Mechanisms of influenza NA drug resistance

流感NA耐药机制

基本信息

批准号：
7257291
负责人：
JENNIFER L MCKIMM-BRESCHKIN
金额：
$ 26.22万
依托单位：
COMMONWEALTH SCIENTIFIC & INDUST RES ORG
依托单位国家：
美国
项目类别：
财政年份：
2006
资助国家：
美国
起止时间：
2006-07-15 至 2011-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7257291
关键词：
Active Sites Address Adult Affect Affinity Chromatography Amino Acids Antibodies Asia Avian Influenza Baculovirus Expression System Baculoviruses Binding Binding Sites Biological Biological Assay Birds Carbohydrates Cells Cercopithecine Herpesvirus 1 Characteristics Chemical Structure Child Cleaved cell Clinical Committee Members Complex Condition Country Crystallization Cultured Cells Data Development Disease Outbreaks Dose Drug Exposure Drug Kinetics Drug resistance Engineering Enzymes Epidemic Equilibrium Europe Fab Immunoglobulins Future Gel Chromatography Genes Genotype Growth Head Human Image Immunocompetent In Vitro Influenza Influenza A Virus, H1N1 Subtype Influenza A Virus, H5N1 Subtype Influenza B virus Insecta International Investigation Japan Kinetics Knowledge Length Light Lung Medical Surveillance Membrane Minor Molecular Monitor Monoclonal Antibodies Mus Mutate Mutation Neuraminidase Neuraminidase inhibitor Oseltamivir Patients Pharmaceutical Preparations Phenotype Play Population Predisposition Principal Investigator Production Pronase Proteins Purpose Rate Recombinant Proteins Recombinants Reliance Reporting Research Personnel Resistance Resistance development Resources Robot Roentgen Rays Role Sampling Screening procedure Sialic Acids Site Structure Surveillance Program System Technology Testing Thea Plant Thinking Tokyo Trypsin Vaccines Variant Virion Virus Virus Replication Work anti-influenza drug base c-myc Genes clinically relevant drug resistant virus drug sensitivity egg fitness in vivo influenzavirus inhibitor/antagonist monoclonal antibody production mouse model mutant pandemic disease programs protein function receptor binding sugar thermostability transmission process zanamivir zanamivir resistance

项目摘要

DESCRIPTION (provided by applicant): There are two specific anti-influenza drugs which are effective against all strains of influenza, zanamivir and oseltamivir. Since they bind to highly conserved residues in the active site of the virus neuraminidase (NA) they are effective against all strains of influenza. With the spread of the highly pathogenic H5N1 avian influenza in Asia and now across into Europe, the possibility of a pandemic strain emerging is more likely. Since it will take several months to produce a vaccine the NA inhibitors, especially oseltamivir, are being stockpiled as the first line of defense. While resistance to zanamivir has not yet been seen in immunocompetent patient, there are 3 common mutations in conserved residues in the NA active site which confer oseltamivir resistance. However, these mutations do not confer resistance in all NA subtypes. Resistance rates of up to 18% have now been seen after oseltamivir treatment, and there are already reports of H5N1 oseltamivir resistant viruses, hence there are increasing concerns about drug resistance. Through the Neuraminidase Inhibitor Susceptibility Network (NISN) global surveillance program we have also identified several other viruses which have mutations in non-conserved residues outside of the active site. There must therefore be secondary structural effects outside the active site which can impact on drug binding. Hence our central hypothesis is that "Additional secondary structural characteristics in the NA play a critical role in determining drug sensitivity and whether a mutation confers drug resistance". In addition to the NA mutations, we also have HA mutations in one of the isolates which confer resistance in vitro. Hence we also have a minor hypothesis that "HA mutations in clinical isolates can play a role in resistance to the NA inhibitors in vivo". The long term aim of this project is to use structural and biological studies to understand the mechanisms of influenza resistance which contribute to differences in N1/N2 drug sensitivity, N1/N2 subtype specific resistance, how mutations in non-conserved residues outside the active site affect inhibitor binding, as well as the role of HA mutations in resistance in vivo. We aim to determine the structures of the wild type and mutant NAs from either egg-grown or baculovirus expressed protein. We will assess the impacts of the NA and HA mutations on growth and drug sensitivity in vitro and in vivo as an indication of the "fitness" of the viruses , to understand the likelihood of them being transmissible and drug resistant in the clinical situation. Relevance Understanding the mechanisms of drug resistance, either in naturally occurring or drug selected variants, is critical for appropriate management of epidemic and pandemic outbreaks when resistant virus is circulating, and for the potential long term development of alternative inhibitors.

描述（申请人提供）：有两种特异性抗流感药物对所有流感株均有效，扎那米韦和奥司他韦。由于它们与病毒神经氨酸酶 (NA) 活性位点的高度保守残基结合，因此对所有流感病毒株均有效。随着高致病性H5N1禽流感在亚洲蔓延并现已蔓延到欧洲，出现大流行毒株的可能性更大。由于生产疫苗需要几个月的时间，因此正在储备 NA 抑制剂，尤其是奥司他韦，作为第一道防线。虽然在免疫功能正常的患者中尚未发现对扎那米韦的耐药性，但 NA 活性位点的保守残基中有 3 个常见突变，导致对奥司他韦耐药。然而，这些突变并不赋予所有 NA 亚型耐药性。目前奥司他韦治疗后的耐药率已高达18%，且已有H5N1奥司他韦耐药病毒的报道，因此人们对耐药性的担忧日益增加。通过神经氨酸酶抑制剂敏感性网络 (NISN) 全球监测计划，我们还发现了其他几种在活性位点之外的非保守残基中发生突变的病毒。因此，活性位点之外必定存在会影响药物结合的次级结构效应。因此，我们的中心假设是“NA 中额外的二级结构特征在确定药物敏感性以及突变是否会产生耐药性方面发挥着关键作用”。除了 NA 突变之外，我们还在其中一种分离株中发现了 HA 突变，这些突变在体外产生了耐药性。因此，我们还有一个小假设，即“临床分离株中的 HA 突变可能在体内对 NA 抑制剂的耐药性中发挥作用”。该项目的长期目标是利用结构和生物学研究来了解流感耐药机制，这些机制导致 N1/N2 药物敏感性差异、N1/N2 亚型特异性耐药性、活性位点外非保守残基的突变如何发生影响抑制剂结合，以及 HA 突变在体内耐药中的作用。我们的目标是从鸡蛋生长的或杆状病毒表达的蛋白质中确定野生型和突变型 NA 的结构。我们将评估 NA 和 HA 突变对体外和体内生长和药物敏感性的影响，作为病毒“适应性”的指标，以了解它们在临床情况下传播和耐药的可能性。相关性了解自然发生的或药物选择的变异体的耐药机制对于在耐药病毒传播时适当管理流行病和大流行病爆发以及替代抑制剂的潜在长期开发至关重要。