Recovery of West Nile Virus Envelope Protein Domain III Chimeras with Altered Antigenicity and Mouse Virulence.

UNLABELLED: Flaviviruses are positive-sense, single-stranded RNA viruses responsible for millions of human infections annually. The envelope (E) protein of flaviviruses comprises three structural domains, of which domain III (EIII) represents a discrete subunit. The EIII gene sequence typically encodes epitopes recognized by virus-specific, potently neutralizing antibodies, and EIII is believed to play a major role in receptor binding. In order to assess potential interactions between EIII and the remainder of the E protein and to assess the effects of EIII sequence substitutions on the antigenicity, growth, and virulence of a representative flavivirus, chimeric viruses were generated using the West Nile virus (WNV) infectious clone, into which EIIIs from nine flaviviruses with various levels of genetic diversity from WNV were substituted. Of the constructs tested, chimeras containing EIIIs from Koutango virus (KOUV), Japanese encephalitis virus (JEV), St. Louis encephalitis virus (SLEV), and Bagaza virus (BAGV) were successfully recovered. Characterization of the chimeras in vitro and in vivo revealed differences in growth and virulence between the viruses, within vivo pathogenesis often not being correlated within vitro growth. Taken together, the data demonstrate that substitutions of EIII can allow the generation of viable chimeric viruses with significantly altered antigenicity and virulence. IMPORTANCE: The envelope (E) glycoprotein is the major protein present on the surface of flavivirus virions and is responsible for mediating virus binding and entry into target cells. Several viable West Nile virus (WNV) variants with chimeric E proteins in which the putative receptor-binding domain (EIII) sequences of other mosquito-borne flaviviruses were substituted in place of the WNV EIII were recovered, although the substitution of several more divergent EIII sequences was not tolerated. The differences in virulence and tissue tropism observed with the chimeric viruses indicate a significant role for this sequence in determining the pathogenesis of the virus within the mammalian host. Our studies demonstrate that these chimeras are viable and suggest that such recombinant viruses may be useful for investigation of domain-specific antibody responses and the more extensive definition of the contributions of EIII to the tropism and pathogenesis of WNV or other flaviviruses.

Evaluation of molnupiravir (EIDD-2801) efficacy against SARS-CoV-2 in the rhesus macaque model.

Molnupiravir (EIDD-2801) is a prodrug of a ribonucleoside analogue that is currently being used under a US FDA emergency use authorization for the treatment of mild to moderate COVID-19. We evaluated molnupiravir for efficacy as an oral treatment in the rhesus macaque model of SARS-CoV-2 infection. Twenty non-human primates (NHPs) were challenged with SARS-CoV-2 and treated with 75 mg/kg (n = 8) or 250 mg/kg (n = 8) of molnupiravir twice daily by oral gavage for 7 days. The NHPs were observed for 14 days post-challenge and monitored for clinical signs of disease. After challenge, all groups showed a trend toward increased respiration rates. Treatment with molnupiravir significantly reduced viral RNA levels in bronchoalveolar lavage (BAL) samples at Days 7 and 10. Considering the mild to moderate nature of SARS-CoV-2 infection in the rhesus macaque model, this study highlights the importance of monitoring the viral load in the lung as an indicator of pharmaceutical efficacy for COVID-19 treatments. Additionally, this study provides evidence of the efficacy of molnupiravir which supplements the current ongoing clinical trials of this drug.

The Fc-mediated effector functions of a potent SARS-CoV-2 neutralizing antibody, SC31, isolated from an early convalescent COVID-19 patient, are essential for the optimal therapeutic efficacy of the antibody.

Although SARS-CoV-2-neutralizing antibodies are promising therapeutics against COVID-19, little is known about their mechanism(s) of action or effective dosing windows. We report the generation and development of SC31, a potent SARS-CoV-2 neutralizing antibody, isolated from a convalescent patient. Antibody-mediated neutralization occurs via an epitope within the receptor-binding domain of the SARS-CoV-2 Spike protein. SC31 exhibited potent anti-SARS-CoV-2 activities in multiple animal models. In SARS-CoV-2 infected K18-human ACE2 transgenic mice, treatment with SC31 greatly reduced viral loads and attenuated pro-inflammatory responses linked to the severity of COVID-19. Importantly, a comparison of the efficacies of SC31 and its Fc-null LALA variant revealed that the optimal therapeutic efficacy of SC31 requires Fc-mediated effector functions that promote IFNγ-driven anti-viral immune responses, in addition to its neutralization ability. A dose-dependent efficacy of SC31 was observed down to 5mg/kg when administered before viral-induced lung inflammatory responses. In addition, antibody-dependent enhancement was not observed even when infected mice were treated with SC31 at sub-therapeutic doses. In SARS-CoV-2-infected hamsters, SC31 treatment significantly prevented weight loss, reduced viral loads, and attenuated the histopathology of the lungs. In rhesus macaques, the therapeutic potential of SC31 was evidenced through the reduction of viral loads in both upper and lower respiratory tracts to undetectable levels. Together, the results of our preclinical studies demonstrated the therapeutic efficacy of SC31 in three different models and its potential as a COVID-19 therapeutic candidate.

Cross-neutralisation of viruses of the tick-borne encephalitis complex following tick-borne encephalitis vaccination and/or infection.

The tick-borne encephalitis complex contains a number of flaviviruses that share close genetic homology, and are responsible for significant human morbidity and mortality with widespread geographical range. Although many members of this complex have been recognised for decades, licenced human vaccines with broad availability are only available for tick-borne encephalitis virus. While tick-borne encephalitis virus vaccines have been demonstrated to induce significant protective immunity, as determined by virus-neutralisation titres, vaccine breakthrough (clinical infection following complete vaccination), has been described. The aim of this study was to confirm the cross-neutralisation of tick-borne flaviviruses using mouse immune ascitic fluids, and to determine the magnitude of cross-neutralising antibody titres in sera from donors following tick-borne encephalitis vaccination, infection, and vaccine breakthrough. The results demonstrate that there is significant cross-neutralisation of representative members of the tick-borne encephalitis complex following vaccination and/or infection, and that the magnitude of immune responses varies based upon the exposure type. Donor sera successfully neutralised most of the viruses tested, with 85% of vaccinees neutralising Kyasanur forest disease virus and 73% of vaccinees neutralising Alkhumra virus. By contrast, only 63% of vaccinees neutralised Powassan virus, with none of these neutralisation titres exceeding 1:60. Taken together, the data suggest that tick-borne encephalitis virus vaccination may protect against most of the members of the tick-borne encephalitis complex including Kyasanur forest disease virus and Alkhumra virus, but that the neutralisation of Powassan virus following tick-borne encephalitis vaccination is minimal.

Plasticity of a critical antigenic determinant in the West Nile virus NY99 envelope protein domain III.

West Nile virus (WNV) is a mosquito-borne flavivirus that causes febrile illness, encephalitis, and occasionally death in humans. The envelope protein is the main component of the WNV virion surface, and domain III of the envelope protein (EIII) is both a putative receptor binding domain and a target of highly specific, potently neutralizing antibodies. Envelope E-332 (E-332) is known to have naturally occurring variation and to be a key determinant of neutralization for anti-EIII antibodies. A panel of viruses containing all possible amino acid substitutions at E-332 was constructed. E-332 was found to be highly tolerant of mutation, and almost all of these changes had large impacts on antigenicity of EIII but only limited effects on growth or virulence phenotypes.

La observancia de la bioética en el cuidado paliativo de enfermos con esclerosis lateral amiotrófica / The observance of bioethics in the palliative care of patients with amyotrophic lateral sclerosis / A observância da Bioética no cuidado paliativo de doentes com esclerose lateral amiotrófica

Resumen La esclerosis lateral amiotrófica (ELA) es una enfermedad neurodegenerativa de etiología desconocida, curso progresivo y pronóstico desfavorable, para la cual no existe tratamiento curativo. Muchos son los profesionales de la salud involucrados en la atención de un paciente con ELA. El manejo apropiado de los problemas éticos es de gran importancia en todas las etapas de la ELA. El objetivo del siguiente trabajo es identificar las categorías éticas más vulneradas que atentan la calidad de la atención a estos pacientes. Se realizó una investigación cualitativa, cualitativa, observacional, descriptiva, longitudinal; se empleó la observación participativa y se tomó como base el espectro completo de cuestiones éticas en la atención de pacientes con ELA. Como resultado se encontró cumplimiento incompleto del respeto al derecho del paciente a una información completa, poca consideración de las consecuencias de la información insuficiente, así como irrespeto por las preferencias de la persona enferma en el tratamiento de situaciones de emergencia. Las categorías más vulneradas por los profesionales de salud fueron: el proceso de toma de decisiones, la toma de decisiones al final de la vida y la indicación médica.

Abstract Amyotrophic lateral sclerosis is a neurodegenerative disease of unknown etiology, progressive course and unfavorable prognosis, for which there is no curative treatment. Many are the health professionals involved in the care of a patient with ALS. The proper handling of ethical problems is of great importance in all stages of ELA. The objective of the following work is to identify the most vulnerable ethical categories that threaten the quality of care for these patients. A qualitative longitudinal descriptive observational research was carried out, participatory observation was used and the full spectrum of ethical issues in the care of patients with ALS was taken as a basis. As a result, incomplete compliance with the patient's right to complete information was found, little consideration of the consequences of insufficient information, and disrespect for the preferences of the sick person in the treatment of emergency situations. The categories most affected by health professionals were: The decision-making process, decision making at the end of life and medical indication.

Resumo A esclerose lateral amiotrófica (ELA) é uma doença neurodegenerativa de etiologia desconhecida, curso progressivo e prognóstico desfavorável, para a qual não há tratamento curativo. Muitos profissionais da saúde estão envolvidos na atenção de um paciente com ELA. O manejo apropriado dos problemas éticos é de grande importância em todas as etapas dessa doença. Este trabalho tem como objetivo identificar as categorias éticas mais vulneradas que atentam a qualidade da atenção a esses pacientes. Foi realizada uma pesquisa qualitativa, observacional, descritiva e longitudinal; foram utilizados a observação participativa e, como base, o espectro completo de questões éticas na atenção de pacientes com ELA. Como resultado, verificou-se que não se cumpriu totalmente o direito do paciente a uma informação completa, que houve pouca consideração das consequências da informação insuficiente, bem como desrespeito das preferências da pessoa doente no tratamento de situações de emergência. As categorias mais vulneradas pelos profissionais da saúde foram: o processo de tomada de decisões; a tomada de decisões no final da vida e a indicação médica.

Consentimiento informado en los cuidados paliativos de pacientes con esclerosis lateral amiotrófica / Informed consent in the palliative care for patients with amyotrophic lateral sclerosis

Introducción: El consentimiento informado es un proceso comunicativo primordial para preservar y promover la autonomía moral de las personas en el ámbito socio-sanitario. Es imprescindible el conocimiento pleno de dicho proceso por los profesionales de salud involucrados en la toma de decisiones, tanto asistenciales como investigativas, para el logro de una atención de alta calidad. Objetivo: fundamentar la importancia del consentimiento informado en el cuidado paliativo de las personas que padecen esclerosis lateral amiotrófica. Material y Métodos: Se realizó una revisión bibliográfica donde se hicieron búsquedas en sitios de información científica certificada como PubMed, Cochrane y LILACS, con los términos consentimiento informado, consentimiento informado and cuidados paliativos consentimiento informado and derechos humanos y Consentimiento informado and esclerosis lateral amiotrófica. Resultados: el ejercicio del consentimiento informado en el marco de la relación médico paciente constituye expresión de solidaridad, respeto y humanismo, derecho humano inviolable, responsabilidad profesional. El consentimiento informado en cuidados paliativos merece una reflexión más sensata y juiciosa que permita a los profesionales hacer uso adecuado de este proceso para garantizar una atención de salud de máxima calidad. Conclusiones: el consentimiento informado reviste gran importancia pues permite la toma conjunta de decisiones autónomas, competentes, razonables y moralmente válidas y constituye expresión de solidaridad, respeto y humanismo, derecho humano inviolable y responsabilidad profesional(AU)

Introduction: Informed consent is an essential communicative process aimed at preserving and promoting the moral autonomy of the people in the social-sanitary level. A full knowledge of this process is indispensable for healthcare professionals involved in making decisions in both the healthcare and research areas in order to achieve a good quality care. Objective: To establish the importance of informed consent in the palliative care for people suffering from amyotrophic lateral sclerosis. Material and Methods: A literature review was carried out through the search in databases such as PubMed, Cochrane and LILACS, using keywords such as informed consent, informed consent and palliative care, informed consent and human rights, and informed consent and amyotrophic lateral sclerosis. Results: The practice of the informed consent within the framework of the doctor-patient relationship is an expression of solidarity, respect and humanism, an inviolable human right, and a professional responsibility. Informed consent in the palliative treatments merits a more sensible thinking that allows professionals to make an adequate use of this process to guarantee a high quality healthcare. Conclusions: Informed consent is of great importance because it provides joint decision-making, which should be autonomous, competent, reasonable, and morally valid, representing an expression of solidarity, respect, humanism, inviolable human right, and professional responsibility(AU)