RESUMEN
BACKGROUND & AIMS: SCY-635 is a non-immunosuppressive analog of cyclosporin A that inhibits cyclophilins A and B and hepatitis C virus (HCV) replication in vitro. In a phase 1b multi-dose escalation study, we evaluated the safety, plasma pharmacokinetics, and antiviral activity of 15 days of monotherapy with SCY-635 in adults with chronic genotype 1 HCV infection. METHODS: Twenty adults with chronic HCV genotype 1 were randomized to SCY-635 oral doses of 100, 200, or 300 mg three times daily for 15 days. RESULTS: No dose-limiting clinical or laboratory toxicities were identified. On day 15, the mean decline in plasma viremia was 2.24±1.74 log(10) IU/ml with SCY-635 900 mg/d. Individual antiviral responses correlated with host IL28B genotype. Post hoc analyses indicated treatment with SCY-635 increased plasma protein concentrations of interferon α (IFNα), IFNs λ(1) and λ(3), and 2'5' oligoadenylate synthetase 1 (2'5'OAS-1), with the greatest increases in IL28B CC and CT subjects. Changes in plasma concentrations for all markers were coincident with changes in the plasma concentration of SCY-635. Peaks of IFNs α, λ(1), and λ(3) and 2'5'OAS-1 were observed within 2 h after drug administration. In replicon cells, SCY-635 enhanced secretion of type I and type III IFNs and increased the expression of IFN-stimulated genes (ISG). CONCLUSIONS: These studies establish clinical proof of concept for SCY-635 as a novel antiviral agent and suggest that restoration of the host innate immune response to chronic hepatitis C infection may represent a major mechanism through which cyclophilin inhibitors exert clinical antiviral activity.
Asunto(s)
Antivirales/administración & dosificación , Ciclofilina A/antagonistas & inhibidores , Ciclofilinas/antagonistas & inhibidores , Ciclosporinas/administración & dosificación , Hepacivirus/efectos de los fármacos , Hepatitis C Crónica/tratamiento farmacológico , Adulto , Anciano , Antivirales/efectos adversos , Antivirales/farmacocinética , Carcinoma Hepatocelular , Línea Celular Tumoral , Ciclosporinas/efectos adversos , Ciclosporinas/farmacocinética , Relación Dosis-Respuesta a Droga , Femenino , Genotipo , Hepacivirus/genética , Hepacivirus/crecimiento & desarrollo , Hepatitis C Crónica/inmunología , Humanos , Interferón-alfa/sangre , Interferón beta/sangre , Interferón gamma/sangre , Interferones , Interleucinas/genética , Neoplasias Hepáticas , Masculino , Persona de Mediana Edad , Resultado del TratamientoRESUMEN
An important challenge for primary or secondary analysis of cytometry data is how to facilitate productive collaboration between domain and quantitative experts. Domain experts in cytometry laboratories and core facilities increasingly recognize the need for automated workflows in the face of increasing data complexity, but by and large, still conduct all analysis using traditional applications, predominantly FlowJo. To a large extent, this cuts domain experts off from the rapidly growing library of Single Cell Data Science algorithms available, curtailing the potential contributions of these experts to the validation and interpretation of results. To address this challenge, we developed FlowKit, a Gating-ML 2.0-compliant Python package that can read and write FCS files and FlowJo workspaces. We present examples of the use of FlowKit for constructing reporting and analysis workflows, including round-tripping results to and from FlowJo for joint analysis by both domain and quantitative experts.
Asunto(s)
Citometría de Flujo/métodos , Programas Informáticos , Flujo de Trabajo , Algoritmos , Biología Computacional , Humanos , Aprendizaje Automático , Análisis de la Célula IndividualRESUMEN
An efficient synthesis of [D-lysine](8)cyclosporin A has been developed. Several analogs of [D-lysine](8)cyclosporin A have been synthesized and show promising anti-HCV activity, particularly compounds 39 and 43, which each exhibit an anti-HCV EC(50)<200 nM, and are each ≥50-fold less immunosuppressive than cyclosporin A.
Asunto(s)
Antivirales/síntesis química , Antivirales/farmacología , Ciclosporina/síntesis química , Ciclosporina/farmacología , Hepacivirus/efectos de los fármacos , Cristalografía por Rayos X , Modelos MolecularesRESUMEN
Enfuvirtide (ENF), the first human immunodeficiency virus type 1 (HIV-1) fusion inhibitor approved for clinical use, acts by binding to gp41 heptad repeat 1 (HR1) and preventing its interaction with the viral HR2 region. Treatment-emergent resistance to ENF has been mapped to residues within HR1, and these mutations decrease its susceptibility to ENF and may reduce viral fitness and pathogenesis, although the mechanism for these effects is not clear. N43D, a common ENF resistance mutation, was found in in vitro assays to cause a 5-50-fold in antiviral activity. We introduced this mutation into peptide models and determined the impact of this mutation by circular dichroism and X-ray crystallography. We find that the mutation results in a decrease in the thermal stability of the six-helix bundle and causes a significant change in the HR1-HR2 interface, including a loss of HR2 helicity. These data form a mechanistic basis for the decrease in ENF sensitivity and six-helix bundle stability. The E137K polymorphism, generally present at baseline in patients who develop N43D, partially compensates for the loss of stability, and we show that these residues likely form an ion pair. These data form a framework for understanding the impact of resistance mutations on viral fitness and pathogenesis and provide a pathway for the development of novel fusion inhibitor peptides.
Asunto(s)
Farmacorresistencia Viral/genética , Proteína gp41 de Envoltorio del VIH/farmacología , VIH-1/efectos de los fármacos , Fragmentos de Péptidos/farmacología , Sitios de Unión , Dicroismo Circular , Cristalografía por Rayos X , Enfuvirtida , Proteína gp41 de Envoltorio del VIH/química , Proteína gp41 de Envoltorio del VIH/genética , Inhibidores de Fusión de VIH/farmacología , VIH-1/genética , Humanos , Enlace de Hidrógeno , Concentración 50 Inhibidora , Modelos Moleculares , Mutación , Polimorfismo Genético , Estructura Secundaria de Proteína , Estructura Terciaria de ProteínaRESUMEN
BACKGROUND: Enfuvirtide (ENF) is the first of a novel class of drugs that block HIV gp41-mediated viral fusion to host cells. Viruses with mutations at positions 36-38 in HIV-1 gp41 and/or reduced susceptibility to ENF have been selected both in vitro and in vivo. METHODS: An analysis of baseline and on-treatment ENF susceptibility in virus samples from Phase II clinical trial patients treated with ENF as functional monotherapy for 28 days (TRI-003) or in combination with oral antiretrovirals for >/= 48 weeks (T20-205, T20-206 and T20-208). Population sequencing identified amino acid (aa) substitutions at positions 36-45 of gp41 in plasma HIV-1. ENF susceptibility of virus isolates was tested in the cMAGI assay and viral DNA was sequenced for selected isolates. RESULTS: HIV-1 gp41 aa 36-45 were highly conserved in virus from ENF-naive patients, except for a 15% incidence of N42S which did not reduce sensitivity to ENF. Virus from patients experiencing viral load rebound exhibited reduced susceptibility to ENF and substitutions in gp41 aa 36-45. The most common substitutions observed on treatment were at positions 36, 38, 40, 42 and 43. On-treatment changes in the phenotypic susceptibility of virus isolates to ENF were generally associated with genotypic changes in aa 36-45. There was a relatively lower incidence of ENF resistance in patients with baseline sensitivity to more oral antiretrovirals in comparison to patients sensitive to fewer antiretrovirals. CONCLUSIONS: These data identify the importance of HIV-1 gp41 aa 36-45 in the emergence of resistance to ENF.