Dissertations / Theses on the topic 'Antivirals'
To see the other types of publications on this topic, follow the link: Antivirals.
Create a spot-on reference in APA, MLA, Chicago, Harvard, and other styles
Consult the top 50 dissertations / theses for your research on the topic 'Antivirals.'
Next to every source in the list of references, there is an 'Add to bibliography' button. Press on it, and we will generate automatically the bibliographic reference to the chosen work in the citation style you need: APA, MLA, Harvard, Chicago, Vancouver, etc.
You can also download the full text of the academic publication as pdf and read online its abstract whenever available in the metadata.
Browse dissertations / theses on a wide variety of disciplines and organise your bibliography correctly.
1
Chen, Qian. "Caracterización molecular del perfil de resistencias del virus de la hepatitis C después del fallo terapéutico a antivirales de acción directa mediante secuenciación masiva." Doctoral thesis, Universitat Autònoma de Barcelona, 2018. http://hdl.handle.net/10803/666656.
Full textAbstract:
La infección crónica por el virus de la hepatitis C constituye un problema de salud pública a nivel mundial por su contribución al desarrollo de enfermedades hepáticas avanzadas y carcinoma hepatocelular. Actualmente, gracias a la disponibilidad de terapias antivirales altamente eficaces y bien toleradas basadas en combinaciones de antivirales de acción directa (DAAs) se han logrado tasas de respuesta virológica sostenida (RVS) en el 95% de los pacientes. A pesar de la excelente eficacia de los DAAs, sigue habiendo un porcentaje considerable de pacientes que no alcanzan la curación virológica. Tras el fallo terapéutico, es frecuente observar la selección de sustituciones asociadas a resistencia (RASs) a frecuencias altas en la población viral. Mientras que la selección de RASs tiene un papel importante en el fracaso terapéutico, el impacto clínico de las RASs, así como su relevancia en el retratamiento sigue siendo una cuestión abierta. Hoy en día, se disponen de pocos estudios en vida real sobre RASs, estando la mayoría basados en secuenciación Sanger. En la presente Tesis Doctoral, se ha realizado un estudio de resistencias de una cohorte de 220 pacientes con fracaso terapéutico a combinaciones de DAAs mediante secuenciación masiva. En los resultados obtenidos, se observaron patrones de resistencia específicos de subtipo viral tras el fallo a combinaciones de DAAs, lo cual realza la importancia del uso de cebadores específicos de subtipo para evitar sesgos en la amplificación. Se caracterizaron varias combinaciones de RASs altamente prevalentes, sugiriendo la importancia de su detección antes del retratamiento por los elevados niveles de resistencia que confieren. Además, considerando la prevalencia de las RASs en las regiones que mapean fuera de las regiones frente a las que van dirigidos los antivirales de acción directa, es recomendable analizar todas las regiones terapéuticas para decidir el tratamiento de rescate. En resumen, la alta prevalencia de RASs tras el fallo terapéutico, la gran cantidad de RASs minoritarias y las combinaciones de RASs en el mismo genoma, reafirman la importancia de su análisis antes del retratamiento usando secuenciación masiva ultra profunda para maximizar las tasas de RVS. Sería de gran interés conocer los resultados de aquellos pacientes sometidos a retratamiento para caracterizar las RASs clínicamenteChronic hepatitis C infection is considered as a major public health issue worldwide due to its linkage to the development of advanced liver diseases and hepatocellular carcinoma. Currently, the availability of highly efficient and well-tolerated antiviral therapies based on combinations of direct acting antivirals (DAAs) has provided sustained virological response (SVR) in nearly 95% of patients. Despite the excellent efficacy of DAAs, still a non-negligible percentage of patients do not achieve virological cure. At treatment failure, resistance-associated substitutions (RASs) are usually selected at high frequencies in the viral population. While selection of RASs has an important role in treatment failure, the clinical impact of RASs and its relevance in retreatment still remain unknown. Few real-life data on RASs testing are available, mainly performed by Sanger sequencing. In this PhD Thesis, we have performed a RASs analysis in a cohort of 220 patients who experienced treatment failure to several DAA combinations using next generation sequencing. In our analysis, the RASs profile that emerge after each DAA-based treatment was subtype-specific, which strongly suggests the use of subtype-specific primers to avoid amplification bias. Also, several high prevalent RASs combinations were characterized, suggesting the importance of their detection before retreatment due to their high level of resistance. Moreover, attending to the high occurrence of extra-target RASs detected, testing all genomic regions for RASs analysis is strongly recommended for treatment decision making. In summary, the high prevalence of RASs at treatment failure, the high amount of minority RASs and the combination of RASs at the same genome, reinforce the importance of RASs analysis before retreatment using ultra-deep sequencing in order to maximize SVR. The outcome of patients who undergo retreatment should be also analysed in order to characterize clinically meaningful RASs.
2
BASILE, TERESA. "Pericyclic Reaction for Antivirals." Doctoral thesis, Università degli studi di Pavia, 2020. http://hdl.handle.net/11571/1318326.
Full textAbstract:
Questa tesi di dottorato ha lo scopo di sintetizzare nuovi nucleosidi carbonilici sfruttando la chimica degli intermedi nitrosocarbonilici e dei nitrilossidi.
La tesi può essere suddivisa in tre argomenti principali:
a) I primi studi sono stati condotti sugli analoghi nucleosidici del 4-idrossi-2-ciclopentenone, uno scaffold presente in numerosi prodotti naturali e biologicamente attivi. Questo può essere facilmente funzionalizzato utilizzando opportuni gruppi uscenti al fine di realizzare una sostituzione nucleofila per linserimento di diverse eterobasi.
b) La seconda parte è dedicata alla generazione di intermedi nitrosocarbonilici usando un approccio fotochimico. Lo starting material, un acido idrossamico o un suo sale, viene ossidato per mezzo del TBADT. Questi intermedi nitrosocarbonilici possono essere poi intercettati da opportuni dieni in una cicloaddizione [4 + 2] nota come nitroso-Diels-Alder (NDA) per generare una varietà di cicloaddotti.
c) Infine, lo studio e la sintesi di diversi acidi idrossamici per la generazione degli intermedi nitrosocarbonili ci ha condotti ad una nuova sintesi step-economy del Panobinostat. Il Panobinostat o commercialmente chiamato Farydak, è un acido idrossamico complesso con importanti proprietà biologiche e chemioterapiche.The synthesis of new nucleoside analogues is a new approach in viral chemotherapy showing a remarkable activity towards different types of viruses. The aim of this work is to synthetize new carbocyclic nucleosides by taking advantage of the chemistry of nitrosocarbonyl intermediates and nitrile oxides. The thesis can be divided in three main topics: a) First of all, were conducted synthetic studies of nucleoside analogues of racemic 4-hydroxy-2-cyclopentenone, a scaffold often found in natural products and biologically active compounds. This core can be easily functionalized with suitable leaving groups to perform nucleophilic substitution reaction or metal-catalyzed synthesis to access nucleoside analogues by insertion of several heterobases. The reaction can be also evaluated in its optically pure version. Docking studies guided the choice of the functional groups in order to increase or modify properly the biological activity. Apoptotic activities were evaluated for the most promising compounds. b) In this part, nitrosocarbonyls were generated using a photochemical reaction method. The starting material, an hydroxamic acid or its corresponding salt, was tested in oxidant room of TBADT. The tetrabutylammonium decatungstate (TBADT) is an efficient and robust photocatalyst able to promote photoredox reactions, as well as hydrogen atom transfer processes, starting from different classes of organic substrates. The [4+2] cycloaddition of dienes with nitrosocompounds, namely the nitroso-Diels-Alder (NDA) reaction, is a versatile method to generate highly reactive acylnitroso species from hydroxamic acid derivatives. Since nitrosocarbonyl intermediates participate in a variety of organic reactions, the in situ formation of this highly reactive species using photoredox conditions furnished a general procedure for patterning surfaces bearing a range of properties. c) Finally, we proposed a new synthesis of Panobinostat, a complex hydroxamic acid with important biological properties. Specifically, it is an orally administered drug for the treatment of patients with multiple myeloma. In this context, for its relevant pharmacological role, is essential to identify new step-economy and waste-economy approach to access this important compound.
3
Jayawardena, Shanthi. "Control of influenza detection and antivirals /." Thesis, Click to view the E-thesis via HKUTO, 2007. http://sunzi.lib.hku.hk/hkuto/record/B40039742.
Full text
4
Fleta, Soriano Eric 1983. "Broad-spectrum host-acting antivirals: identification and characterization of anti-HIV drugs." Doctoral thesis, Universitat Pompeu Fabra, 2015. http://hdl.handle.net/10803/402212.
Full textAbstract:
Hundreds of host factors related to viral infections like HIV, hepatitis C virus, dengue virus or West Nile virus have been identified. As many of these host factors are shared by different viruses, chemical blockade of key virus-associated cellular components may effectively act as broad-spectrum antiviral treatment. Broad-spectrum host-acting antivirals (HAAs) may reduce treatment complexity and costs, increase adherence to the therapy and may pose a higher barrier to develop resistance. In this thesis a high-throughput anti-HIV assay was used to screen for virus inhibitory effects of a library of secondary metabolites derived from myxobacteria. Compounds with high anti-HIV activity and low toxicity were classified as hits and two of them (ratjadone A and soraphen A) were selected for further analysis. The mechanism of HIV inhibition of both compounds is described here. The results presented in this thesis show that broad-spectrum HAAs are a feasible option for antiviral treatment and that the compounds identified can be further studied for hit-to-lead compound development.Cientos de factores del huésped relacionados con infecciones virales por VIH, hepatitis C, dengue o virus del Nilo occidental han sido identificados. Como muchos de esos factores del huésped son compartidos por diferentes virus, el bloqueo químico de un componente celular clave asociado al virus podría actuar de forma efectiva como un tratamiento antiviral de amplio espectro. Antivirales de amplio espectro contra factores del huésped podrían reducir la complejidad y el coste del tratamiento, incrementar el cumplimiento de la terapia y pueden suponer una barrera mayor al desarrollo de resistencia. En esta tesis un cribado de alta capacidad anti-VIH fue aplicado a una librería de metabolitos secundarios de myxobacteria. Compuestos con alta actividad anti-VIH y baja toxicidad fueron clasificados como hits y dos de ellos (ratjadone A y soraphen A) fueron seleccionados para posteriores estudios. El mecanismo de inhibition de VIH de ambos compuestos es descrito aquí. Los resultados presentados en esta tesis muestran que usar antivirales de amplio espectro contra factores del huésped es un opción viable para tratamientos antivirales y que los compuestos identificados pueden ser estudiados para el desarrollo de fármacos.
5
Meister, Gabriel T. "Antiviral mechanism(s) of the experimental immunosuppressive agent leflunomide against human cytomegalovirus and polyomavirus." Connect to this title online, 2005. http://rave.ohiolink.edu/etdc/view?acc%5Fnum=osu1111428519.
Full textAbstract:
Thesis (Ph. D.)--Ohio State University, 2005.Title from first page of PDF file. Document formatted into pages; contains xiii, 127 p.; also includes graphics (some col.) Includes bibliographical references (p. 113-127). Available online via OhioLINK's ETD Center
6
Hussain, S. "Iminosugars as antivirals against human influenza A viruses." Thesis, University College London (University of London), 2013. http://discovery.ucl.ac.uk/1383772/.
Full textAbstract:
Influenza A viruses cause seasonal epidemics and pandemics in the human population, resulting in significant morbidity and mortality. Influenza can be controlled by vaccination and antiviral therapy. However, antigenic drift, reducing vaccine effectiveness, and the development of antiviral resistance can result in reduced efficacy of the control measures. Drugs that target host cell processes, such as glycosylation, may be employed to complement drugs that target the virus, and iminosugar compounds which inhibit α-glucosidases have been reported to show antiviral activity against some viruses. Here, I have examined the effect of two iminosugars on human influenza A viruses. I have shown that two α-glucosidase inhibitors, N-butyl deoxynojirimycin (NB-DNJ) and N-nonyl deoxynojirimycin (NN-DNJ), show antiviral activity in cell culture against three human influenza A viruses: a recently circulating seasonal H3N2 virus strain, A/Brisbane/10/2007, an older H3N2 strain, A/Udorn/307/72, and a representative of the currently circulating pandemic H1N1 virus, A/Lviv/N6/2009. Of the two, NN-DNJ was the more potent drug. The virus target and mode of action of NN-DNJ has been examined. The effect of the drug was most marked after infection. Consistent with its action as an α-glucosidase inhibitor, NN-DNJ treatment resulted in altered glycan processing, as shown by a reduction in electrophoretic mobility of both influenza virus glycoproteins, haemagglutinin (HA) and neuraminidase (NA). NN-DNJ treatment was found to reduce cell surface expression of H3 HA. The level of sialidase activity of NA was reduced in the infected cell, however addition of exogenous sialidase to cells did not complement NN-DNJ mediated inhibition of virus replication. Using reassortant viruses, the drug susceptibility profile correlated with the HA. Reverse genetics was used to determine the effect of altering the glycosylation status of the HA; engineered viruses carrying modified sites seemed slightly more sensitive to the inhibitor than the parent virus. These results show that NN-DNJ inhibits influenza A virus replication in a strain-specific manner which is dependent on the HA.
7
Hadpech, Sudarat. "Nouveaux agents antiviraux dérivés de protéines cellulaires à motifs répétés et ciblant l’assemblage du VIH." Thesis, Lyon, 2017. http://www.theses.fr/2017LYSE1139/document.
Full textAbstract:
Au cours de notre programme de thèse, nous avons isolé et caractérisé des molécules protéiques à activité antivirale intracellulaire dirigée contre le VIH-1. Ces protéines, appelées aRep, ont été obtenues par criblage d'une banque de protéines artificielles de nouvelle génération, construites de façon combinatoire à partir de protéines naturelles constituées de motifs alpha-hélicoidaux répétés. La cible virale, ou "appât", utilisé pour ce criblage a été une région de la polyprotéine Gag du VIH-1 identifiée comme une cible privilégiée de nouvelles thérapeutiques antivirales, car essentielle à l'assemblage viral, l'empaquetage du génome viral et le clivage de maturation de Gag aboutissant à la formation de virions infectieux. Deux molécules d'aRep à affinité élevée pour la cible virale, l'aRep4E3 (32 kDa; 7 motifs répétés) et l'aRep9A8 (28 kDa; 6 motifs répétés) ont ainsi été identifiées, isolées et clonées. L'étude de l'activité anti-VIH intracellulaire de ces aRep a été réalisée dans différents systèmes d'expression cellulaire, nécessitant la construction de lignées stables de cellules d'insecte et de cellules épithéliales humaines, et leur infection par différents types de vecteurs viraux recombinants, baculovirus ou lentivirus, porteurs du gène rapporteur luciférase. Mais surtout, cette étude a été menée sur des cellules lymphocytaires-T (SupT1), cibles naturelles du virus, exprimant ces aRep et infectées par du VIH-1 naturel infectieux. Nos résultats ont montré que l'aRep4E3 et l'aRep9A8 ont toutes deux un effet négatif significatif sur le cycle réplicatif du VIH-1, mais ciblent des fonctions virales différentes. L'aRep4E3 bloque l'empaquetage du génome viral, tandis que l'aRep9A8 inhibe la maturation et diminue l'infectivité virale. De plus, l'aRep9A8, exprimée de façon constitutive dans les cellules SupT1, leur confère une résistance au VIH: une lignée de SupT1 chroniquement infectée par le VIH-1 a pu être ainsi isolée et maintenue en culture pendant plusieurs semaines, sans effet cytopathique viro-induit apparent. Ces nouvelles données auront des implications non-négligeables dans le choix et la conduite de futures stratégies de thérapie cellulaire anti-VIHHIV-1 infection is a long-term disease which required a long-life treatment. Besides the standard HAART regiment, HIV gene therapy is a promising alternative strategy which give rise to hope for the better HIV-1 treatment. Protein therapeutics is one another technique that represent high impact results in curing various types of disease. It is already become a significant part of current medical treatments. In this study we first designed aRep, a non-immunoglobulin scaffold protein which target two domains of HIV-1 Pr55Gag, SP1-NC and investigated their roles as an intracellular therapeutic agents. Phage display technology was used for the specific isolation of aRep against a critical C-terminal region of the HIV-1 Pr55Gag precursor from a large and diverse library. The antiviral activity of these two Pr55Gag binders was investigated using different cell systems. Two aRep scaffolds aRep4E3 and aRep9A8 were isolated and characterized. aRep4E3 contains 7 repeat motifs (32 kDa), meanwhile aRep9A8 has 6 repeat motifs (28 kDa). These two scaffold molecules found to be able to display antiviral effects on the late stage of HIV-1 replication, by reducing and delaying the viral progeny production. The difference in the molecular mechanism was observed between these two aRep proteins: aRep4E3 mainly interferes with the packaging of the viral genome, meanwhile aRep9A8 interferes with the proteolytic processing of Gag, and performs as a protease inhibitor to prevent the PR cleavage required for the production of newly infectious mature virus. Interestingly, aRep9A8 is able to survive in the chronical HIV-1 infected cells up to D38 pi with the low level of HIV-1 replication. Taken together, results suggested that aRep, a new type of scaffold protein could serve as a promising alternative agent in protein therapy, not only the HIV-1 infection but also the others pathogens or disorders
8
Storm, Rickard. "Early host cell interactions and antivirals against ocular adenoviruses." Doctoral thesis, Umeå universitet, Virologi, 2015. http://urn.kb.se/resolve?urn=urn:nbn:se:umu:diva-99907.
Full textAbstract:
Viruses are common causative agents of ocular infection among humans. Epidemic keratoconjuntivitis (EKC) is a severe and contagious ocular disease with reported outbreaks worldwide. It is estimated that this disease affects 20-40 million individuals every year, which leads to huge socioeconomic costs for the affected countries. EKC is characterized by keratitis and conjunctivitis but is also associated with pain, edema, lacrimation, and decreased vision that can prolong for months after the infection and in rare cases years. This disease is caused by human adenoviruses (HAdVs), which belong to the family of Adenoviridae. Currently, there is no available treatment against EKC. EKC is mainly caused by HAdV-8, HAdV-19, HAdV-37, HAdV-53, HAdV-54, and HAdV-56, which belong to species D HAdVs. HAdV-8, HAdV-19 and HAdV-37 have previously been shown to use sialic acid (SA)-containing glycans as cellular receptors to bind to and infect human corneal epithelial (HCE) cells. To characterize the receptor in more detail, we performed a glycan array, which included SA-containing glycans. A branched hexasaccharide terminating with SA in each arm was identified as a candidate receptor. This glycan corresponds to the glycan motif found on a ganglioside, GD1a. By performing a series of biological and biochemical experiments we confirmed the function of the GD1a glycan as a cellular receptor for EKC-causing HAdVs. However, the glycan used as a receptor was linked to plasma membrane protein(s) through O-glycosidic bonds, rather than to a lipid (as in the ganglioside). X-ray crystallography analysis showed that the two terminal SA:s interacted with two of the three previously identified SA-binding sites on the knob domain of the HAdV-37 capsid protein known as the fiber. Based on the structural features of the GD1a:HAdV-37 knob interaction, we assumed that a three-armed molecule with each arm terminating with SA would be an efficient inhibitor. Such molecules were designed, synthesized and found to efficiently prevent HAdV-37 binding to and infection of corneal cells. These results indicate that trisialic acids-containing compounds may be used for treatment of EKC. After binding to its primary receptor, most HAdVs have been shown to interact with αVβ3 and αVβ5 integrins to enter human cells. This interaction occurs through the RGD (arginine-alanine-aspartic acid) motif in the capsid protein known as the penton base. However, it was not clear if corneal epithelial cells express αVβ3 and αVβ5 integrins. Thus, to better understand additional early steps of infection by EKC-causing HAdVs, we performed binding and infection competition experiments using human corneal epithelial cells and siRNA, integrin specific antibodies, peptides and RGD-containing ligands indicating that α3, αV, β1 affected HAdV-37 infection of but not binding to HCE cells. We could also see that HAdV-37 co-localize with α3 and αV at after entry into HCE cells. In situ histochemistry confirmed that the expression of α3 and αV in human corneal tissue. Overall, our results suggest that αV and α3 integrins are important for HAdV-37 infection of corneal cells. Altogether, these results provide further insight into the biology of HAdVs and open up for development of novel antiviral drugs.
9
Khedr, Mohammed Abdou. "Computer-aided drug design and synthesis of novel antivirals." Thesis, Cardiff University, 2010. http://orca.cf.ac.uk/54378/.
Full textAbstract:
The Flaviviridae is a family of 66 viruses of which almost half have been associated with human disease. The most well-known members are: Hepatitis C virus (HCV), Dengue virus (DV), and West Nile virus (WNV). Diseases caused by these viruses are a global health problem that put an estimated 2.5 billion people at risk. At present, there are neither vaccines nor other treatments available to prevent or cure these diseases. Potential targets for the development of therapeutics against the virus are the viral protease and polymerase. The aims of this project are to design and synthesize compounds that can be used as inhibitors for these two key enzymes for Dengue. Structure-based drug design methods utilize knowledge of a three dimensional structure of an enzyme/receptor to develop small molecules able to bind to the desired target, generating a specific biological response. These computer-based methodologies are now becoming an integral part of the drug discovery process and, although the principles of molecular recognition are far from being completely understood, some marketed compounds (i.e. Zanamivir, Lopinavir) have been developed with the help the of successful application of structure-based design techniques. Different structure-based drug design approaches have been used to identify putative new inhibitors for the Dengue protease and polymerase. A pharmacophore query has been built based on the active site of the Dengue protease enzyme and then used for screening different databases for identification of potential inhibitors. For the polymerase, a fragment-based approach has been used to find the fragments that would interact more efficiently with a specific binding pocket on the enzyme. The virtual library obtained by linking the best scored fragment was then docked to identify the most promising structures to be synthesized. The identification of potent small molecules that bind to receptors and enzymes is one of the major goals of chemical and biological research.
10
Bhave, Sukhada. "INVESTIGATING SYNERGY BETWEEN RIBONUCLEOTIDE REDUCTASE INHIBITORS AND CMV ANTIVIRALS." VCU Scholars Compass, 2012. http://scholarscompass.vcu.edu/etd/2838.
Full textAbstract:
Cytomegalovirus (CMV) infections remain a significant problem in congenitally infected infants and immunocompromised individuals. Modest antiviral activities of currently approved drugs coupled with dose-limiting toxicities restrict effectiveness and promote development of resistance. The potential for ribonucleotide reductase (RR) inhibitors hydroxyurea (HU), Didox, and Trimidox to synergize, through reduction of nucleotide pools, with the deoxynucleotide analog Ganciclovir (GCV) was examined. A yield reduction assay that utilizes luciferase expressed by a recombinant virus as a surrogate measure of viral infectious units was developed and used to determine effective dose ranges for each drug. RR inhibitors exhibited intrinsic anti-CMV activities on their own with IC50 values well below toxic levels. Moreover, RR inhibitors significantly synergized with GCV. These findings provide a rationale for exploration of RR inhibitors and deoxynucleotide analogs in anti-CMV combination therapy.
11
Jun, Min Medical Sciences Faculty of Medicine UNSW. "Analysis of human cytomegalovirus susceptibility to novel antiviral agents." Publisher:University of New South Wales. Medical Sciences, 2008. http://handle.unsw.edu.au/1959.4/41443.
Full textAbstract:
Human cytomegalovirus (CMV) is a significant infectious agent causing disease in immunocompromised HIV-infected patients, transplant recipients, and neonates. The current antiviral therapeutic strategy against CMV is limited in its utility due to the inherent toxicity and lack of bioavailability of currently available anti-CMV agents, ganciclovir (GCV), cidofovir (CDV), and foscarnet (FOS). The development of the prodrug of GCV, valganciclovir (val-GCV), has vastly improved the bioavailability profile of GCV. However, val-GCV demonstrates limited effectiveness against tissue-invasive CMV diseases as side effects involved with traditional intravenously administered GCV such as haematologic and reproductive toxicities remain. In addition, the emergence of antiviral resistant CMV mutant strains due to prolonged treatment with currently available antivirals necessitates the development of novel anti-CMV agents with reduced toxicity and improved bioavailability. In this study, select groups of novel compounds were analysed for their potential for further development as anti-CMV agents. Three groups of compounds were identified based on two screening methods which included the computer simulated screening process of compounds known as in silico screening and the traditional method of random screening. The first group of compounds (CATi) were identified by in silico screening against the CMV DNA polymerase catalytic aspartate triad, resulting in the identification of 31 compounds with the potential for inhibitory activity against CMV. The second group of compounds (PRO-i) were identified through in silico screening against the CMV protease, identifying a total of 18 lead compounds exhibiting structural complementarity with CMV protease. The third and final group of compounds (TPEX) were identified through random screening and consisted of plant extracts purified from tropical plants. All three compounds were initially screened for cytotoxicity against human fibroblasts. Plaque reduction assays were performed using compounds with acceptable levels of cytotoxicity to determine the ability of the compounds to inhibit the replication of the laboratory antiviral sensitive CMV strain, Towne. Two of the PRO-i compounds demonstrated good antiviral activity against CMV. Eleven percent (2/18) of the PRO-i compounds inhibited CMV replication, with PRO-i-43 and PRO-i??-44 displaying mean 50% inhibitory concentrations (IC50) of 4.8 ?? 1.2 ??M and 8.04 ??M, respectively. PRO-i-43 and PRO-i-44 are thus good candidates for further development as novel antiviral agents against CMV. The majority of CATi and TPEX compounds displayed significant cytotoxicity against human fibroblasts and compounds with acceptable levels of cytotoxicities did not significantly inhibit CMV replication. However, the identification of compounds with low cytotoxicities provides a good foundation for further development of novel anti-CMV agents with superior antiviral activity. In silico screening against three-dimensional viral protein models is a useful strategy for the identification of novel antiviral agents with the potential for inhibitory activity against CMV. Structural modification to produce potent derivatives of the identified anti-CMV compounds (PRO-i-43 and PRO-i-44) is a good option for the further development of novel antiviral agents against CMV. Such further examination of the identified compounds with anti-CMV activity is required to investigate their activity against not only antiviral sensitive CMV strains but also resistant CMV strains. Further investigations will yield new insights into their target, allowing further identification of compounds with potential anti-CMV activity with pharmaceutical application.
12
Lang, Yuekun. "Identification and evaluation of antivirals for Rift Valley fever virus." Diss., Kansas State University, 2017. http://hdl.handle.net/2097/38195.
Full textAbstract:
Doctor of PhilosophyDepartment of Diagnostic Medicine/Pathobiology
Wenjun Ma
Rift Valley fever virus (RVFV) is an enveloped, negative-sense, ssRNA virus with a tripartite genome that causes morbidity and mortality in both livestock and humans. Although RVFV is mainly circulating in mainland Africa, this arthropod-borne virus is a potential threat to the other parts of the world. No fully licensed vaccines for human or animal use in the U.S., and effective antiviral drugs have not been identified. As virulent RVFV strains are only handled in biosafety level (BSL) 3 or higher level facilities in the U.S., few laboratories have access to RVFV which limits antiviral development. However, it is crucial to develop effective antivirals to protect public and animal health. Animal models that reproduce Rift Valley fever are vital to identifying and developing antiviral compounds. The currently available attenuated RVFV strain, MP12, provides a BSL-2 challenge model virus for preliminary investigations of RVFV prior to using the virulent RVFV strains. All strains of RVFV have a highly conserved genome, indicating that antivirals or vaccines effective against any RVFV strain will most likely be effective for all RVFV strains. Therefore, we hypothesize that the MP12 is a suitable model virus that can be used for identification and evaluation of effective RVF antivirals. The first objective of this project was to establish a mouse model susceptible to MP12 infection. Based on the literature, we selected and screened six different strains of mice to test their susceptibilities to MP12. We found the STAT-1 knockout mice are the most susceptible to MP12 infection based on clinical symptoms, mortality, viremia, virus replication, histopathological, and immunochemical analyses. Importantly, these mice displayed acute-onset hepatitis and delayed-onset encephalitis similar to severe cases of human RVFV infection. Our second objective was to identify potential antiviral drugs in vitro. We developed and employed a cell-based assay using the recombinant MP12 virus expressing Renilla luciferase to screen a library of 727 small compounds purchased from National Institutes of Health. Of the compounds, 23 were identified and further tested for their inhibitory activities on the recombinant MP12 virus expressing green fluorescent protein. Further plaque reduction assays confirmed that two compounds inhibited replication of parental RVFV MP12 strain with limited cytotoxic effects. The 50% inhibitory concentrations using an MP12 multiplicity of infection (MOI) of 2 were 211.4 µM and 139.5 µM, respectively. Our third objective was to evaluate these two candidates, 6-azauridine and mitoxantrone, in vivo using our mouse model. After one-hour post MP12 infection via an intranasal route, treatment was given intranasally twice daily. Mice treated with placebo and 6-azauridine displayed severe weight loss and reached the threshold for euthanasia with obvious neurological signs, while mice treated with ribavirin (a known antiviral drug) or mitoxantrone showed delayed onset of disease. This result indicates that the mitoxantrone can improve the outcome of RVFV infection in our mouse model. The underlying mechanism of mitoxantrone to inhibit RVFV replication remains to be investigated. Our studies build the foundation for identification and development of antivirals against RVFV in a BSL-2 environment.
13
Richard, Mathilde. "Diversité des mécanismes de résistance aux inhibiteurs de la neuraminidase des virus influenza A : implications de résidus conservés dans le site actif de la neuraminidase et de la balance fonctionnelle entre la neuraminidase et l’hémagglutinine." Thesis, Lyon 1, 2010. http://www.theses.fr/2010LYO10323.
Full textAbstract:
Chaque année, les épidémies de grippe, dont les principaux agents étiologiques sont les virus influenza de type A, ont un impact considérable sur la population en terme de morbidité et de mortalité. Le virus influenza A comporte à sa surface deux glycoprotéines, la neuraminidase et l’hémagglutinine. Ces deux protéines ont des fonctions antagonistes : l’hémagglutinine permet l’entrée du virus dans la cellule hôte et la neuraminidase, par son activité sialidase, libère les nouveaux virions formés. Bien que la prophylaxie du virus grippal repose essentiellement sur la vaccination, les antiviraux jouent un rôle important dans la lutte contre les épidémies de grippe et dans la stratégie développée en prévision d'une pandémie grippale. Les inhibiteurs de la neuraminidase (INAs) sont des antiviraux efficaces contre la grippe. Ils inhibent l’activité enzymatique de la neuraminidase et empêchent la libération des nouveaux virions formés. La démarche méthodologique qui a conduit à l’élaboration de molécules ciblant la neuraminidase laissait espérer une apparition limitée de résistance. Cependant, des cas de résistances aux INAs ont été mis en évidence lors d’études cliniques. Outre la nécessité d’une surveillance étroite, il est donc important d’étudier et de comprendre les diverses mécanismes susceptibles d’induire une résistance aux INAs. Le travail de cette thèse s’est ainsi porté sur la compréhension de la diversité des mécanismes de résistance. Dans un premier temps, nous avons étudié l’impact de mutations sur l’ensemble des résidus structuraux du site actif de la neuraminidase. Nous avons observé que la plupart de ces mutations n’altéraient pas les caractéristiques du virus et induisaient une légère baisse de sensibilité aux INAs. Par la suite, nous avons cherché à explorer les possibilités de synergie dans la résistance aux INAs par la combinaison de deux mutations structurales du site actif de la neuraminidase. Sur quatre virus produits, seul le virus possédant la double mutation E119V+I222L était viable, malgré une capacité réplicative in vitro altérée. La combinaison de ces deux mutations induit une synergie dans la résistance à l’oseltamivir. Enfin, nous avons voulu intégrer l’interaction fonctionnelle de la neuraminidase avec l’hémagglutinine. Nous avons montré que la combinaison d’une hémagglutinine de faible affinité pour les récepteurs sialylés permettait de restaurer un virus possédant une neuraminidase déficiente. Ainsi, un virus influenza peut se libérer de la fonction de la neuraminidase, cible des seuls antiviraux efficaces disponibles à l’heure actuelle. Les mécanismes de résistances aux inhibiteurs de la neuraminidase sont multiples. L’émergence durant les deux dernières saisons hivernales de virus résistants aux INAs sans pression de sélection a remis en question les hypothèses développées sur l’infectivité et la transmissibilité de souches résistantes, ouvrant de nouvelles perspectives quant à l’étude des mécanismes permettant l’obtention de virus épidémiogènes résistants aux INAsEach winter, influenza epidemics have a considerable impact on the population in terms of morbidity and mortality. Influenza A virus is the main etiologic agents of influenza. They present at their surface two glycoproteins, the neuraminidase and the hemagglutinin. These two proteins have antagonist functions : the hemagglutinin allows the virus to enter the host cell and the neuraminidase, through its sialidase activity, releases progeny virions from host cells. Although prophylaxis of influenza is mainly based on vaccination, antiviral drugs play a very important role in the fight against epidemics of influenza and the strategy developed in anticipation of a flu pandemic. The neuraminidase inhibitors are effective antiviral against influenza. Through the inhibition of the neuraminidase enzymatic activity, they prevent the release of new virions formed. The introduction into clinical practice of new drugs requires monitoring in order to detect the potential emergence of resistance. Although the approach to the design of neuraminidase inhibitors has provided hope that resistance will be limited, resistance to NAIs already been observed in clinical, encouraging close monitoring. It is therefore important to continue to study and understand the various mechanisms of resistance to neuraminidase inhibitors. The work of this thesis has thus focused on understanding the diversity of resistance mechanisms. Initially, we studied the impact of mutations in all structural residues of the active site of neuraminidase. We observed that most of these mutations did not alter the characteristics of the virus and induced very limited resistance to antivirals. Subsequently, we then sought to explore opportunities for synergy in resistance by the combination of two structural mutations of the active site of neuraminidase. On four viruses produced, only the virus with the double mutation E119V+I222L in the active site of neuraminidase was viable, although its in vitro replicative capacity was impaired. The combination of these two mutations induced a synergistic resistance to oseltamivir. Finally, we wanted to integrate the functional interaction of neuraminidase with hemagglutinin. We have shown that the combination of a hemagglutinin low affinity for sialylated receptors allowed to rescue a virus with a deficient neuraminidase. Thus an influenza virus may discharge the function of neuraminidase, the target of the only available effective antivirals. The mechanisms of resistance to neuraminidase inhibitors are numerous. Plus, the circulation in the last two seasons of resistant viruses without selective pressure challenges the assumptions developed on the possible emergence of resistance in clinic. This opens new issues to consider in order to understand the mechanisms that allowed this emergence and transmission
14
Sheehy, Noreen. "Analysis of AZT sensitive and resistant HIV-1 strains." Thesis, University of Warwick, 1995. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.308439.
Full text
15
Talló, Parra Marc 1992. "Circular RNAs : from host RNA molecules to novel broad-spectrum antivirals." Doctoral thesis, Universitat Pompeu Fabra, 2019. http://hdl.handle.net/10803/668309.
Full textAbstract:
La rellevància clínica dels virus transmesos per mosquits, com el virus del dengue (DENV), el virus del zika (ZIKV), el virus del chikungunya (CHIKV) i el virus del Nil Occidental (WNV), ha augmentat dràsticament durant els darrers anys, provocant un problema de salut global. Actualment, no hi ha cap tractament disponible ni cap vacuna efectiva per tractar aquestes infeccions. Tots aquests virus causen infeccions agudes que han de ser tractades ràpidament després de l’aparició dels símptomes inicials perquè els medicaments siguin efectius. Tot i això, el diagnòstic precoç continua sent un repte no resolt. Això evidencia la necessitat de descobrir noves interaccions essencials entre el virus i la cèl·lula que podrien ser utilitzades com a noves dianes terapèutiques; i la necessitat de desenvolupar teràpies antivirals d’ampli espectre, eficients, que puguin ser administrades abans que s’aconsegueixi un diagnòstic precís. En aquesta tesi hem abordat aquests dos grans problemes centrant-nos en els ARNs circulars (circRNAs).
Els circRNAs són una classe d’ARN generats a partir de progenitors lineals d'ARN mitjançant un mecanisme alternatiu de splicing anomenat back-splicing. En comparació amb els seus homòlegs lineals, els circRNAs són molt estables a causa de la seva resistència a les exonucleases. Actualment, s’ha descrit la implicació dels circRNAs en les infeccions virals, tanmateix, no es coneix el seu rol precís. El primer capítol de la tesi intenta respondre a aquest buit de coneixement utilitzant el virus de l’hepatitis C (HCV) com a sistema model i analitza l'efecte dels circRNAs identificats en altres virus de la mateixa família, en concret, els virus transmesos per mosquits. Mitjançant anàlisis de seqüenciació d’ARN, hem identificat 73 circRNAs cel·lulars induïts pel HCV. Aquest canvi en l’expressió dels circRNAs no pot ser explicat a través de canvis paral·lels en els ARNs lineals. A més a més, hem identificat que el silenciament de cinc d’aquests circRNAs provoca canvis en la infectivitat viral, actuant com a molècules pro- o anti- virals. Un d’aquests circRNAs, cPSD3, és clau per a la infectivitat del virus del dengue.
El segon capítol de la tesi se centra en desenvolupament d’una nova plataforma basada en circRNAs que sigui versàtil, dificulti l’aparició de mutants resistents i permeti desenvolupar antivirals d’ampli espectre. En contrast amb altres teràpies basades en ARN, els circRNAs són molècules altament estables, una característica que simplificarà el seu ús terapèutic. Els circRNAs sintètics dissenyats contenen seqüències llargues que s’hibriden a regions del genoma viral implicades en formar estructures d’RNA essencials per a la supervivència del virus. Com a prova de concepte, hem validat amb èxit circRNAs que inhibeixen el HCV, el DENV, el CHIKV o el WNV. A més, hem generat circRNAs amb capacitat antiviral d’ampli espectre i hem optimitzat la producció in vitro d’aquestes molècules per obtenir quantitats elevades a baix preu.
En conclusió, els nostres resultats (i) emfatitzen la complexitat de la interacció entre els circRNAs cel·lulars i els virus i (ii) descobreixen el gran potencial dels circRNAs artificials com a noves plataformes per al desenvolupament de fàrmacs.The clinical importance of the mosquito-borne viruses, such as dengue virus (DENV), zika virus (ZIKV) chikungunya virus (CHIKV) and West Nile virus (WNV), has dramatically increased over the last years, resulting in a global health problem. Currently, there are no available treatments or effective vaccines to treat these infections. All these viruses produce acute infections that require to be treated early after the onset of the symptoms for drugs to be effective. However, an early diagnosis remains still as an unsolved challenge. This brings to the spotlight the need to uncover novel fundamental virus-cell interactions that could be targeted and to develop efficient broad-spectrum antiviral therapies that could be administered before an accurate diagnosis is achieved. In this thesis we addressed these two major concerns with a focus in circular RNAs (circRNAs). CircRNAs are a class of RNAs generated from linear RNA progenitors by an alternative splicing mechanism termed back splicing. They are highly stable relative to their linear spliced counterparts due to exonuclease resistance. Currently, cellular circRNAs are described to be involved in viral infections. However, their precise role is mainly unknown. The first chapter of the thesis addresses this intriguing issue using HCV as a model system and analyzing the effect of the identified circRNAs in mosquito-borne viruses that belong to the same viral group. By RNA-Seq analyses we identified 73 HCV-differentially expressed circRNAs whose changes could not be explained by parallel changes in linear mRNAs. Silencing of five selected HCV-induced up-regulated circRNAs altered viral infectivity, acting either as anti-viral or pro-viral molecules. Further characterization of one of the selected circRNAs, cPSD3, show, that it also impaired DENV infections. The second chapter focuses on the generation of a novel circRNA-based platform that is versatile, hampers the emergence of resistant mutants, and allows developing broad-spectrum antivirals. In contrast to other RNA-based therapies, circRNAs are highly stable molecules, a trait that will simplify their therapeutic use. The designed synthetic circRNAs contain long sequences that hybridize to multiple target sequences in the viral RNA genome involved in forming RNA structures essential for virus survival. As a proof of concept, we have successfully validated circRNAs that inhibit HCV, DENV, CHIKV or WNV. Furthermore, we have generated circRNAs with broad-spectrum antiviral capacity and optimized the production in vitro of these molecules to obtain high amounts at low price. In conclusion, our results (i) emphasize the complexity of the interaction between cellular circRNAs and viruses and (ii) uncover the great potential of artificial circRNAs as novel platforms for drug development.
16
Lacey, Simon F. "Herpesvirus enzymes involved in nucleotide metabolism as targets for novel antivirals." Thesis, Open University, 1990. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.255214.
Full text
17
Abdurakhmanov, Eldar. "Discovery and evaluation of direct acting antivirals against hepatitis C virus." Doctoral thesis, Uppsala universitet, Biokemi, 2015. http://urn.kb.se/resolve?urn=urn:nbn:se:uu:diva-265299.
Full textAbstract:
Until recently, the standard therapy for hepatitis C treatment has been interferon and ribavirin. Such treatment has only 50% efficacy and is not well tolerated. The emergence of new drugs has increased the treatment efficacy to 90%. Despite such an achievement, the success is limited since the virus mutates rapidly, causing the emergence of drug resistant forms. In addition, most new drugs were developed to treat genotype 1 infections. Thus, development of new potent antivirals is needed and drug discovery against hepatitis C is continued. In this thesis, a FRET-based protease assay was used to evaluate new pyrazinone based NS3 protease inhibitors that are structurally different to the newly approved and currently developing drugs. Several compounds in this series showed good potencies in the nanomolar range against NS3 proteases from genotype 1, 3, and the drug resistance variant R155K. We assume that these compounds can be further developed into drug candidates that possess activity against above mentioned enzyme variants. By using SPR technology, we analyzed interaction mechanisms and characteristics of allosteric inhibitors targeting NS5B polymerases from genotypes 1 and 3. The compounds exhibited different binding mechanisms and displayed a low affinity against NS5B from genotype 3. In order to evaluate the activity and inhibitors of the NS5B polymerase, we established an SPR based assay, which enables the monitoring of polymerization and its inhibition in real time. This assay can readily be implemented for the discovery of inhibitors targeting HCV. An SPR based fragment screening approach has also been established. A screen of a fragment library has been performed in order to identify novel scaffolds that can be used as a starting point for development of new allosteric inhibitors against NS5B polymerase. Selected fragments will be further elaborated to generate a new potent allosteric drug candidate. Alternative approaches have successfully been developed and implemented to the discovery of potential lead compounds targeting two important HCV drug targets.
18
Prachanronarong, Kristina L. "Understanding Drug Resistance and Antibody Neutralization Escape in Antivirals: A Dissertation." eScholarship@UMMS, 2016. https://escholarship.umassmed.edu/gsbs_diss/840.
Full textAbstract:
Antiviral drug resistance is a major problem in the treatment of viral infections, including influenza and hepatitis C virus (HCV). Influenza neuraminidase (NA) is a viral sialidase on the surface of the influenza virion and a primary antiviral target in influenza. Two subtypes of NA predominate in humans, N1 and N2, but different patterns of drug resistance have emerged in each subtype. To provide a framework for understanding the structural basis of subtype specific drug resistance mutations in NA, we used molecular dynamics simulations to define dynamic substrate envelopes for NA to determine how different patterns of drug resistance have emerged in N1 and N2 NA. Furthermore, we used the substrate envelope to analyze HCV NS3/4A protease inhibitors in clinical development. In addition, influenza hemagglutinin (HA) is a primary target of neutralizing antibodies against influenza. Novel broadly neutralizing antibodies (BnAbs) against the stem region of HA have been described and inhibit several influenza viral subtypes, but antibody neutralization escape mutations have emerged. We identified potential escape mutations in broadly neutralizing antibody F10 that may impact protein dynamics in HA that are critical for function. We also solved crystal structures of antibody fragments that are important for understanding the structural basis of antibody binding for influenza BnAbs. These studies can inform the design of improved therapeutic strategies against viruses by incorporating an understanding of structural elements that are critical for function, such as substrate processing and protein dynamics, into the development of novel therapeutics that are robust against resistance.
19
Foca, Adrien. "Identification of PLK1 as a proviral factor for the hepatitis B virus replication : A possible target for antiviral and anticancerous drug development." Thesis, Lyon, 2018. http://www.theses.fr/2018LYSE1310/document.
Full textAbstract:
Dans les régions de fortes endémicités, 70-80% des carcinomes hépatocellulaires sont induits par le VHB. Bien qu’un vaccin prophylactique très efficace existe, il n’est d’aucune utilité pour les 250 millions de personnes chroniquement infectées. Les traitements actuels pour contrôler l'infection chronique par le VHB montrent des limites et le besoin de nouvelles thérapies se fait ressentir. La Polo-like kinase-1 (PLK1), qui joue un rôle essentiel dans la mitose et est surexprimée dans de nombreux cancers, représente une cible prometteuse. Outre son rôle lors de la division cellulaire, PLK1 est impliquée dans la régulation de l'expression des gènes en interphase. Il a été montré que la protéine X du VHB (HBx) active PLK1 dans des modèles de cellules murines. Cependant, il restait à déterminer si PLK1 jouait un rôle au niveau de la réplication du VHB dans des hépatocytes quiescents. Des études récentes ont mis en évidence un lien positif entre l'activation de PLK1 et la réplication du VHB. Le but de ce projet de thèse a été d'étudier le(s) mécanisme(s) par le(s)quel(s) PLK1 jouait un rôle positif sur la réplication virale, avec pour objectif futur d'explorer l’inhibition de PLK1 comme cible antivirale. L'interaction entre PLK1 et la réplication du VHB a d'abord été décrite à l'aide du modèle HepAD38. Dans ce contexte, l'ADN viral est intégré dans le génome hôte, sous le contrôle d'un système d'expression Tet-off. La transcription de l'ARN prégénomique (pgRNA), à la base de la réplication virale, est initiée par la suppression de tétracycline. Dans ce contexte, l'augmentation de l'expression de PLK1 est corrélée avec la régulation négative de deux protéines; SUZ12 et ZNF198, faisant partie de complexes de remodelage de la chromatine. L'inhibition de PLK1 bloque la réplication du VHB, en agissant au niveau de la transcription virale. D'autre part, dans les modèles de réplication du VHB qui miment au mieux une infection, comprenant les hépatocytes primaires humains (PHH) et les cellules non transformées/différenciées HepaRG (dHepaRG), où le VHB se réplique dans des cellules quiescentes, nous avons mis en évidence que: 1) L'inhibition pharmacologique de PLK1 bloque la réplication virale, semblablement en perturbant l’encapsidation du pgRNA via une interaction avec la protéine core du VHB (HBc). 2) Un knocking-down de PLK1 en utilisant des ARN interférents délivrés par nanoparticules lipidiques résulte en une forte baisse de la production de pgRNA et dans la sécrétion des antigènes HBeAg/HBsAg, sans impact sur la viabilité cellulaire. Ce projet a donc permis la preuve de concept que PLK1 pouvait être une cible thérapeutique afin de controler la réplication du VHB. De plus, grâce à la technologie de délivrance par nanoparticules lipidiques d’ARN interférents, nous avons pu cibler spécifiquement les hépatocytes, augmentant de ce fait la spécificité et l’efficacité de nos traitements. Un travail sur la compréhension précise des méchanismes cellulaires impliqués permettra de mieux cerner cette interaction hôte/virus afin de poursuivre le développement de stratégies antivirales innovantes portant sur l’inhibition de PLK1. De manière significative, l'inhibition de PLK1 est non toxique pour les cellules quiescentes par rapport à des cellules cancéreuses à fort taux réplicatif, ce qui fait de PLK1 une cible thérapeutique attrayante. Des inhibiteurs spécifiques sont déjà en essais cliniques pour certains cancers (e.g., Volasertib pour le traitement de la leucémie myéloïde aiguë) et pourraient servir de thérapie bimodale dans le cadre de patients infectés par le VHB, en inhibant la réplication virale, ainsi qu’en prévenant l'émergence de cellules néoplasiques. L'inhibition de la PLK1 est une approche antivirale innovante, qui, en combinaison avec les thérapies actuelles de type IFN-α ou analogues nucléotidiques offre de grandes promesses pour endiguer l'infection chronique par le VHB mais également prévenir les événements carcinogéniquesIn highly HBV endemic regions, 70-80% of hepatocellular carcinoma cases are attributable to this virus. Despite the existence of an HBV vaccine, the World Health Organization estimates 240 million individuals are chronically infected with HBV worldwide. Current antiviral treatments to control chronic HBV infections, and consequently reduce the incidence of liver cancer, are ineffective. New and effective therapies are needed not only for fighting the virus but also to prevent HCC emergence or progression. The polo-like-kinase 1 (PLK1), which plays pivotal roles in mitosis and is over-expressed in many human cancers, represents a promising druggable target in oncology. Beside its role during cell division, PLK1 is also thought to be involved in gene expression regulation during interphase. It was shown that the X protein (HBx) could activate PLK1 in murine cell transformation models. Yet it remained to be determined whether PLK1 could also play a role for HBV replication in non-dividing hepatocytes. Our, and collaborators, recent studies have identified a positive link between PLK1 activation and HBV replication. The goal of this thesis project was to investigate the mechanism(s) by which PLK1 exerts a positive effect on HBV replication, with the future goal of exploring PLK1 as an antiviral target. The interplay between PLK1 and HBV replication was firstly described using the HepAD38 cellular model of HBV replication. In this context, the HBV DNA is stably integrated into the host genome, under control of a Tet-off expression system. Transcription of HBV pregenomic RNA (pgRNA), the template of viral replication, is initiated by tetracycline removal. It has been shown that in HBV-replicating HepAD38 cells, increased PLK1 expression correlates with down-regulation of two proteins that are components of chromatin modifying complexes; SUZ12 protein of the PRC2 complex, and ZNF198 of the LSD1-CoREST-HDAC1 complex. PLK1 inhibition was described to inhibit HBV replication by reducing viral transcription. How PLK1 regulates HBV transcription remains unknown. On the other hand, in HBV replication models that resemble physiologic HBV infection, comprised of Primary Human Hepatocytes (PHH) and non-transformed/differentiated HepaRG cells (dHepaRG), where HBV replicates in non-transformed and non-dividing cells, thus enabling the study of the inter-phasic role of PLK1, irrespective of its well-established cell division implication, we have demonstrated that: 1) A pharmacological inhibition of PLK1 suppressed HBV replication by a different mechanism, likely targeting the packaging of pgRNA by the HBV core antigen (HBc). 2) Knocking-down PLK1 using siRNA delivered by lipid nanoparticles (LNP siPLK1) results in a strong drop of HBV DNAs, RNAs and HBe/HBsAg secretion without affecting the cell viability. This thesis project brought the proof of concept that PLK1 could be a drug target in HBV infection. Furthermore, the use of LNP allowed us to improve the delivery of siPLK1 to hepatocytes. Significantly, PLK1 inhibition is not toxic to quiescent cells in comparison to fast growing cancer cells, rendering PLK1 an attractive therapy target. High level of viremia in chronic HBV patients is a risk factor for progression to liver cancer. PLK1 specific inhibitors are already in clinical trials for other types of cancer (e.g., acute myeloid leukaemia) and could serve as bimodal therapy in HBV infected patients, by inhibiting virus replication as well as preventing emergence and spreading of neoplastic cells. This project was part of a full-working group of experts and thus, has beneficiated of a strong support. The proximity of the oncology-specialized hospital, the Centre Léon Bérard provided us with fresh hepatic biopsy [etc...]
20
Nastasja, Palombi. "Broad antiviral compounds targeting viral envelope: synthesis and pre-clinical studies." Doctoral thesis, Università di Siena, 2020. http://hdl.handle.net/11365/1096254.
Full textAbstract:
The majority of viral pathogens that cause emerging and re-emerging infectious diseases are membrane-enveloped viruses.
In this thesis work a new series of antiviral compounds, with 1H-pyrrol-methylene thioxodihydropyrimidine structure, were designed and synthesized.
They were endowed with antiviral activity on enveloped viruses, likely ZIKAV, DENV-2, and five influenza virus strains, including the pandemic H7N9 model.
The intriguing mechanism of action was pursued and confirmed the hypothesis of compounds able to intercalate in the viral lipid envelope membrane and to oxidize phospholipids, impairing the physical and chemical structure of the viral membrane and impacting the fusion of viral and cell membranes.
With the aim of investigating in vitro and in vivo pre-clinical features, compounds were assayed for their chemical-physical properties. Finally, a preliminary in vivo PK was assessed.
21
Satterly, Neal Gilpin. "The mRNA Nuclear Export Machinery is Targeted by Influenza Virus and Antivirals." Scholarly Repository, 2009. http://scholarlyrepository.miami.edu/oa_dissertations/197.
Full textAbstract:
Proper mRNA nuclear export is essential for harmonious growth and maintenance of a cell. An effective weapon influenza virus employs to hijack a host cell is its ability to inhibit such export. Exactly how influenza virus achieves this inhibition is not fully known. Here, we demonstrate that upon infection, influenza virus degrades two nucleopore proteins (Nup98 and Nup96), which play a key role in mRNA nuclear export. Also, a main virulence factor of influenza virus (non-structural protein 1, NS1) binds directly to NXF1 and E1B-AP5, two key constituents of the mRNA export pathway (NXF1/NXT pathway) responsible for exporting bulk (~70%) mRNA from the nucleus. By increasing the expression levels of members of the NXF1/NXT pathway, we were able to reverse NS1-mediated inhibition of gene expression. On the other hand, by decreasing the levels of members of the NXF1/NXT pathway, we demonstrated that host cells become more sensitive to influenza virus infection and produce more viral particles. These results demonstrate undiscovered influenza-mediated host interactions that may be used to medicinally inhibit influenza virus. To this end, high-throughput screens were designed to identify small molecule antagonists of both NS1-mediated inhibition of gene expression and influenza virus-mediated cell death. Seventy-one compounds were identified, and the most potent molecule (named compound #8) was examined further. We found that compound #8 releases influenza virus-mediated mRNA nuclear export blockage and decreased viral replication and viral gene expression. Thus, the bulk mRNA nuclear export machinery is vital to antiviral response, and compound #8 enhances its ability to fight the cytopathic effects of NS1 and influenza virus. In conclusion, our data demonstrate that the mRNA export machinery is disrupted by influenza virus, and that this machinery also facilitates an antiviral function. We have also shown that these two events can be manipulated chemically to attenuate the negative effect of the virus and enhance the positive antiviral effect of the mRNA export machinery, thereby providing a powerful, new strategy against the ever-present, global threat of influenza virus.
22
Wing, Peter Alexander Cornelius. "Reduced sensitivity of Genotype 3 hepatitis C virus to direct acting antivirals." Thesis, Queen Mary, University of London, 2018. http://qmro.qmul.ac.uk/xmlui/handle/123456789/44044.
Full textAbstract:
Sofosbuvir is a uridine based nucleotide inhibitor of the hepatitis C viral (HCV) polymerase that is the backbone of many treatment regimens. In combination with drugs targeting other viral enzymes (including the poorly potent guanosine analogue ribavirin or highly potent inhibitors of viral NS5A or protease) most patients clear virus and resistance to sofosbuvir is rare, allowing effective retreatment with sofosbuvir. Patients with Genotype 3 HCV respond less well than other genotypes and response is reduced in those previously exposed to interferon. Here we show that patientderived virus from patients with Genotype 3 HCV who relapse to sofosbuvir-based therapies have a reduced sensitivity to SOF in an in-vitro phenotyping assay. Analysis of viral sequencing data revealed two distinct polymorphisms (A150V and K206E) in the HCV polymerase that are associated with treatment failure and in-vitro; they reduce sofosbuvir sensitivity against genotype 3 hepatitis C virions. However both polymorphisms modify the cellular response to type I interferon and in cells lacking response to interferon the impact on sofosbuvir sensitivity is minimal. The A150V polymorphism reduces the response to interferon 70 fold whereas the K206E substitution has minimal effects on interferon in isolation but in combination with A150V reduces the response 100 fold. Preliminary data indicates that the A150V polymorphism interferes with the late response to type I interferons enabling the virus to overcome the induction of interferon-stimulated genes. These data indicate a complex interaction between direct acting antiviral drugs and the innate antiviral response.
23
Pereira, Joana Duarte da Rocha. "Searching for antivirals agaist norovirus: 2-Styrylchromones as potential anti-norovirus agents." Master's thesis, Faculdade de Farmácia da Universidade do Porto, 2008. http://hdl.handle.net/10216/20864.
Full text
24
Pereira, Joana Duarte da Rocha. "Searching for antivirals agaist norovirus: 2-Styrylchromones as potential anti-norovirus agents." Dissertação, Faculdade de Farmácia da Universidade do Porto, 2008. http://hdl.handle.net/10216/20864.
Full text
25
VILLANI, ROSANNA. "Direct-acting antivirals (daas) increase the serum vegf level in patients with chronic hepatitis c: a rationale for tumor recurrence." Doctoral thesis, Università di Foggia, 2017. http://hdl.handle.net/11369/363292.
Full textAbstract:
The currently available novel combinations of DAAs have completely changed the panorama of hepatitis C therapy. As a result, current HCV infection cure rates have exceeded 90% in a very short time. Data on compensated cirrhosis show rates of sustained virological response (SVR) of around 95-97%, although this is a little lower in more advanced liver diseases.
However, some authors reported that hepatocellular carcinoma (HCC) seems to develop within weeks or few months of starting treatment with direct-acting antivirals (DAAs).
They reported for the first time that patients with cirrhosis and very early HCCs (single tumor <2 cm) or early HCCs with a low to moderate risk of recurrence showed a probability of recurrence at 4 months of between 17.6 and 21.5%; however, the recurrence rate was double the expected rate.
On the other hand, some authors considered it unlikely that DAAs have an effect on tumor recurrence. A French group analyzed 307 chronic HCV patients with HCC who were treated with DAAs in 3 different cohorts; they concluded that there was no increased risk of HCC recurrence.
It is well established that angiogenesis is a major driver of tumor dissemination and that VEGF is a critical player in liver cancer angiogenesis. VEGF levels in HCC tissues or in circulation correlate with more aggressive disease; therefore this research was aimed at monitoring the serum level of vascular endothelial growth factor (VEGF) and changes in the pattern of circulating interleukins in 103 chronic hepatitis C patients during antiviral treatment with DAA-regimens.
VEGF, epidermal growth factor (EGF), and interleukins (IL-6, IL-10, IL-8, TNF alpha) were assessed at baseline, during treatment, and after treatment.
The biological effect of DAA-treated patient serum on human umbilical vein endothelial cell (HUVEC) and Hepa RG cell proliferation in the presence of anti-VEGF monoclonal antibody, bevacizumab, was also confirmed.
The median log10VEGF was significantly increased at 4 weeks after initiation of therapy (from 2.18 log10 pg/mL to 2.45 log10 pg/mL, p<0.001), remaining higher at the end of treatment (2.44 log10 pg/mL, p<0.001 vs. T0), and decreasing to the baseline value upon treatment discontinuation (2.13 log10 pg/mL at SVR4 and 2.28 log10 pg/mL at SVR12)
On the other hand, EGF levels also increased during DAA treatment and subsequently decreased at SVR12; however, this trend was not significant.
The temporal trends of the 2 main inflammatory cytokines, namely IL-10 and TNF-alpha, were different. At 4 weeks, IL-10 significantly
decreased (from 1.44 pg/mL to 0.83 pg/mL; p=0.03), whereas TNF-alpha levels remained stable (from 4.05 pg/mL to 4.01 pg/mL, p=0.27). Interestingly, while IL-10 levels were substantially suppressed from the end of treatment onwards, the decrease in TNF-alpha started later; it was nearly significant at the end of treatment (2.82 pg/mL, p=0.05) and this was confirmed, although with a smaller decrease, at each consecutive time point (2.19 pg/mL and 2.09 pg/mL at SVR4 and 12, respectively). No difference in IL-8 and IL-6 levels was observed during treatment.
The biological activity of sera collected from patients treated with DAAs on human endothelial cells (HUVEC) and Hepa RG was also investigated.
HUVEC proliferation was measured upon stimulation with cell medium supplemented with sera obtained from patients receiving therapy.
A significant induction of HUVEC proliferation was observed upon supplementation of media with sera from patients receiving DAA therapy (mean: 290±60% vs. 120±30, p<0.0001). Moreover, consistent with the reduction of VEGF upon treatment withdrawal, HUVEC proliferation was comparable to baseline in cell cultures supplemented with sera collected 12 weeks after treatment interruption. Of note, the peak of HUVEC stimulation upon addition of patient sera was comparable to maximal HUVEC stimulation under standard culture conditions
The addition of serum from patients collected during therapy induced HepaRG proliferation that was partially inhibited (decrease of 50%)
by bevacizumab; the effect disappeared 4 weeks after the end of therapy (SVR4). These results suggest that additional factors may be involved in the cellular growth observed after supplementation with sera collected during antiviral therapy.
26
Farleigh, Laura Elizabeth. "Development of a new class of antivirals active against pox and measles viruses." Thesis, Cardiff University, 2014. http://orca.cf.ac.uk/63804/.
Full textAbstract:
In this PhD project we show for the first time that novel dideoxy bicyclic pyrimidine nucleoside analogues (ddBCNAs) with L-chirality represent promising antiviral candidates for use against pox and measles viruses. We suggest a mechanism of action based on a cellular target. Our lead compound (Cf2642, with side chain C9H18–O–C5H11) is active against vaccinia virus (a surrogate poxvirus for smallpox) and measles virus, with IC50 concentrations of 0.19 and 7.5 µM, respectively. This is a 60-fold enhancement over cidofovir (viral DNA polymerase inhibitor; IC50 of 11.5 µM against VACV). A structure activity relationship was established, which was similar for both viruses, indicating a common and specific mechanism of action. Cf2642 does not inhibit HSV-1/2, influenza, adeno or yellow fever viruses. The mechanism of action for the ddBCNAs has been investigated and, though not defined, has been narrowed down. Based on our observations of drug activity in cell lines derived from various sources, we have suggested a cellular target for the ddBCNAs, most likely cellular membrane compartments or the proteins located therein. Though inhibition of vaccinia is observed within two hours of infection, we have shown that the ddBCNAs are unlikely to be entry inhibitors. Acidification of the extracellular medium was observed but, whilst it may be linked to the mechanism of action, this is not the cause of the antiviral effects. With a possible cellular target, toxicity was carefully evaluated. We have not observed significant cytotoxicity in any of our cell models. Antivirals active against cellular targets are less subject to viral resistance, which may develop rapidly with virus-targeting drugs. This could be critical since, there are currently no effective measles antiviral drugs available on the market, and resistance to measles RNA polymerase inhibitors and the potential antipoxviral drug cidofovir has already been described.
27
Berry, Michael. "Massively-Parallel Computational Identification of Novel Broad Spectrum Antivirals to Combat Coronavirus Infection." University of the Western Cape, 2015. http://hdl.handle.net/11394/8321.
Full textAbstract:
Philosophiae Doctor - PhDGiven the significant disease burden caused by human coronaviruses, the discovery of an effective antiviral strategy is paramount, however there is still no effective therapy to combat infection. This thesis details the in silica exploration of ligand libraries to identify candidate lead compounds that, based on multiple criteria, have a high probability of inhibiting the 3 chymotrypsin-like protease (3CUro) of human coronaviruses. Atomistic models of the 3CUro were obtained from the Protein Data Bank or theoretical models were successfully generated by homology modelling. These structures served the basis of both structure- and ligand-based drug design studies. Consensus molecular docking and pharmacophore modelling protocols were adapted to explore the ZINC Drugs-Now dataset in a high throughput virtual screening strategy to identify ligands which computationally bound to the active site of the 3CUro . Molecular dynamics was further utilized to confirm the binding mode and interactions observed in the static structure- and ligand-based techniques were correct via analysis of various parameters in a IOns simulation. Molecular docking and pharmacophore models identified a total of 19 ligands which displayed the potential to computationally bind to all 3CUro included in the study. Strategies employed to identify these lead compounds also indicated that a known inhibitor of the SARS-Co V 3CUro also has potential as a broad spectrum lead compound. Further analysis by molecular dynamic simulations largely confirmed the binding mode and ligand orientations identified by the former techniques. The comprehensive approach used in this study improves the probability of identifying experimental actives and represents a cost effective pipeline for the often expensive and time consuming process of lead discovery. These identified lead compounds represent an ideal starting point for assays to confirm in vitro activity, where experimentally confirmed actives will be proceeded to subsequent studies on lead optimization.
28
Nilsson, Emma C. "Cellular receptors for viruses with ocular tropism." Doctoral thesis, Umeå universitet, Virologi, 2011. http://urn.kb.se/resolve?urn=urn:nbn:se:umu:diva-42818.
Full textAbstract:
Several viruses from different virus families are known to cause ocular infections, e.g. members of the Adenoviridae, Picornaviridae and the Herpesviridae families. These infections are spread by contact and in the case of adenoviruses (Ads) and picornaviruses they are also highly contagious. The ocular infections caused by Ads and picornaviruses are called epidemic keratoconjunctivitis (EKC) and acute hemorrhagic conjunctivitis (AHC), respectively. Historically, EKC is caused mainly by three types of Ads from species D: Ad8, Ad19 and Ad37. The infection is characterized by keratitis and conjunctivitis but also involves pain, edema, lacrimation and blurred vision. AHC is caused mainly by two types of picornaviruses: coxsackievirus A24v (CVA24v) and enterovirus 70 (EV70), and is characterized by pain, redness, excessive tearing, swelling and subconjunctival hemorrhages. In addition, blurred vision, keratitis, malaise, myalgia, fever, headache and upper respiratory tract symptoms can also be experienced. Both infections are problematic in many parts of the world, affecting millions of people every year. Despite the great need, the only treatment available today is supportive treatment; no antiviral drugs are available to combat these common viral infections. Ad37 has previously been reported to use sialic acid (SA) as its cellular receptor. Since there is no antiviral treatment available against EKC we wanted to evaluate the inhibitory effect of SA-based antiviral compounds on Ad37 binding to and infection of ocular cells. We found that multivalent compounds consisting of SA linked to a globular carrier molecule, in this case human serum albumin, efficiently blocked Ad37 binding and infection at low concentrations. Further attempts were then made to improve the effect by chemically modifying SA monosaccharides. However, no enhanced inhibitory effect was accomplished and the conclusion was that the best inhibitors are based on unmodified SA. We next hypothesized that development of efficient SA-based binding inhibitors may require detailed knowledge about the structure of the SA-containing receptor. Using a battery of biological and biochemical experiments, including glycan array, binding and infection assays, X-ray crystallography and surface plasmon resonance (SPR); we identified a specific glycan involved in the binding and infection of Ad37. This glycan turned out to be a branched, di-SA-containing motif corresponding to the glycan motif of the ganglioside GD1a. However, the ganglioside itself did not function as a cellular receptor, as shown by a number of binding and infection assays. Instead, the receptor consisted of one or more glycoproteins that contain the GD1a glycan motif. This glycan docked with both its SAs into the trimeric Ad37 knob resulting in a very strong interaction as compared to most other protein-glycan interactions. Hopefully, this finding will aid development of more potent inhibitors of Ad37 binding and infection. The receptor for CVA24v, one of the main causative agents of AHC, has been unknown until now. We showed that this ocular virus, like Ad37, is also able to use SA as a receptor on corneal cells but not on conjunctival cells. This suggested that CVA24v may use two different receptors. As for Ad37, the receptor used by CVA24v on corneal cells also appears to be one or more sialic acid-containing glycoproteins. We believe that these findings may be a starting point for design and development of candidate drugs for topical treatment of AHC.
29
Imhof, Ingrid. "Development of an intra- and intergenotypic HCV cell culture method to phenotype and assess antiviral susceptibilities and resistance development of HCV NS3 protease genes from HCV genotypes 1-6." Thesis, University of Edinburgh, 2010. http://hdl.handle.net/1842/6207.
Full textAbstract:
The development of specific antiviral drugs directly targeting the hepatitis C virus (HCV) is clinically important, as the current standard interferon/ribavirin combination treatment is only partially effective, expensive and often associated with severe side effects. Inhibitors of the NS3 protease (PI) therefore represent a promising alternative or additional therapy. To date, the development and in vitro evaluation of PIs is restricted to the genotype 1/2 based replicon and the genotype 2a full length viral cell culture system. However, proteases of the different HCV genotypes vary substantially in their amino acid sequence and secondary structure and require separate evaluation of their efficacy before they go into clinical trials. To address this issue, a panel of intra- and intergenotypic recombinants based on the recombinant infectious clone Jc1 (pFK JFH1/J6/C-846) was developed in this work. The viability of these recombinants was assessed in the Huh7.5 cell culture system, where replicating viruses were detected by HCV-NS5A immunostaining. Intergenotypic recombinants containing genotype 1a, 1b, 3a, 4a and 6a derived proteases were replication defective, whereas the recombinant with genotype 5a derived protease replicated efficiently after acquiring cell culture adaptive mutations. The replacement of not only the NS3 protease gene region, but also its cofactor NS4A, allowed the generation of replication competent intra- and intergenotypic recombinants for all 6 major genotypes. Replacing the NS3 protease of the recombinants with that of patientderived proteases also generated replicating recombinants, greatly expanding the panel of intergenotypic recombinants available for phenotyping and PI evaluation. However, intra- and intergenotypic recombinants showed substantial differences in their replication kinetics, which may be influenced by naturally occurring polymorphism between genotypes and the differential requirement of adaptive/attenuating cell culture mutations. Genotype 1a recombinants replicated very poorly, which may be due to incompatibilities between the type 1a NS3/4A protease and the type 2a backbone. 50% inhibitory concentrations (IC50) of different PIs were measured using Foci Forming Units/ml (FFU/ml) reductions and replication inhibition assays. The different recombinants showed consistent, genotype-associated differences in their susceptibility to the PI BILN 2061, with genotypes 2a, 3a and 5a derived recombinants showing approximately 100-fold lower susceptibility than genotype 1b, 4a and 6a derived recombinants. These observations are consistent with major differences in response rates found in recent treatment trials of genotype 1, 2 and 3 infected patients. Differences in susceptibility were also observed for VX-950, with genotype 1b, 2a and 6a derived recombinants being twice as susceptible than genotype 3a, 4a and 5a derived recombinants. Passaging the intra- and intergenotypic recombinants under increasing concentrations of PI allowed the identification of PI resistance mutations. Resistance mutations to BILN 2061 mapped to the previously identified positions 156 and 168 within the NS3 protease, with a great diversity of amino acid substitutions observed within each genotype. Reintroduction of the identified resistance mutations into the original recombinant viruses conferred increased resistance towards BILN 2061 and some mutations also affected replication kinetics of the recombinants. The developed system will be of major value for the phenotypic characterisation of naturally occurring and treatment induced resistance mutations within all 6 major HCV genotypes towards different PIs. This will allow treatment response predictions for newly developed PIs before they enter clinical trials and the development of individually tailored antiviral treatment regimes.
30
LUCIA, FALSITTA. "DDX3, a new frontier in broad-spectrum antiviral therapy: synthesis of potential inhibitors." Doctoral thesis, Università di Siena, 2020. http://hdl.handle.net/11365/1095615.
Full textAbstract:
Viral infections inflict many serious human diseases with very high mortality rates. New drug-resistant strains are continually emerging due to the high viral mutation rate, which makes necessary to develop novel potent antivirals.
Targeting cellular cofactor essential for the replication of different viruses but not for the cells represents a new strategy to combat infectious diseases and offers a higher genetic barrier to the development of the resistance.
The DEAD-box RNA helicase DDX3 is a multifunctional protein involved in many aspects of RNA metabolism, including transcription, splicing, mRNA nuclear export, translation, RNA decay and ribosome biogenesis.
DDX3 is a human host factor required for the replication of several DNA and RNA such as herpes virus, human immunodeficiency virus type 1, hepatitis C virus, Dengue virus and West Nile virus.
Given the multifaceted functions of DDX3, this host factor represents a promising target to develop compounds with broad spectrum antiviral activity.
In the last few years Prof. Botta’s research group has been identified several inhibitors of DDX3 proteins. From a medicinal chemistry point of view, DDX3 has multiple enzymatic activities, ATPase and RNA helicase, and functional domains that may be targeted by potential inhibitors.
Prof. Botta’s research group, designed and validated the first small molecule DDX3 inhibitors specifically designed to target its RNA binding site (16d with anti-helicase activity against DDX3 IC50 = 0.3 μM). Pursuing this research line, a structure-based optimization process was prosecuted, resulting in the identification of a novel compound with the 1,2,4-oxadiazole nucleus UVR40, with anti-helicase activity against DDX3 IC50 = 0.13 μM. Thus, a small library of UVR40 derivatives has been designed by our computation group and synthesized during my PhD with the purpose to enlarge SAR knowledge, enhance its ADME properties and improve its activity profile.
At the same time, with the attempt to enlarge our library of DDX3 inhibitors, Prof. Botta’s group built a novel library of “hybrid” compounds starting from the structures of two hit compounds previously discovered, 16d member of the urea series, and UVR06 characterized by a sulfonamide moiety.
The novel library was synthesized, validated on the target enzyme, and evaluated against the West Nile virus (WNV) infection.
31
Mak, Ka-ki Peter. "The potential trade-offs between treatment and prophylaxis with antivirals in households during a pandemic." View the Table of Contents & Abstract, 2007. http://sunzi.lib.hku.hk/hkuto/record/B38479631.
Full text
32
Mak, Ka-ki Peter, and 麥家麒. "The potential trade-offs between treatment and prophylaxis with antivirals in households during a pandemic." Thesis, The University of Hong Kong (Pokfulam, Hong Kong), 2007. http://hub.hku.hk/bib/B39724517.
Full text
33
王軼 and Yi Jennifer Wang. "The optimal allocation of investment between antivirals and vaccines for influenza pandemic preparedness planning." Thesis, The University of Hong Kong (Pokfulam, Hong Kong), 2008. http://hub.hku.hk/bib/B41712018.
Full text
34
Ng, Sophia, and 吳鈺陪. "The role of antivirals and vaccines in the control of influenza epidemics and pandemics." Thesis, The University of Hong Kong (Pokfulam, Hong Kong), 2012. http://hub.hku.hk/bib/B49617849.
Full textAbstract:
Influenza vaccination is the best preventive measure against influenza virus infection, and antivirals including oseltamivir are effective treatments. From a public health point of view, it is important to evaluate whether vaccination and antiviral treatment reduces transmission of the virus. I analyzed data from a community-based study of influenza virus transmission in households, and identified effectiveness of antiviral treatment in reducing duration of illness and some evidence that treatment reduced transmission to household contacts. I also analyzed data from a community-based placebo-controlled trial of influenza vaccination and confirmed efficacy of vaccination against seasonal influenza but differential efficacy against pandemic influenza possibly because of timing and mediation of seasonal influenza epidemics. In further analyses I found that antibody titers of 1:40 correlated with 50% protection against infection, and repeated vaccination with the same strains tended to be associated with reduced responses to those strains although there was no evidence of reduced efficacy. In the study, one child in each household was randomly allocated to receive vaccine or placebo and I did not identify any evidence of indirect benefits to the household members of vaccinated children. I reviewed vaccine target groups in different countries, and noted that some countries now include school-age children in their target groups based mainly on the principle of herd immunity. My findings did not support the inclusion of school-age children as a target group for vaccination in Hong Kong. Further studies should examine the indirect as well as direct benefits of vaccination in different settings in order to guide optimal influenza vaccination policies.published_or_final_version
Community Medicine
Doctoral
Doctor of Philosophy
35
Wang, Yi Jennifer. "The optimal allocation of investment between antivirals and vaccines for influenza pandemic preparedness planning." Click to view the E-thesis via HKUTO, 2008. http://sunzi.lib.hku.hk/hkuto/record/B41712018.
Full text
36
MAGRI, ANDREA. "Exploration of new uracil-based compounds as novel inhibitors of Hepatitis C Virus replication." Doctoral thesis, Università del Piemonte Orientale, 2016. http://hdl.handle.net/11579/115181.
Full textAbstract:
Hepatitis C Virus (HCV) is a major public health problem worldwide. While highly efficacious directly-acting antiviral agents have been developed in recent years, their high costs and relative inaccessibility make their use limited. In this thesis, new uracil-based compounds have been evaluated as potential antiviral drugs against HCV. Using several biochemical and virological assays to investigate virus infection and replication, it has been shown that these compounds are able to significantly reduce viral genomic replication with their IC50 values in the nanomolar range. Finally, these compounds have been shown to block the de novo RNA synthesis and that effect is dependent on a chemical structure of the compounds.
37
Aillot, Ludovic. "Effets antiviraux de l'agonisation des Toll-like Récepteurs dans les cellules du foie, une nouvelle stratégie immunothérapeutique dans la lutte contre HBV." Thesis, Lyon, 2018. http://www.theses.fr/2018LYSE1139/document.
Full textAbstract:
Le virus de l'hépatite B (HBV) infecte chroniquement près de 240 millions d'individus dans le monde. L'infection chronique par HBV est un souci de santé publique majeur puisque l'infection peut évoluer au cours du temps vers la cirrhose et/ou l'hépatocarcinome (CHC). Malgré l'existence de traitements efficaces à base d'analogues de nucléos(t)ides permettant de diminuer la charge virale chez les patients, ceux-ci nécessitent une prise médicamenteuse à vie. En effet, malgré la diminution importante du risque de développer un cancer du foie, ces traitements ne permettent pas l'élimination définitive du virus. Les cellules infectées par HBV sont les hépatocytes du foie, qui remplissent la majorité des rôles vitaux de cet organe. La formation d'un minichromosome viral au sein de ces cellules infectées appelés ADNccc (pour ADN circulaire-covalemment-clos), est majoritairement responsable de la persistance du HBV. Les traitements actuels utilisés sont principalement des analogues de nucléos(t)ides et ceux-ci n'ont pas ou peu d'effets sur l'ADNccc. La nécessité de développer de nouvelles stratégies antivirales visant à éliminer définitivement HBV a donc conduit de nombreux laboratoires, dont le nôtre, à étudier l'utilisation de stratégies immuno-thérapeutiques incluant des stimulateurs de l'immunité innée (agonistes de TLR7, TLR8, RIG-1.) dans le cadre d'infections chroniques. De nombreuses études ont démontré que l'utilisation de ligands stimulant les récepteurs de l'immunité innée promouvait un fort effet antiviral, médié par la production endogène et locale de cytokines pro-inflammatoires et l'induction de gènes régulés par l'interféron (1SG). Dans ce but, nous nous sommes intéressés plus particulièrement aux potentiels effets antiviraux de l'agonisation des senseurs de l'immunité innée les plus connus, les Toll-like récepteurs (TLR), dans le cadre de l'infection par HBV dans les cellules hépatiques. La stratégie immuno-thérapeutique envisagée, vise à stimuler aussi bien les cellules immunitaires que les hépatocytes infectés. La caractérisation de l'expression de différents senseurs de l'immunité innée, d'une part dans les cellules primaires isolées du foie et d'autre part dans certaines lignées cellulaires correspondantes, nous a permis d'avoir une vue d'ensemble 1) des récepteurs exprimés par les différentes cellules du foie notamment dans les hépatocytes (TLR2/TLR3/TLR4/TLR5) ; 2) d'évaluer la fonctionnalité de ceux-ci pour la production de cytokines (IL-6 ; IP-10) lors de leur agonisation 3) d'évaluer les modèles disponibles parmi les lignées cellulaires les plus proches immunologiquement des cellules hépatiques. Les cellules HepaRG et une nouvelle lignée dérivée des macrophages du foie les iKC par exemple sont plus proches respectivement des hépatocytes et des macrophages primaires hépatiques et sont donc des modèles relevant pour les études immuno-thérapeutiques. L'utilisation de ligands de TLR2 et TLR3 sur des hépatocytes infectés chroniquement par HBV, a montré le plus fort effet antiviral (incluant une médiation par la sécrétion de cytokines et l'induction d'1SG) aussi bien sur la réplication d'HBV que sur l'ADNccc. De plus, cet effet semble stable au cours du temps sans résurgence massive de productions virales. Cette stratégie cible non seulement les hépatocytes infectés, mais également les cellules immunitaires dont les productions cytokiniques ont également un fort effet antiviral. Bien que l'effet in vivo, dans un modèle murin, ait été plus modeste, un ajustement des doses d'agonistes utilisées ainsi qu'un meilleur moyen de délivrance au foie de ligands de TLR2 ou TLR3 pourraient être une stratégie immuno-thérapeutique intéressante. Enfin nous nous sommes intéressés au cas particulier de l'agonisation du TLR9 en présence d'HBV… [etc]HBV chronically infects 240 million peoples around the world. HBV chronic infection is a major public health problem and can lead to cirrhosis or/and hepatocarcinoma (HCC). Even if some efficient treatments are already available, based in particular on the use of nucleos(t)ides analogues that induce a decrease of viral load in patients, these drugs do not lead to a definitive HBV cure They enable an important decrease of liver cancer risk but need to be taken life-long. HBV infects hepatocytes the major liver cells which are involve in many vital mechanisms into the organism. The HBV minichromosome, which is formed into infected cells also called cccDNA (i.e., covalently-closed-circular DNA), is not affected by nucleos(t)ides treatments and thus is responsible for HBV persistence. The use of immune receptors (e.g. Toll-like receptors/TLR) agonists can lead to 1) an important cytokines/interferon (IFN) secretion; 2) promote immune cells activation/recruitment and 3) induction of many Interferon-Stimulated Genes (ISG). These mechanisms could lead to a greater viral clearance by cccDNA degradation or silencing. The need for new strategies to permanently eliminate HBV infection led many laboratories, including ours, to explore the use of immunotherapeutic treatments in a context of chronic infection, including innate immune stimulators (e.g. TLR7, TLR8 or RIG-I agonist are under clinical trials). To this end, we got interested on the potential anti-HBV effects of many TLR agonists in liver cells. Our strategy is to stimulate both infected hepatocytes and immune cells. We first characterized the expression of innate immune sensors in primary liver cells as well as in some liver cell lines. This allowed us to: 1) identify which sensors are expressed by liver cells, especially in hepatocytes (TLR2, TLR3, TLR4, TLR5); 2) evaluate their ability to produce cytokines (IL-6, IP-10) upon agonisation; 3) evaluation of cell lines model which are immunologically closed to the primary liver cells. HepaRG and a new liver macrophage cell line call iKC are immunologically close to their primary cells and appear to be relevant models for immune-therapeutics studies. The use of TLR2 and TLR3 agonists on HBV chronically infected hepatocytes showed a strong antiviral effect (i.e., decrease of HBV replication and cccDNA level) mediated directly by NF- kB-inducible and ISG genes activation and indirectly by cytokines secretion. Furthermore, this effect was shown stable over time without any viral replication rebound. This strategy targets not only infected hepatocytes but also immune cells, whose cytokines production also has a strong antiviral effect. Despite a weak in vivo effect in mice, a tuning in agonist doses used and better liver delivery could be an interesting immune-therapeutic strategy. Finally, we were investigated the particular case of TLR9 agonisation in presence of HBV. We showed an interaction between synthetic or not DNA ligands such as CpG ODN and HBV particles. This interaction leads in one hand, to HBV entry inhibition in hepatocytes, on the other hand, to a blockage of ligand delivery to TLR9 in pDC, which is not due to an inhibition of the TLR9 pathway, but to a lack of access of the ligand to its receptor. These two mechanisms are responsible for a decrease of viral infection during its establishment and a decrease in IFN synthesis by pDC, respectively. A decrease in IFN production, which this time was linked to a bona fide inhibition of the TLR9 pathway, in the presence of the sub-viral particles HBsAg was still observed, without retention of TLR9 ligand of the latter. It would seem, therefore, that use of TLR agonists represent an interesting strategy in setting up new anti-HBV immune-therapeutic approaches. However, their improvement will depend on the evaluation of viro-induced inhibitory mechanisms as well as better ways of in vivo delivering these ligands
38
Amorós, Reboredo Patrícia. "Tractament de l'hepatitis C crònica amb agents antivirals directes en pacients majors de 65 anys." Doctoral thesis, Universitat de Barcelona, 2020. http://hdl.handle.net/10803/671482.
Full textAbstract:
ANTECEDENTS DEL TEMA
En els últims anys els agents antivirals directes aprovats pel tractament de l’hepatitis C han permès l’accés al tractament a grups de pacients que anteriorment no es tractaven pel risc que comportava l’ús d’esquemes amb interferó, incloent als pacients d’edat avançada. Es desconeix si l’alta prevalença de comorbiditats i la polimedicació dels pacients d’edat avançada poden estar associades a més esdeveniments adversos i a una disminució en l’eficàcia del tractament, ja que els pacients majors de 65 anys no es veuen representats en els assaigs clínics. És conegut que els pacients d’edat avançada acostumen a tenir un risc superior de reaccions adverses a medicaments, entre elles les que presenten efectes anticolinèrgics. Per aquest motiu diversos anticolinèrgics s’han llistat com a medicaments potencialment inadequats en pacients geriàtrics i actualment existeixen diverses escales per mesurar la càrrega anticolinèrgica que mostren diferències en els medicaments que inclouen, la puntuació i la potència anticolinèrgica que atribueixen a cada medicament.
HIPÒTESI DE TREBALL
Reconèixer la influència de les comorbiditats i la tolerància als possibles esdeveniments adversos dels nous tractaments amb antivirals d’acció directa pot ajudar en l’optimització del tractament de l’hepatitis C en aquests pacients.
Els pacients geriàtrics que inicien tractament per l’hepatitis C poden estar rebent altres medicaments amb efectes anticolinèrgics que poden influir en la seguretat del tractament. Un enfocament multidisciplinar amb la presència d’un farmacèutic clínic a l’equip pot ajudar a millorar l’atenció als pacients geriàtrics que inicien tractament per l’hepatitis C.
OBJECTIUS
Avaluar l’impacte de les comorbiditats i les interaccions farmacològiques en l’eficàcia i la tolerabilitat dels antivirals d’acció directa pel tractament de l’hepatitis C crònica en la població geriàtrica.
Determinar la prevalença de càrrega anticolinèrgica, relacionada amb medicació crònica, en pacients d’edat avançada tractats amb antivirals d’acció directa, i els factors de risc associats, així com analitzar les conseqüències en la seguretat dels tractaments.
METODOLOGIA
Estudi retrospectiu observacional en un hospital universitari de tercer nivell. Mitjançant el programa informàtic utilitzat per a la prescripció i preparació de medicaments es van identificar tots els pacients que havien estat tractats amb antivirals de acció directa associats o no a ribavirina, al centre, des d’abril de 2015 a març del 2016. Es van recollir variables clíniques i demogràfiques, incloent la presència de comorbiditats. També els factors virològics i clínics relacionats amb la infecció pel virus de l’hepatitis C. Les comorbiditats es van analitzar segons els valors de Clinical Risk Group, que classifiquen als pacients segons la presència de malalties cròniques. Es van recollir variables farmacològiques, com el nombre de medicaments concomitants, el tipus de medicament, i es va classificar als pacients segons el grau d’exposició a polifarmàcia (entesa com la presa de cinc o més medicaments), així com es va considerar el risc d’interaccions farmacològiques. La presència de càrrega anticolinèrgica es va valorar amb tres escales validades diferents: l’Anticholinergic Cognitive Burden scale (ACB), l’Anticholinergic Risk Scale (ARS) i l’Anticholinergic Drug Scale (ADS). L’eficàcia dels tractaments es va avaluar segons la resposta viral sostinguda la setmana 12 després de finalitzar el tractament, classificant els pacients en tres grups d’edat (65 a 74, 75 a 79 i ≥80 anys). L’anàlisi en seguretat va incloure dades especificades a la història clínica i les notificacions d’esdeveniments adversos registrades a la mateixa. Els resultats en seguretat es van analitzar d’acord amb la presència o no de medicaments anticolinèrgics, cirrosi, alta comorbiditat i interaccions farmacològiques.
RESULTATS
Es van identificar 261 pacients, amb una edat promig de 71 anys, sent el 61% dones. La prevalença de cirrosi era alta (74%) i el genotip 1b el predominant (n=232; 89%). Un elevat nombre de pacients (n=126; 48%) havia fracassat a un tractament anterior amb interferó i 22 pacients (8%) havien rebut un trasplantament de fetge prèviament. El 90% dels pacients prenien medicació concomitant. La resposta viral sostinguda global va ser del 96.9%, sense diferències entre les cohorts d’edat. Tampoc es van observar diferències en eficàcia tenint en compte la presència de cirrosi o haver rebut un tractament previ. El 86% dels pacients va presentar algun esdeveniment advers, majoritàriament fatiga i símptomes gastrointestinals o dèrmics. Tots els pacients que van presentar algun esdeveniment advers greu (6.5%) tenien una alta comorbiditat associada. Entre els pacients que prenien almenys un medicament crònic (90%), es va observar una elevada presència de polifarmàcia, amb un 47% d’aquests prenent cinc o més fàrmacs concomitants. Els fàrmacs més observats com a medicació crònica van ser els diürètics i els psicolèptics. Un elevat nombre de pacients presentava risc d’interaccions farmacològiques tot i que aquestes no es van relacionar amb pitjors resultats terapèutics. Pel que fa a la prevalença de càrrega anticolinèrgica, els resultats van ser diferents segons l’escala de mesura utilitzada essent similars les escales ACB i ADS (35.2% i 34.3%) i lleugerament superiors respecte a l’escala ARS (10.6%). Per a totes les escales es va fer palesa la relació entre comorbiditat i presència de medicació amb càrrega anticolinèrgica, posant de manifest que els pacients amb més comorbiditat presenten més risc de rebre tractaments amb efectes anticolinèrgics. En la present tesi les tres escales valorades han mostrat resultats diferents i només l’escala Anticholinergic Risk Scale (ARS) ha mostrat relació significativa entre càrrega anticolinèrgica i esdeveniments adversos.
CONCLUSIONS
Els antivirals d’acció directa són efectius i ben tolerats en pacients majors de 65 anys. Els pacients geriàtrics que inicien tractament per l’hepatitis C són pacients complexes i amb múltiples comorbiditats que reben altres medicaments, alguns dels quals amb efectes anticolinèrgics, que poden influir en la seguretat del tractament. L’eficàcia dels tractaments no s’ha vist influenciada per l’edat, ni per la presència o absència de cirrosi, així com tampoc per haver rebut anteriorment tractament antiviral. El nombre d’esdeveniments adversos greus augmenta amb la comorbiditat i el nombre de medicaments concomitants associats. No es poden extreure conclusions pel que fa a quina mesura de càrrega anticolinèrgica representa el millor valor pronòstic. Per tal d’oferir un tractament antiviral òptim, la coordinació entre els hepatòlegs i els farmacèutics clínics recolzats en un equip multidisciplinari és necessària.
39
Revuelto, Artigas Tamara. "Ateromatosis subclínica en pacientes con infección crónica por virus de la Hepatitis C: Factores de riesgo y modificación tras la terapia con Antivirales de Acción Directa." Doctoral thesis, Universitat de Lleida, 2019. http://hdl.handle.net/10803/667849.
Full textAbstract:
OBJECTIU: Analitzar si la infecció VHC és un factor de risc independent d`ateromatosi subclínica i conèixer les característiques que influeixen en la composició de les plaques, així com la seva modificació després de 12 mesos de la teràpia amb Antivirales d'acció directa (AAD). MATERIAL I MÈTODES: Estudi prospectiu que compara 185 pacients VHC amb diferents genotips i fibrosi hepàtica abans d'AAD, davant de 411 subjectes sense infecció, amb similar risc cardiovascular. L`ateromatosi subclínica (GIM i presència de plaques) i composició de la placa (programari HEMODYN 4) es va avaluar amb ecografia en territori carotidi i femoral a l'inici i després de 12 mesos de AAD. RESULTATS: Es va detectar major GIM (0,83 vs 0,73mm; p = 0,045), placa d'ateroma (63% vs 44%; p <0,001) amb composició lipídica (50% vs 29%; p <0,001) en els pacients VHC que en controls. El percentatge de lípids es va associar amb edat i VHC (p <0,001) .Els factors de risc de ateromatosi van ser la infecció VHC (OR = 2,64), el sexe masculí (OR = 2,79), l'edat (OR = 1,08, amb un RR = 3,11 en pacients infectats menors de 55anys), el tabaquisme (OR = 3,25), la tensió arterial (OR = 1,02) i l'índex de insulinoresistència (TiG, OR = 3 , 18). Pel que fa a les característiques virals, només va influir el genotip (OR = 2,46, amb un risc de placa G2 45,4%, G1 55,3%, G4 78,8% i G3 94,4%) independent de la fibrosi hepàtica. Després de 12 mesos de la resposta viral sostinguda amb AAD, avaluem 85 pacients sense detectar modificació del GIM (0,74 vs 0,81mm; p = 0,068) ni a la placa (66% vs el 72%; p = 0,063). En la composició, es va observar una lleu tendència a la disminució de lípids no significativa (49,5 vs 47%; p = 0,305). Després d'aquest període, vam detectar una millora de l'esteatosi i fibrosi hepàtica, però un augment dels nivells sèrics de colesterol (p <0,001). CONCLUSIONS: La infecció crònica per VHC és factor de risc independent d`ateromatosi subclínica accelerada amb plaques predominantment lipídiques, l'ecografia arterial és un mètode no invasiu per a l'avaluació del risc cardiovascular. Després de 12 mesos de l'eradicació del VHC amb AAD no millora l'ateromatosi globalment ni en ajustar per factors vasculars o severitat fibrosi hepàtica.OBJETIVO: Analizar si la infección VHC es un factor de riesgo independiente de ateromatosis subclínica y conocer las características que influyen en la composición de las placas, así como su modificación tras 12 meses de la terapia con Antivirales de acción directa (AAD). MATERIAL Y MÉTODOS: Estudio prospectivo que compara 185 pacientes VHC con diferentes genotipos y fibrosis hepática antes de AAD, frente a 411 sujetos sin infección, con similar riesgo cardiovascular. La ateromatosis subclínica (GIM y presencia de placas) y composición de la placa (software HEMODYN 4) se evaluó con ecografía en territorio carotídeo y femoral al inicio y tras 12 meses de AAD. RESULTADOS: Se detectó mayor GIM (0,83 vs 0,73mm; p=0,045), placa de ateroma (63% vs 44%; p<0,001) con composición lipídica (50% vs 29%; p<0,001) en los pacientes VHC que en controles. El porcentaje de lípidos se asocio con edad y VHC (p<0,001).Los factores de riesgo de ateromatosis fueron la infección VHC (OR=2,64), el sexo masculino (OR=2,79), la edad (OR=1,08, con un RR=3,11 en pacientes infectados menores de 55años), el tabaquismo (OR=3,25), la tensión arterial (OR=1,02) y el índice de insulinorresistencia (TyG, OR=3,18). Respecto a las características virales, solo influyó el genotipo (OR=2,46, con un riesgo de placa G2 45,4%, G1 55,3%, G4 78,8% y G3 94,4%) independiente de la fibrosis hepática. Tras 12meses de la respuesta viral sostenida con AAD, evaluamos 85 pacientes sin detectar modificación del GIM (0,74 vs 0,81mm; p=0,068) ni en la placa (66% vs 72%; p=0,063). En la composición, se observó una leve tendencia a la disminución de lípidos no significativa (49,5 vs 47%; p=0,305). Tras este periodo, detectamos una mejoría de la esteatosis y fibrosis hepática, pero un aumento de los niveles séricos de colesterol (p<0,001). CONCLUSIONES: La infección crónica por VHC es factor de riesgo independiente de ateromatosis subclínica acelerada con placas predominantemente lipídicas, la ecografía arterial es un método no invasivo para la evaluación del riesgo cardiovascular. Tras 12meses de la erradicación del VHC con AAD no mejora la ateromatosis globalmente ni al ajustar por factores vasculares o severidad de fibrosis hepática.
OBJECTIVE: To analyze whether HCV infection is an independent risk factor for subclinical atheromatosis and to know the characteristics that influence the composition of the plaques, as well as its modification after 12 months of direct action antiviral therapy (DAA). MATERIAL AND METHODS: Prospective study comparing 185 HCV patients with different genotypes and liver fibrosis before AAD, compared to 411 subjects without infection, with similar cardiovascular risk. Subclinical atheromatosis (IMT and presence of plaques) and plaque composition (software HEMODYN 4) was evaluated with ultrasound in the carotid and femoral territory at the beginning and after 12 months of DAA. RESULTS: Higher MIC was detected (0.83 vs 0.73 mm, p = 0.045), atheroma plaque (63% vs 44%, p <0.001) with lipid composition (50% vs 29%, p <0.001) in the HCV patients than in controls. The percentage of lipids was associated with age and HCV (p <0.001). The risk factors for atheromatosis were HCV infection (OR = 2.64), male sex (OR = 2.79), age (OR = 1.08, with RR = 3.11 in infected patients under 55 years of age), smoking (OR = 3.25), blood pressure (OR = 1.02) and the insulin resistance index (TyG, OR = 3 , 18). Regarding the viral characteristics, only the genotype influenced (OR = 2.46, with a risk of G2 plaque 45.4%, G1 55.3%, G4 78.8% and G3 94.4%) independent of fibrosis hepatic After 12 months of the sustained viral response with DAA, we evaluated 85 patients without detecting a change in the IMT (0.74 vs 0.81 mm, p = 0.068) or in the plaque (66% vs 72%, p = 0.063). In the composition, there was a slight tendency to decrease lipids not significant (49.5 vs 47%, p = 0.305). After this period, we detected an improvement in steatosis and liver fibrosis, but an increase in serum cholesterol levels (p <0.001). CONCLUSIONS: Chronic HCV infection is an independent risk factor for accelerated subclinical atheromatosis with predominantly lipid plaques; arterial ultrasound is a non-invasive method for evaluating cardiovascular risk. After 12 months of eradication of HCV with DAA, atheromatosis does not improve globally nor does it adjust for vascular factors or severity of liver fibrosis.
40
Ferreira, Roberta Costa Santos. "Avaliação da atividade antirretroviral de produtos naturais." Universidade Federal de Alagoas, 2010. http://www.repositorio.ufal.br/handle/riufal/2520.
Full textAbstract:
This study aimed to evaluate the antiretroviral activity of natural products. Initially
implemented the cell culture for cytotoxicity tests and some tests of antiviral activity. Were selected based on pre-existing data in the literature of antiviral activity in the genus or family under study and also in a chemotaxonomic approach, 39 extracts (from 12 plant families and four families of Brazilian algae) and 11 pure compounds from the group of quinones and terpenes. All were evaluated for cytotoxicity in order to find a concentration with low cytotoxicity for testing antivirals ( 50%). Cytotoxicity was evaluated by the methods of Trypan Blue exclusion and MTT reduction. The evaluation of antiviral activity was done by two methods: research on the inhibition of cytopathic effects (ECPs) characteristic of retroviruses (syncytium and lysis) using maedi visna virus which is a model in vitro and in vivo of the HIV, and inhibition of activity of reverse transcriptase (RT) of HIV-1. The activity of RT HIV-1 was measured by a colorimetric method quantitative immunoassay (Reverse Transcriptase Assay, Roche , Germany). Was first tested an extract of each plant species on the potential antiviral activity, and when it showed some activity other extracts of the same species were tested. Efavirenz was used as control and showed antiviral inhibition of CPE and TR of 75 and 98%, respectively, at a concentration of 1 μg/mL. Among the pure compounds the best results were those of emotinas D and F, which also inhibit the ECPs in 37.5 and 25% were 24.4 and 20.5% inhibition of TR, respectively, 1 μg/mL. Among the tested plants found seven with inhibitory activity of HIV-1 RT. Among these we find a plant whose activity was very high in the crude extract when compared with control efavirenz and has been even greater in isolated fractions. The crude extracts of leaves (50 μg/mL) and stem bark (100 μg/mL) showed 98 and 67% inhibition of TR, respectively. The acetate (50 μg/mL) and chloroform (100 μg/mL) fractions of the latter, showed 95 and 89% of inhibition, respectively. Our results suggest that we have strong candidates for to combat the HIV that may present as a rich source of inhibitors of RT.Este trabalho objetivou avaliar a atividade antirretroviral de produtos naturais. Inicialmente implementamos o cultivo celular para realização de testes de citotoxicidade e alguns dos testes de atividade antiviral. Foram selecionados, baseados em dados pré-existentes na literatura de atividade antiviral no gênero ou família em estudo e ainda em uma abordagem quimiotaxonômica, 39 extratos (de 12 famílias de plantas e quatro famílias de algas brasileiras) e 11 substâncias puras do grupo das quinonas e dos terpenos. Todos foram avaliados quanto à citotoxicidade a fim de encontrarmos uma dose com baixa citotoxicidade para os testes antivirais (50%). A citotoxicidade foi avaliada pelos métodos de exclusão do Azul de Tripan e redução do MTT. A avaliação da atividade antiviral foi feita por duas metodologias: pesquisa da inibição de efeitos citopáticos (ECPs) característicos de retrovírus (sincício e lise) utilizando-se o vírus maedi visna que é um modelo in vitro e in vivo do HIV e inibição da transcriptase reversa (TR) do HIV-1. A atividade da TR do HIV-1 foi medida por um método colorimétrico quantitativo imunoenzimático (Reverse Transcriptase Assay, Roche , Germany). Foi inicialmente testado um extrato de cada espécie de planta quanto à potencial atividade antiviral e quando o mesmo mostrava alguma atividade os outros extratos da mesma espécie eram testados. O efavirenz foi utilizado como controle antiviral e apresentou 75 e 98% de inibição do ECP e da TR, respectivamente, em uma concentração de 1 μg/mL. Dentre as substâncias puras os melhores resultados encontrados foram os das emotinas D e F que além de inibirem os ECPs em 37,5 e 25%, apresentaram 24,4 e 20,5% de inibição da TR, respectivamente, a 1 μg/mL. Entre as plantas testadas encontramos sete com atividade inibidora da TR do HIV-1. Entre estas encontramos uma planta cuja atividade foi muito elevada no extrato bruto quando comparada ao controle efavirenz e tem se mostrado ainda maior nas frações isoladas. Os extratos brutos das folhas (50 μg/mL) e da casca do caule (100 μg/mL) apresentaram 98 e 67% de inibição da TR, respectivamente. E as frações acetato (50 μg/mL) e clorofórmio (100 μg/mL) deste último, 95 e 89% de inibição, respectivamente. Nossos resultados sugerem que temos fortes candidatos para o combate ao HIV que podem se apresentar como uma rica fonte de substâncias inibidoras da TR.
41
Egmond, Elfi. "Health-related quality of life and risk factors in hepatitis C patients treated with direct-acting antivirals." Doctoral thesis, Universitat Autònoma de Barcelona, 2017. http://hdl.handle.net/10803/458523.
Full textAbstract:
El virus de la hepatitis C (VHC) causa una de las infecciones crónicas más importantes, afectando a una población estimada de 71 millones de personas. El tratamiento antiviral clásico con (peg)interferón-alfa y ribavirina (PR) provoca un deterioro en la calidad de vida relacionada con la salud (CVRS) de los pacientes con hepatitis C crónica (HCC). Recientemente, se han introducido antivirales de acción directa (AAD) que se han asociado a una mayor ratio de respuesta al tratamiento, una reducción de los efectos secundarios y un impacto mínimo relacionada con la CVRS. Sin embargo, las evidencias aún son escasas debido a que los ensayos clínicos investigando los AAD están en desarrollo.
Para esta tesis doctoral, se han llevado a cabo dos estudios: (I) una revisión sistemática y un meta-análisis de ensayos clínicos randomizados (ECR) que han evaluado la CVRS y los factores de riesgo que podrían predecir su deterioro en pacientes en su deterioro con HCC tratados con AAD; (II) un estudio de cohorte naturalístico longitudinal con el fin de evaluar la CVRS y la incidencia de depresión durante el tratamiento antiviral, teniendo en cuenta posibles factores de riesgo para un deterioro en la CVRS y la aparición de depresión.
Los resultados de la revisión sistemática sugieren que los nuevos regímenes antivirales tienen un impacto mínimo en la CVRS, e incluso pueden mejorar el componente de salud mental a 12 semanas de postratamiento (MD=2.88; 95%CI=2.24, 3.53). La co-administración de ribavirina a los AAD mostró deterioro en el componente mental (MD=-1.7; 95%CI=-2.5, -0.91). Cualquier combinación de AADs con PR empeoró la CVRS tanto en el componente mental como físico (MD= -0.13; 95%CI=-0.15, -0.11). A nivel basal, la CVRS fue menor en pacientes con VHC sin empleo, en aquellos con cirrosis, anemia, edad avanzada, o con antecedentes de depresión o ansiedad. Además, el género femenino, la edad avanzada, y los antecedentes de depresión pudieron predecir una menor CVRS durante AAD. Asimismo, los acontecimientos adversos y la no-respuesta al tratamiento con AAD y PR fueron factores de riesgo.
En el segundo estudio, se observó una incidencia acumulada de depresión mayor en el 13.7% (95%CI: 5.7 - 26.3), y de cualquier trastorno depresivo en el 51% (95%CI: 36.6 - 65.2) durante la administración de AAD. El análisis de regresión logístico multivariado mostró la puntuación a nivel basal del PHQ-9 (p=0.002) como predictor para incidencia de depresión mayor, con una tendencia para historia familiar de depresión (p=0.079). Además, los pacientes con cirrhosis descompensada fue predictor para experimentar más dolor y disconfort durante el tratamiento (p=0.045). No se pudo descartar la presencia de cambios significativos de la media (DS) en puntuaciones del EQ-VAS durante el tratamiento antiviral con AAD (67.2±20.3), al final de tratamiento (71.3±19.6), o a las 12 semanas (76.1±18.7) comparado a nivel basal, después de controlar por otras variables.
La presente investigación tiene algunas limitaciones. Pocos ECR en la literatura han replicado los resultados, o han estudiado la CVRS en grupos específicos, como por ejemplo con coinfección por VIH, con uso de sustancias u otros trastornos psiquiátricos. Por otro lado, el tamaño de la muestra, la inclusión de pacientes con enfermedad hepática avanzada y sin coinfección con VIH, y la no inclusión de un grupo control sin HCC, limitan la generalización de nuestros resultados.
En conclusión, los resultados de la investigación apoyan que la calidad de vida puede mejorar después de un régimen antiviral exitoso. Es importante poder detectar pacientes con factores de riesgo, especialmente ellos con síntomas de depresión, antes de empezar cualquier tratamiento antiviral. En general, un enfoque multidisciplinar continúa siendo recomendable para mejorar la CVRS de los pacientes infectados por VHC.Hepatitis C virus (HCV) affects physical and mental health in 71 million persons worldwide. Classic antiviral treatment with interferon and ribavirin (PR) causes considerable impairment on chronic hepatitis C (CHC) patients’ life quality. Recently, direct-acting antivirals (DAAs) have been introduced, which have been associated with high cure rates (over 95%), reduced side effects, and are suggested to have a minimal impact on health-related quality of life (HRQL). However, the amount of evidence is still limited, as trials on these new regimens are ongoing. In this doctoral thesis, two studies were conducted in order to assess HRQL in CHC patients treated with DAAs: (I) a systematic review and meta-analysis of RCT studies that have assessed HRQL and possible risk factors that may predict life quality impairment, in CHC patients treated with any type or combination of DAAs; (II) a longitudinal naturalistic cohort study assessing HRQL and incidence of depression during antiviral treatment, taking in account possible risk factors that may predict life quality impairment and depression. Findings from the systematic review suggest that the new antiviral regimens have a minimal impact on HRQL, and may even improve in terms of mental wellbeing at 12 weeks of post-treatment follow-up. With regard to DAAs alone, a slight improvement in patients’ mental life quality was observed (MD=2.88; 95%CI=2.24, 3.53). However, ribavirin co-administration with DAAs showed significant impairment on mental HRQL (MD=-1.7; 95%CI=-2.5, -0.91). Any combination of DAAs with PR seemed to impair significantly both mental and physical health quality (MD= -0.13; 95%CI=-0.15, -0.11). At baseline, HRQL was more impaired in CHC patients who are unemployed, have cirrhosis, anaemia, or history of depression, anxiety, fatigue, or insomnia, than those who do not. Furthermore, female gender, older age, and history of depression seemed to predict HRQL impairment during DAAs plus ribavirin treatment. Also, adverse events and treatment non-response at post-treatment were identified risk factors for DAAs plus ribavirin or PR. In the second study, the cumulative incidence of major depression was 13.7% (95%CI: 5.7 to 26.3), and of any depressive disorder, 51% (95%CI: 36.6 to 65.2) during DAA treatment. Multivariate logistic regression analysis showed that the PHQ-9 score at baseline was a predictive factor for incidence of major depression (p=0.002), with a tendency for family history of depression (p=0.079). Also, decompensated cirrhotic patients reported a impaired pain and discomfort (p=0.045) compared to those without (decompensated) cirrhosis during DAA treatment. We could not exclude the presence of significant mean (SD) changes in EQ-VAS scores during DAA treatment (67.2±20.3), nor at EOT (71.3±19.6), or 12 weeks after treatment cessation (76.1±18.7) related to baseline after controlling by age, gender, comorbidity, history of depression, or ribavirin co-administration. This research has some limitations. Few RCTs in the literature have replicated their findings, and of those studies, only few of them have studied HRQL in specific groups such as co-infection, substance use disorder, and other psychiatric comorbidities. Other limitations of the study include the relatively small sample size, and inclusion of patients with more advanced liver disease and without HIV co-infection, which are factors that limit the generalization of our results. In summary, the results from this dissertation support that HRQL may improve after successful treatment. It is important to detect those patients with risk factors, especially for those with baseline symptoms of depression, before starting antiviral treatment. Altogether, findings suggest the use of a holistic, multidisciplinary approach to manage both physical and mental health.
42
Swaminathan, Kavya. "Novel anthocyanin inhibitors to influenza neuraminidase and monitioring antiviral resistance by mass spectrometry." Thesis, The University of Sydney, 2013. http://hdl.handle.net/2123/10220.
Full textAbstract:
A novel matrix assisted laser desorption ionization (MALDI) mass spectrometry based approach to study the binding of inhibitors to the influenza virus neuraminidase is described. The approach was shown to successfully be able to localize the binding of the known inhibitors of the viral neuraminidase – zanamivir and 2-deoxy-2,3-didehydro-N-acetylneuraminic acid, confirmed from the analysis of available X-ray crystal structures. The approach was extended to study the binding of an elderberry anthocyanin – cyanidin-3-sambubiocide to the neuraminidase, in parallel with computational approaches. Results revealed for the first time the molecular basis for the anthocyanidin’s neuraminidase inhibitory and by showing its binding within the neuraminidase 430-cavity, remote from residues known to regulate neuraminidase resistance. The results obtained herein provide a framework for the development of a new class of antivirals against influenza without this susceptibility. As an integral part of the anti-influenza drug development strategy a new phylogenetic approach for the surveillance of drug resistance and newly emerging strains is presented. It utilizes mass spectral data produced from proteolytic digestion of proteins, rather than gene/translated gene sequences to chart the evolutionary history of organisms. The concept and validity of the approach is demonstrated using theoretical and experimental mass data of the influenza hemagglutinin and neuraminidase. The ability of these trees to accurately cluster viral proteins from drug resistant strains is also shown and its relevance for surveillance of novel strains and drug-resistant mutants is also established by demonstrating their ability to accurately place experimentally derived mass data on mass trees. Given that the mass data can be generated more rapidly than gene sequences, mass trees offer new opportunities and advantages for phylogenetic analysis.
43
Giammarino, Federica. "PHENOTYPIC CHARACTERIZATION OF NOVEL ANTIVIRALS FOR THE TREATMENT OF MULTIDRUG RESISTANT HIV-1 AND EMERGING VIRUSES." Doctoral thesis, Università di Siena, 2023. https://hdl.handle.net/11365/1224634.
Full textAbstract:
Abstract
Phenotypic characterization of novel antivirals for the treatment of multidrug resistant HIV-1 and emerging viruses
Doctoral Research School of Medical Biotechnologies – Cycle XXXV
Supervisor: Maurizio Zazzi; Candidate: Federica Giammarino
Background
The need for new antiviral drugs has increased overtime due to the worldwide circulation of different viruses together with the increased frequency and diversity of new outbreaks. The ideal option for a prompt response against both emerging and re-emerging viruses is represented by the use and the development of direct acting antiviral agents. During my PhD I was involved in several projects focused on the evaluation of the antiviral activity of licensed and investigational antiviral drugs against Human Immunodeficiency (HIV-1), West Nile (WNV), Dengue (DENV) and SARS-CoV-2 viruses.
Results and discussion
Doravirine
The antiviral activity of the NNRTI doravirine was evaluated against viruses harbouring different patterns of NNRTI resistance mutations in two studies. Globally, our data confirmed that the antiviral activity of doravirine may be compromised by the presence of multiple NNRTI resistance mutations, even in the absence of specific doravirine mutations.
A third study was focused on the role of the natural polymorphism of the reverse transcriptase V106I. Our results indicate that it minimally affects the susceptibility to doravirine in clinical isolates and that it does not impact the genetic barrier to resistance as compared to reference wild-type virus, while viruses including the NNRTI resistant mutation V106A or V106M rapidly showed viral breakthrough under doravirine pressure due to the reduced susceptibility.
Islatravir
Our study confirmed the decrease of susceptibility of the investigational NRTTI islatravir due to the presence of M184V mutation. The clinical impact of NRTI mutations in the activity of islatravir has still to be defined and the threshold of fold-change values associated to reduced activity in vivo remains to be established.
Ibalizumab
The combinatorial activity of ibalizumab together with other antivirals, both approved and investigational, was evaluated through a newly developed cell-based assay consisting in the infection of the MOLT4-R5 cell line with the wild-type strains NL4-3 and AD8, and by the analysis of the results using the innovative software SynergyFinderPlus. Our data suggest that ibalizumab positively interacts with other antivirals with possible synergistic effects in select cases. Further studies are needed to determine the impact of Env variability and viral tropism in combination with other entry inhibitors.
Development of a Cell-Based Immunodetection Assay for Simultaneous Screening of Antiviral Compounds Inhibiting Zika and Dengue Virus Replication
An easy-to-perform and fast flavivirus immunodetection assay (IA) was developed to determine antiviral activity of promising compounds against ZIKV and DENV. The system, validated with references compounds against both viruses, was able to distinguish between the inhibitory effect of molecules targeting the early and the post-budding phase of viral replication cycle.
Evaluation of sofosbuvir activity and resistance profile against West Nile virus in vitro
Since the activity of sofosbuvir has been documented against different flaviviruses, we investigated whether it may exert an activity also against WNV. In both cell-based and enzymatic assays sofosbuvir was able to inhibit WNV replication in the low micromolar range. Moreover, in vitro selection and molecular docking experiments indicated that HCV and WNV share a similar sofosbuvir resistance pattern.
ORIGINALE CHEMIAE in Antiviral Strategy - Origin and Modernization of Multi-Component Chemistry as a Source of Innovative Broad Spectrum Antiviral Strategy
The “ORIGINALE CHEMIAE in Antiviral Strategy” project aims to identify promising broad-spectrum antivirals by taking advantage of the Multi-Component Chemistry strategy. Following the synthetization of molecules, their antiviral activity was determined in in vitro standardized virus-cell systems against DENV, WNV, HIV-1 and SARS-CoV-2. We identified eight molecules able to inhibit at least one of the viruses tested. However, their low selectivity indexes indicate the need to further improve the design of these molecules to increase the antiviral activity and/or reduce the cell toxicity in order to identify candidates for preclinical testing in animal models.
Monoclonal antibodies and antivirals vs. SARS-CoV-2
After the development of a quantitative live-virus microneutralization assay, we evaluated the efficacy of licensed monoclonal Antibodies (mAbs) and the antiviral drugs remdesivir, nirmaltrevir and molnupiravir against different circulating SARS-CoV-2 variants. Our results showed that these drugs, contrary to the mAbs, retained activity against all tested variants.
Conclusions
A continuous challenge for public health is represented by the control of viral infections. Both vaccines and antiviral drugs may synergistically help to reduce the spread and the fatality of acute viral diseases and chronic infections. All the studies described in this thesis emphasize the role of the laboratory of virology within all the steps of the in vitro investigation of antiviral drugs, from the identification of molecules endowed with antiviral activity to the definition of the mechanism of action.
44
Gaspareto, Karine Vieira. "Sequenciamento de nova geração para rastreamento de mutações de resistência aos novos medicamentos utilizados no tratamento da hepatite C." Universidade de São Paulo, 2017. http://www.teses.usp.br/teses/disponiveis/87/87131/tde-25092018-163244/.
Full textAbstract:
O presente estudo realizou o sequenciamento de nova geração do vírus da hepatite C genótipo 1, incluindo os subtipos 1a (n=51) e 1b (n=49), e identificou variantes associadas com resistência (RAV) aos antivirais de ação direta em pacientes sem tratamento prévio. No subtipo 1a, foram encontradas RAV para as regiões NS3-4A, NS5A e NS5B em 10%, 22% e 8% dos pacientes, respectivamente. RAV detectadas foram: T54S (2%), V55A (2%), Q80K (4%) e R155K (2%) na protease NS3-4A; Q30H (4%), H58P (10%) e Q30H/R+Y93C/H/N (8%) na região NS5A; e A421V (8%) na polimerase NS5B. As frequências das RAV para o subtipo 1b foram 12%, 53% e 31% para as regiões NS3-4A, NS5A e NS5B, respectivamente. Foram encontradas as RAV F43I (2%), T54S (4%), Q80H (2%), D168E (2%) e M175L (2%) na região NS3-4A; L28M (2%), R30Q (2%), L31M (2%), Q54H (27%), A92T (2%), Y93H (4%), Q54H+A92T (6%), Q54H+Y93H (6%) e A92T+Y93H (2%) na região NS5A e, L159F (2%), C316N (4%), A421V (7%), L159F+C316N (9%) e S556G (9%) na polimerase. Utilizando esta metodologia, um recombinante inter-subtipo 1a/1b foi identificado.This study performed the next-generation sequencing of the hepatitis C virus genotype 1, including subtypes 1a (n = 51) and 1b (n = 49), and identified resistance-associated variants (RAVs) to direct-acting antivirals in previously untreated patients. In subtype 1a, RAVs were found for NS3-4A, NS5A, and NS5B regions in 10%, 22% and 8% of patients, respectively. RAVs detected were: T54S (2%), V55A (2%), Q80K (4%) and R155K (2%) in NS3-4A protease; Q30H (4%), H58P (10%) and Q30H/R+Y93C/H/N (8%) in NS5A region; and A421V (8%) in NS5B polymerase. Frequencies of RAV for subtype 1b were 12%, 53% and 31% for NS3-4A, NS5A and NS5B regions, respectively. RAVs F43I (2%), T54S (4%), Q80H (2%), D168E (2%) and M175L (2%) were found in NS3-4a region; L28M (2%), R30Q (2%), L31M (2%), Q54H (27%), A92T (2%), Y93H (4%), Q54H+A92T (6%), Q54H+Y93H (6%) and A92T+Y93H (2%) in NS5A region and, L159F (2%), C316N (4%), A421V (7%), L159F+C316N (9%) and S556G (9%) in polymerase. By using this methodology, a recombinant inter-subtype 1a/1b was identified.
45
Freitas, Ferdinando. "Functional characterization of unassigned african swine fever virus proteins putatively involved in transcription and replication towards an efficient vaccine design." Doctoral thesis, Universidade de Lisboa, Faculdade de Medicina Veterinária, 2019. http://hdl.handle.net/10400.5/18129.
Full textAbstract:
Tese de Doutoramento em Ciências Veterinárias na especialidade de Ciências Biológicas e BiomédicasAfrican swine fever (ASF) is an infectious disease of domestic pigs and wild boars with mortality rates reaching up to 100% and is endemic in most of the Sub-Saharan countries. In 2007 it was introduced in Georgia and spread to neighbouring countries, reaching the Russian Federation, several European countries and, more recently, China and Vietnam (February 2019). Currently, there is neither a vaccine nor a treatment against ASF and the control of the disease depends strictly on sanitary measures, including stamping out and trade bans of animals and pork products leading to devastating socio-economic losses to affected countries. The etiologic agent of the disease is African swine fever virus (ASFV), a large (approx. 190 nm) double-stranded DNA (170 to 193 kbp) enveloped virus. ASFV genome encloses more than 150 open reading frames (ORFs) and to this date most of them lack any known or predictable function. ASFV is quite independent from cellular machinery encoding enzymes required for replication, transcription and virion assembly, including the putative I215L E2 Ubiquitin-conjugating enzyme, QP509L, Q706L RNA Helicases and the P1192R type II topoisomerase. The E2 ubiquitin-conjugating enzymes are part of the essential cellular post-transcriptional regulation ubiquitin-proteasome pathway. In this study, the pI215L binding activity was characterized as being mono and poly-ubiquitinated in the Cys85 at different temperatures and pH values. Moreover, I215L gene is transcribed from 2 hours post infection (hpi), and immunoblot analysis confirmed that pI215L is expressed from 4 hpi being detected all over the cell specially in the viral factories from 8 hpi. Downregulation assays by siRNA suggested that pI215L plays a critical role in the transcription of late viral genes and in viral DNA replication. RNA helicases are described as essential for infections, modulating RNA-RNA and RNA-protein interactions, gene expression, viral egress and host antiviral responses. In the present work, we found that QP509L, Q706L are conserved between ASFV virulent and non-virulent isolates. Furthermore, ASFV-QP509L and Q706L are actively transcribed from 2 hours post infection, and both proteins are localized in the viral factories at 12 hours post infection. However, QP509L was also detected in the cell nucleus. Transcript downregulation uncovered the essential role of these proteins during viral cycle progression, in particular for the late transcription. Type II topoisomerases are involved in resolving DNA tangles and supercoils by cutting the duplex and allowing the DNA replication to proceed. In this study, we report that P1192R is actively transcribed throughout infection, being detected from 2 hpi and reaching a maximum concentration around 16 hpi. P1192R knockdown experiments revealed its critical role for viral infection, given by a reduction in viral transcripts, cytopathic effect, the number of viral factories per cell, and virus yields. We also demonstrated that enrofloxacin exposure during the late phase of infection induces viral genomes fragmentation, whereas, when added at early phase of infection completely abolishes replication. The data obtained from I215L, QP509L. Q706L and P1192R characterization studies opens new venues to the rational design of a mutant virus lacking these genes, and also points new pathways to be targeted by antiviral drugs.
RESUMO - Caracterização funcional de proteínas do vírus da peste suína Africana putativamente envolvidas na transcrição e replicação com o intuito de desenvolvimento de uma vacina. - A peste suína africana é uma doença viral infeciosa que afeta os suínos domésticos e os selvagens, com taxas de mortalidade perto dos 100%, originando perdas económicas elevadas nos países afetados. A doença é endémica na maioria dos países subsaarianos, e desde 2007, assistiu-se uma expansão nos países Europeus, incluindo membros da União Europeia, e mais recentemente, na China e Vietname. Atualmente não existe vacina ou tratamento para esta infeção e o controlo da doença baseia-se no diagnóstico rápido, na eliminação compulsiva dos suínos e no bloqueio ao comércio de suínos e produtos derivados. O agente etiológico é o vírus da peste suína africana (VPSA), um vírus composto por uma molécula de ADN de cadeia dupla (170 to 193 kbp) contendo mais de 150 grelhas de leitura. Algumas destas estão devidamente caracterizadas codificando para proteínas estruturais ou regulatórias, contudo, a grande maioria foi identificada por homologia de sequência com outros vírus não se conhecendo, até à data, qual a sua função durante a infeção. Apesar dos inúmeros esforços ao longo dos anos, a complexidade viral, a falta de conhecimento sobre muitos dos aspetos da biologia do vírus e das suas interações com o hospedeiro invalidaram a obtenção de uma vacina segura e eficaz. Por um lado, as abordagens clássicas embora promissoras não garantem proteção contra estirpes heterólogas, enquanto a produção de vacinas de ADN ou proteína, mesmo com adjuvantes, não induzem imunidade contra uma segunda infeção. No entanto, a identificação de suínos previamente infetados e que resistem a novas infeções reforça a ideia da possibilidade de se obter uma imunidade protetora. Dadas as circunstâncias atuais de expansão da doença, estudos recentes apontam a necessidade de se aprofundar o conhecimento sobre os aspetos da biologia do VPSA com vista a identificação de novas estratégias para o desenvolvimento racional de vacinas ou de identificação de novos alvos para o uso de fármacos com vista a controlar a infeção. Neste contexto, os estudos apresentados neste trabalho caracterizam a I215L, QP509L, Q706L e P1192R, identificadas inicialmente, por homologia de sequência com outras proteínas tipicamente envolvidas na replicação e transcrição de outros vírus. A I215L foi identificada por partilhar identidade com as enzimas E2 de conjugação da ubiquitina. Estas enzimas pertencem a uma cadeia de sinalização do sistema de regulação pós-transcricional ubiquitina-proteossoma. Os estudos realizados revelaram que a pI215L tem a capacidade de receber uma ou duas ubiquitinas (mono e di-ubiquitinada) no resíduo Cisteína-85, a diferentes temperaturas e valores de pH, evidenciando a sua plasticidade em participar em diferentes fases da infeção quer no hospedeiro quer no vetor. Além disto, o gene é transcrito precocemente (2 horas após infecção, hpi) e a proteína expressa desde as 4h, sugerindo que esta deverá ser necessária desde o início da infecção. Paralelamente, os nossos estudos por imunofluorescência revelaram uma distribuição da pI215L por toda a célula, e em especial, nas fábricas virais, sugerindo um papel ativo na regulação de vários processos, incluindo replicação de ADN e da transcrição. Os ensaios de ARN de interferência (siRNA) contra o I215L demonstraram um papel essencial desta proteína durante a infeção, originando uma redução dos transcritos tardios, do número de genomas (-63 a -68%) e na libertação de partículas infeciosas (até -94%). [...]
N/A
46
Duwal, Sulav [Verfasser]. "A multiscale systems pharmacology framework to assess the prophylactic utility of antivirals against HIV-1 / Sulav Duwal." Berlin : Freie Universität Berlin, 2019. http://d-nb.info/1180387937/34.
Full text
47
Sarkar, Aurijit. "DEVELOPMENT AND APPLICATIONS OF THE HINT FORCEFIELD IN PREDICTION OF ANTIBIOTIC EFFLUX AND VIRTUAL SCREENING FOR ANTIVIRALS." VCU Scholars Compass, 2010. http://scholarscompass.vcu.edu/etd/2266.
Full textAbstract:
This work was aimed at developing novel tools that utilize HINT, an empirical forcefield capable of quantitating both hydrophobic and hydrophilic (hydropathic) interactions, for implementation in theoretical biology and drug discovery/design. The role of hydrophobicity in determination of macromolecular structure and formation of complexes in biological molecules is undeniable and has been the subject of research across several decades. Hydrophobicity is introduced, with a review of its history and contemporary theories. This is followed by a description of various methods that quantify this all-pervading phenomenon and their use in protein folding and contemporary drug design projects – including a detailed overview of the HINT forcefield. The specific aim of this dissertation is to introduce our attempts at developing new methods for use in the study of antibacterial drug resistance and antiviral drug discovery. Multidrug efflux is commonly regarded as a fast growing problem in the field of medicine. Several species of microbes are known to have developed resistance against almost all classes of antibiotics by various modes-of-action, which include multidrug transporters (a.k.a. efflux pumps). These proteins are present in both gram-positive and gram-negative bacteria and extrude molecules of various classes. They protect the efflux pump-expressing bacterium from harmful effects of exogenous agents by simply evacuating the latter. Perhaps the best characterized mechanism amongst these is that of the AcrA-AcrB-TolC efflux pump. Data is available in literature and perhaps also in proprietary databases available with pharmaceutical companies, characterizing this pump in terms of the minimum inhibitory concentration ratios (MIC ratios) for various antibiotics. We procured a curated dataset of 32 β-lactam and 12 antibiotics of other classes from this literature. Initial attempts at studying the MIC ratios of β-lactam antibiotics as a function of their three dimensional topology via 3D-quantitative structure activity relationship (3D-QSAR) technology yielded seemingly good models. However, this methodology is essentially designed to address single receptor-ligand interactions. Molecules being transported by the efflux pump must undoubtedly be involved in multiple interactions with the same. Notably, such methods require a pharmacophoric overlap of ligands prior to the generation of models, thereby limiting their applicability to a set of structurally-related compounds. Thus, we designed a novel method that takes various interactions between antibiotic agents and the AcrA-AcrB-TolC pump into account in conjunction with certain properties of the drugs. This method yielded mathematical models that are capable of predicting high/low efflux with significant efficiency (>93% correct). The development of this method, along with the results from its validation, is presented herein. A parallel aim being pursued by us is to discover inhibitors for hemagglutinin-neuraminidase (HN) of human parainfluenza virus type 3 (HPIV3) by in silico screening. The basis for targeting HN is explored, along with commentary on the methodology adopted during this effort. This project yielded a moderate success rate of 34%, perhaps due to problems in the computational methodology utilized. We highlight one particular problem – that of emulating target flexibility – and explore new avenues for overcoming this obstacle in the long run. As a starting point towards enhancing the tools available to us for virtual screening in general (and for discovering antiviral compounds in specific), we explored the compatibility between sidechain rotamer libraries and the HINT scoring function. A new algorithm was designed to optimize amino acid residue sidechains, if provided with the backbone coordinates, by generating sidechain positions using the Dunbrack and Cohen backbone-dependent rotamer library and scoring them with the HINT scoring function. This rotamer library was previously used by its developers previously to design a very successful sidechain optimization algorithm called SCWRL. Output structures from our algorithm were compared with those from SCWRL and showed extraordinary similarities as well as significant differences, which are discussed herein. This successful implementation of HINT in our sidechain optimization algorithm establishes the compatibility between this forcefield and sidechain rotamer libraries. Future aims in this project include enhancement of our current algorithm and the design of a new algorithm to explore partial induced-fit in targets aimed at improving current docking methodology. This work shows significant progress towards the implementation of our hydropathic force field in theoretical modeling of biological systems in order to enhance our ability to understand atomistic details of inter- and intramolecular interactions which must form the basis for a wide variety of biological phenomena. Such efforts are key to not only to understanding the said phenomena, but also towards a solid basis for efficient drug design in the future.
48
Trivisani. "APPLICATION OF COMPUTATIONAL METHODS FOR THE IDENTIFICATION OF NEW DDX3X INHIBITORS." Doctoral thesis, Università di Siena, 2021. http://hdl.handle.net/11365/1127108.
Full textAbstract:
The search for new antiviral drugs for the treatment of clinical and emerging viruses is a delicate task. Drugs available today are developed to target a specific virus or viral strain, and only few prophylactics show a broad-spectrum activity that can be used to treat drug-resistant infections or in case of emergencies. Among the strategies that can be pursued to search for new antiviral drugs, there is the inhibition of a host protein involved in the viral replication cycle. The inhibition of DDX3X, a human ATP-dependent RNA helicase, allowed to discover the first broad-spectrum antiviral compound able to inhibit the replication of HIV resistant strains, HCV and of emerging viruses like West Nile Virus, Japanese Encephalitis Virus, Dengue Virus.
In this project, several computational strategies have been applied to improve the biodistribution and pharmacokinetic properties of this compound and a fluorescent inhibitor was designed to understand the mode of action of DDX3X inhibitors in DENV infected cells.
The selective inhibition of DDX3X can be pursued targeting a small pocket, peculiar to the human protein, called unique motif (UM). The study of the interactions established by the first active compound within UM, allowed to identify the amino acids responsible of its activity. Considering these findings, a small library of derivatives able to establish the fundamental interactions with the UM was designed. Moreover, a pharmacophore-based virtual screening procedure allowed to discover new compounds that will be biologically evaluated as new UM inhibitors.
PROTAC is instead a potent strategy to target protein degradation. A PROTAC molecule, that is constituted by two active moieties hold together by a linker, allow the selective ubiquitination and degradation of the protein of interest by the proteasome. In this contest, the application of computational procedures on known SOCS2 binders, allowed to establish a SAR that will be used to design novel derivatives that can be used both as inhibitors of the protein and as binders of the E3 ligase. A pharmacophore-based virtual screening performed on the Elongin C allowed to discover new compounds that will be biologically evaluated to establish if they can be used as E3 ligase binders.
49
Bouet, Gabrielle. "Identification d'une molécule antivirale active in-vivo chez le cheval contre le virus de l'artérite virale équine." Electronic Thesis or Diss., Normandie, 2023. http://www.theses.fr/2023NORMC422.
Full textAbstract:
Depuis plus de 60 ans, le virus de l’artérite virale équine (EAV) induit des problèmes respiratoires et reproducteurs des équidés. L’EAV appartient à la sous-famille des Equarterivirinae, famille Arteriviridae de l’ordre des Nidovirales comme le SARS-CoV2. L’avortement de juments gravides et l’établissement du statut d’excréteur viral chez les étalons font de l’EAV un problème majeur pour les éleveurs et le développement de la filière équine. Malgré la commercialisation de 2 vaccins, la couverture vaccinale est aujourd’hui insuffisante pour prévenir les épizooties mondiales. Récemment au laboratoire, nous avons identifié la première molécule commerciale antivirale inhibant la réplication de l’EAV in vitro, la ribavirine. Malheureusement, son utilisation est uniquement approuvée chez l’homme et les petits animaux. Cette découverte met en évidence qu’un traitement contre l’EAV est possible.Au cours de cette thèse, nous avons i) validé un modèle in vitro de criblage haut-débit (HTS) permettant d’évaluer la cytotoxicité et la cytoprotection cellulaire induites par 1250 molécules issues de 2 chimiothèques testées à différentes concentrations. ii) Sur les 75 molécules identifiées comme potentiellement antivirales, nous avons confirmé que 49 molécules sont antivirales à des concentrations moins cytotoxiques et plus cytoprotectrices que la ribavirine. iii) Nous avons cherché à caractériser le mécanisme d’action de la molécule présentant la meilleure activité et nous avons iv) de déterminer les caractéristiques pharmacologiques et pharmacodynamiques in vitro des 2 molécules sélectionnées. Ces travaux in vitro renforcent la faisabilité d’un traitement antiviral contre l’EAV et ouvre la possibilité de tester ces molécules in vivoFor over 60 years, the equine viral arteritis (EAV) virus has been causing respiratory and reproductive problems in equids. EAV belongs to the Equarterivirinae subfamily, Arteriviridae family of the order Nidovirales, like SARS-CoV2. The abortion of pregnant mares and the establishment of viral excretor status in stallions make EAV a major problem for breeders and the development of the equine industry. Despite the marketing of 2 vaccines, vaccination coverage is currently insufficient to prevent worldwide epizootics. Recently in the laboratory, we identified the first commercial antiviral molecule to inhibit in vitro EAV replication, ribavirin. Unfortunately, it is only approved for use in humans and small animals. This discovery shows that a treatment for AVE is possible. In the course of this thesis, we i) validated an in vitro high-throughput screening (HTS) model to assess the cytotoxicity and cellular cytoprotection induced by 1250 molecules from 2 chemical libraries tested at different concentrations. ii) Of the 75 molecules identified as potentially antiviral, we confirmed that 49 molecules are antiviral at concentrations less cytotoxic and more cytoprotective than ribavirin. iii) We sought to characterize the mechanism of action of the molecule with the best activity, and to determine the in vitro pharmacological and pharmacodynamic characteristics of the 2 selected molecules. This in vitro work reinforces the feasibility of an antiviral treatment against EAV and opens up the possibility of testing these molecules in vivo
50
Ruoff, Kerstin [Verfasser], and Martin [Akademischer Betreuer] Müller. "Analysis of synthetic compounds and natural extracts as potential antivirals against human Noroviruses / Kerstin Ruoff ; Betreuer: Martin Müller." Heidelberg : Universitätsbibliothek Heidelberg, 2020. http://d-nb.info/1226428541/34.
Full text