Highly specific anti-toxin therapies for severe bacterial gut infections

针对严重细菌性肠道感染的高度特异性抗毒素疗法

• 批准号: 9055644
• 负责人: Brett D Welch
• 金额: $ 29.74万
• 依托单位: NAVIGEN, INC.
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-05-01 至 2017-10-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9055644
• 关键词: Acute; Affinity; Amino Acids; Antibiotics; Antibodies; Applications Grants; Avidity; Bacteriophages; Binding; Biochemical; Bioinformatics; Biological Assay; Blood Circulation; Categories; Cell Surface Proteins; Cells; Centers for Disease Control and Prevention (U.S.); Child; Childhood; Circular Dichroism; Clinic; Communicable Diseases; Complication; Consensus; Consensus Sequence; Coupled; Developing Countries; Disease Outbreaks; Diversity Library; Drug Design; Ebola virus; Endocytosis; Enzyme-Linked Immunosorbent Assay; Escherichia coli; Escherichia coli EHEC; Escherichia coli Infections; Escherichia coli O157:H7; Escherichia coli Vaccines; Face; Food; Food Contamination; Food Poisoning; Fruit or Vegetable; Grant; HIV; Health; Hemolytic-Uremic Syndrome; Hemorrhagic colitis; Image; In Vitro; Individual; Infection; Kidney Failure; L Forms; Lead; Libraries; Life; Measures; Membrane Glycoproteins; Molecular Conformation; Pain; Peptide Hydrolases; Peptides; Phage Display; Phase; Positioning Attribute; Preclinical Drug Evaluation; Production; Protein Engineering; Proteins; Random Peptide Libraries; Resolution; Risk; Route; Sampling; Severities; Shiga Toxin; Shiga-Like Toxin I; Shigella dysenteriae; Source; Specificity; Structure; Supportive care; Surface Plasmon Resonance; Technology; Therapeutic Uses; Toxicology; Toxin; Viral; Virulence Factors; Virulent; Water; X-Ray Crystallography; animal efficacy; base; combat; contaminated water; cross reactivity; deep sequencing; design; drug discovery; effective therapy; env Gene Products; foodborne; global health; immunogenicity; in vitro activity; in vivo; inhibitor/antagonist; innovation; mutant; novel; pathogen; pathogenic bacteria; peptide L; peptide drug; preclinical trial; prevent; public health priorities; receptor binding; screening; small molecule; success; waterborne

 DESCRIPTION (provided by applicant): Shiga toxins 1 and 2 (Stx1/2) are the primary virulence factors of Shiga toxin-producing E. coli (STEC), which are major food-, and waterborne pathogens afflicting both developed and developing countries. An estimated 265,000 STEC infections occur annually in the US. In addition to severe and often bloody diarrhea, the life-threatening complication hemolytic uremic syndrome (HUS) occurs in 2-7% of victims (primarily children). No STEC vaccines or therapies beyond supportive care are available. Antibiotics are not used since they can increase toxin production and risk of HUS. The severity of STEC infection, propensity for outbreaks, and lack of effective treatments make STEC concerning potential bioterror agents and a public health priority. Here, we describe an innovative strategy to discover D-peptide inhibitors of Stx1/2 to combat STEC infection. D-peptides, the mirror images of natural L-peptides, cannot be digested by proteases and, therefore, have the potential for long in vivo half-lives and low immunogenicity. They can readily disrupt protein interfaces with high potency and specificity compared to small molecules and are much less expensive to produce than antibodies. D-peptides are ideal candidates for Stx1/2 neutralization in the gut and/or systemic circulation. Navigen's drug discovery platform employs an enantiomeric screening technology (mirror-image phage display) coupled with protein design. We have successfully validated this platform technology by identifying D- peptide inhibitors of HIV, RSV, and Ebola. Our anti-HIV D-peptide was the first potent and specific D-peptide inhibitor to be discovered and is in advanced preclinical trials. It binds a functionally critical and conserved hydrophobic "pocket" on HIV's trimeric surface glycoprotein, gp41. A trimeric version of this D-peptide binds to all three gp41 pockets, providing a strong avidity boost. Stx1/2 is each composed of a single enzymatically active A subunit and five receptor-binding B subunits that form a pentameric ring. Each of the B pentamer subunits contains a vulnerable pocket analogous to those of our viral targets. The Stx1/2 B subunit pockets are excellent targets for us to next apply our expertise in D- peptide drug design given 1) our success in targeting analogous pockets at functionally critical interfaces, 2) the pentameric Stx1/2 target, which inspires design of pentameric D-peptides with strong avidity, and 3) the likelihood of D-peptide stability and activity in the gut without disturbing native flora. Furthermore, the B subunits of Stx1 and Stx from Shigella dysenteriae are identical, enabling dual therapeutic use for an anti- Stx1B D-peptide. These benefits have generated strong enthusiasm for this project from GI infectious disease clinicians, who understand the dramatic potential impact an anti-Stx1/2 D-peptide would have in the clinic. In this two-year grant, we propose to discover, structurally characterize, and optimize pentameric D-peptide inhibitors that will neutralize Stx1/2 with high potency. Success in this project will launch a new class of inhibitors against pathogenic bacteria important to global health.

 说明(申请人提供)：志贺毒素1和2(STX1/2)是产志贺毒素的大肠杆菌(STEC)的主要毒力因子，STEC是困扰发达国家和发展中国家的主要食源性和水源性病原体。据估计，美国每年有26.5万人感染STEC。除了严重且经常是血性腹泻外，2-7%的受害者(主要是儿童)还会出现危及生命的溶血性尿毒症综合征(HUS)。除了支持性护理之外，没有STEC疫苗或治疗方法可用。抗生素不被使用，因为它们会增加毒素的产生和HUS的风险。STEC感染的严重性、暴发的倾向以及缺乏有效的治疗方法使STEC成为潜在的生物恐怖因子和公共卫生优先事项。在这里，我们描述了一种创新的策略来发现STX1/2的D-肽抑制剂来对抗STEC感染。D-肽是天然L多肽的镜像，不能被酶消化，因此具有较长的体内半衰期和低免疫原性。与小分子相比，它们可以很容易地破坏蛋白质界面，具有更高的效力和特异性，而且生产成本比抗体低得多。D-肽是肠道和/或体循环中STX1/2中和的理想候选者。Navigen的药物发现平台采用了一种结合蛋白质设计的对映体筛选技术(镜像噬菌体展示)。我们已经通过识别艾滋病毒、呼吸道合胞病毒和埃博拉病毒的D-肽抑制剂成功地验证了这一平台技术。我们的抗HIV D-肽是第一个被发现的有效和特异的D-肽抑制剂，正在进行高级临床前试验。它在HIV的三聚体表面糖蛋白gp41上结合了一个功能关键且保守的疏水“口袋”。这种D-肽的一个三聚体版本与所有三个gp41口袋结合，提供强大的亲和力提升。STX1/2每个都由一个具有酶活性的A亚基和五个与受体结合的B亚基组成，形成一个五聚环。每个B五聚体亚基都有一个脆弱的口袋，类似于我们的病毒靶标。STX1/2 B亚单位口袋是我们下一步应用我们在D-肽药物设计方面的专业知识的极佳目标，因为1)我们成功地针对功能关键界面的类似口袋，2)五聚体STX1/2靶，它激发了具有强大亲和力的五聚体D-肽的设计，以及3)D-肽在肠道中稳定和活性而不干扰本地菌群的可能性。此外，志贺氏菌的STX1和STX的B亚基是相同的，这使得抗STX1B D肽能够双重治疗。这些好处激发了胃肠道传染病临床医生对这一项目的强烈热情，他们了解抗STX1/2 D肽在临床上将产生的巨大潜在影响。在这项为期两年的拨款中，我们建议发现、结构表征和优化将以高效力中和STX1/2的五聚体D-肽抑制剂。该项目的成功将推出一类对全球健康重要的病原菌抑制物。