interactions during HCV infection and endocannabinoid metabolism in SARS-CoV-2 infection.

Unexpected high mutations detected in new emerging variants of concern (VOCs) of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), especially in the case of omicron, raises concerns and efforts to understand their evolutionary trajectory. Several hypotheses have been discussed in literature to conceptualize the source of their emergence, including intra-host viral evolution in immunocompromised patients. These patients grant opportunities for the emergence of new variants through a persisting virus winning against host immunity, and selection for viral mutations driven by treatment interventions. VOCs have in common high mutation rate exceeding the average rate of 1-2 mutations per month. Not many studies have investigated the evolutionary rate of SARS-CoV-2 in immunocompromised candidates. Therefore, the purpose of this study is to reveal potential mechanisms underlying the emergence of VOCs by exploring substitution rate of SARS-CoV-2 genomes from surveyed COVID-19 immunocompromised patient’s studies. First, SARS-CoV-2 genome sequences were collected at sequential time series throughout host infection, which were reported in the previous studies. Filtration criteria was applied to reanalyze patients with prolonged infection documented for ≥ 2 months, and comprehensive sequenced samples for ≥ 6 time points. Then, phylogenetic analysis was conducted using Nextclade (https://clades.nextstrain.org/), followed by mutation rate analysis using two substantial similar approaches to calculate the rate in i) substitutions per month and ii) substitutions per site (per year). The mutation tendency of SARS-CoV-2 in immunocompromised hosts was compared to reported VOCs, particularly to omicron. The highest observed mutation rate accounted for approximately 2.2 mutations per month, which is higher than the average rate. High mutation rate was due to prolonged infection and selection pressure by treatment interventions (i.e., convalescent plasma and antibodies). Here, higher rate of intra-host viral evolution in immunocompromised patients is detected, potentially leading to the emergence of VOC. Hence, this research highlights the need for sequencing efforts in high-risk individuals, updating treatment strategies along with further analysis on adaptive mutants pronounced due to intra-host evolution. Together, such findings provide an ultimate synergy for future public health guidelines and infection control measures.

<p>  </p> <p>Seven-membered ring structures are one of the most important structural motifs found widely in natural products and bioactive molecules. Although several [5+2]-cycloaddition reactions have been developed to construct these seven membered cores, asymmetric cycloaddition reactions are less explored. Described in Chapter-1 is a base mediated intramolecular diastereoselective [5+2]-cycloaddition reaction that afforded highly functionalized seven membered rings in good yields and excellent diastereoselectivities. The high diastereoselectivity is controlled by the alkyl stereocenter and the chain length of the alkene tether. The existing chirality of the substrate can direct the stereochemical outcome of the [5+2]-cycloaddition reaction. Furthermore, this methodology has been applied to synthesize various potent HIV-1 protease inhibitors. </p> <p>COVID-19 pandemic has profoundly affected life around the globe and costed us 6 million lives. Therefore, there is an urgent need for rational design of new drug candidates to specifically target different SARS-CoV-2 proteins. Recently, Pfizer developed an FDA-approved antiviral therapeutic agent, Paxlovid, targeting SARS-CoV-2 3CLpro. Chapter-2 discusses a concise synthetic route to synthesize active component of Paxlovid, Nirmatrelvir, in 6-steps without any epimerization. We have also developed a series of potent covalent inhibitors targeting SARS-CoV-2 3CLprotease and compared the antiviral activities of these inhibitors with that of Nirmatrelvir. </p> <p>In the final chapter, a concise partial synthesis of the segment A of (-)-Rafonin is discussed. (-)-Rasfonin is an antitumor agent that induces apoptosis in <em>ras</em>-dependent cells. We have proposed an asymmetric chiron approach to install the α-pyranone ring of rasfonin. Our goal is to perform SAR studies on this natural product by designing various analogues. </p>

All cells depend on properly folded proteins for survival and function. Misfolding of proteins results in loss of critical functions and may trigger the misfolding of other nearby proteins leading to toxic aggregation. While many proteins can fold on their own, others with complicated domain structures require assistance from protein folding machines called chaperones. The most complex and highly specialized of all chaperones is the eukaryotic chaperonin complex CCT which is necessary for the folding of a wide variety of essential proteins. These include the cytoskeletal proteins actin and tubulin as well as the Gβ subunit of the G protein heterotrimer. However, CCT activity can also drive diseases such as cancer and viral infections and could represent a high value therapeutic target if the mechanisms by which it folds its different client proteins were better understood. To this end, we identified and characterized an interaction between CCT and the RNA-dependent RNA polymerase of SARS-CoV-2, the virus responsible for the deadly global pandemic that began in 2019. We showed that SARS-CoV-2 replication is impaired by loss of CCT and that the polymerase, designated Nsp12, interacts with CCT upon synthesis and quickly releases - the hallmark pattern of a CCT substrate. Furthermore, we solved a 3.3 Å cryo-EM structure of Nsp12 bound to open CCT showing Nsp12 binding between the two rings of CCT and extending up through of the folding chambers and out of CCT. At 107 kD, Nsp12 is the largest substrate ever visualized inside of CCT and answers a long-standing question in the field of how CCT could accommodate substrates larger than its 70 kD folding chamber. Given that CCT is known to fold proteins with WD40-repeat domains, we also investigated a potential relationship between CCT and RPE65. RPE65, which contains a WD40 domain, is the retinyl ester isomerase that converts all-trans retinyl esters into 11-cis retinol, a key step in the visual cycle, and its mutation is a common cause of hereditary retinal dystrophies. We showed that CCT interacts with RPE65 and that nascent RPE65 binds and releases from CCT albeit with relatively slow kinetics. However, RPE65 is not dependent on CCT for expression or activity. This suggests that the relationship between RPE65 and CCT may represent a novel CCT function distinct from the canonical obligate substrate dynamic.