Overcoming Pyrazinamide Resistance with Pyrazinoate-Cephalosporin Conjugates

用吡嗪酸-头孢菌素缀合物克服吡嗪酰胺耐药性

• 批准号: 9895968
• 负责人: Courtney C Aldrich
• 金额: $ 22.96万
• 依托单位: UNIVERSITY OF MINNESOTA
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-01-25 至 2021-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9895968
• 关键词: AIDS-Related Opportunistic Infections; Atypical Mycobacteria; Bacillus; Basic Science; Bioavailable; Biochemical; Caco-2 Cells; Cephalosporins; Clinical; Communicable Diseases; Data; Development; Drug Kinetics; Drug resistance; Enzymes; Esters; Etiology; Future; Genetic; Human; Immunology; In Vitro; Infection; Infectious Diseases Research; Liquid substance; Measures; Mediating; Microbiology; Minnesota; Modeling; Mus; Mycobacterium tuberculosis; Oral; Patients; Permeability; Pharmaceutical Chemistry; Pharmaceutical Preparations; Population; Predisposition; Prodrugs; Pyrazinamide; Pyrazinamide resistance; Relapse; Research; Resistance; Serum; Site; Stomach; Treatment Failure; Treatment Protocols; Tuberculosis; Universities; Work; amidase; analog; antimicrobial; beta-Lactamase; beta-Lactams; collaborative environment; commensal microbes; drug discovery; drug disposition; esterase; gut microbiome; in vivo; loss of function mutation; macrophage; microorganism; mortality; mycobacterial; novel; pathogen; preclinical development; pyrazinoic acid; research facility; therapy duration; tuberculosis chemotherapy; tuberculosis drugs; tuberculosis treatment

SUMMARY Mycobacterium tuberculosis (Mtb), the principal etiological agent of tuberculosis (TB), infects over one-third of humanity and is now the leading cause of infectious disease mortality by a single pathogen. Pyrazinamide (PZA) is a first-line sterilizing anti-tubercular drug that is anticipated to be an irreplaceable component of future TB treatment regimens. Previous studies have demonstrated that PZA is a pro-drug which is converted to the active form of pyrazinoic acid (POA) by the M. tuberculosis amidase PncA, and that loss of function mutations in pncA account for the vast majority of PZA resistance. Resistance to PZA in turn is associated with significantly higher treatment failures and relapse rates. Attempts to directly administer POA in humans and mice have been unsuccessful because POA cannot attain sufficient concentrations within Mtb at the infection site. Prodrug approaches employing simple POA esters have been unable to overcome the intrinsic drug disposition liabilities of POA as conventional prodrugs are rapidly hydrolyzed by serum esterases. In preliminary data, we have developed a novel prodrug strategy for POA through conjugation to a β-lactam promoiety leading to selective release of POA by the unique mycobacterial beta-lactamase BlaC and not by serum esterases. This strategy overcomes PZA-resistance by circumventing PncA-mediated activation and renders blaC conditionally essential since loss of function mutations to blaC, restores susceptibility of Mtb to the β-lactam promoiety. Consequently, we hypothesize the genetic barrier for development of spontaneous resistance to our POA-β-lactam conjugates will be high. The objectives of this application are to synthesize and evaluate β-lactam prodrugs of POA, which are orally bioavailable and selectively release POA within Mtb- infected macrophages. We will accomplish the overall objectives of this application by pursuing two specific aims. In aim, we will synthesize β-lactam prodrugs of POA, which are selectively activated by Mtb to release POA and not by commensal microbiota. The prodrugs will be biochemically and microbiologically characterized with a panel of β-lactamases and microorganisms. In aim 2, we will measure prodrug stability in serum and stimulated gastric fluid and permeability in Caco-2 cells. We will next determine complete pharmacokinetic parameters and the extent of POA release by the gut microbiome. Finally, we plan to evaluate one compound in a murine TB infection model.

总结 结核分枝杆菌（Mtb）是结核病（TB）的主要病原体，感染超过三分之一的 目前，单一病原体是导致传染病死亡的主要原因。Pyrazinamide (PZA)是一线杀菌抗结核药物，有望成为未来抗结核药物不可替代的组成部分， 结核病治疗方案。先前的研究已经证明，PZA是一种前体药物， 活性形式吡嗪酸（POA）的M。结核病酰胺酶PncA的功能缺失突变 pncA中的大多数占PZA抗性的绝大多数。对PZA的耐药性反过来又与 治疗失败率和复发率明显更高。尝试在人类中直接施用POA， 小鼠一直不成功，因为POA在感染时不能在Mtb内达到足够的浓度 绝佳的价钱采用简单的POA酯的前药方法已经不能克服内在药物的缺点。 POA作为常规前药的处置负债被血清酯酶迅速水解。在 初步数据，我们已经开发了一种新的POA前药策略，通过共轭β-内酰胺 导致POA被独特的分枝杆菌β-内酰胺酶BlaC选择性释放，而不是被 血清酯酶。该策略通过绕过PncA介导的活化来克服PZA抗性， 由于blaC的功能丧失突变，使得blaC条件性必需，恢复Mtb对 β-内酰胺前体部分。因此，我们假设自发性糖尿病发生的遗传障碍 对我们的POA-β-内酰胺缀合物的抗性将是高的。本申请的目的是综合和 评价POA的β-内酰胺前体药物，这些药物具有口服生物利用度，并在Mtb内选择性释放POA。 感染的巨噬细胞我们将通过追求两个具体目标来实现本申请的总体目标 目标。本论文的目标是合成POA的β-内酰胺类前体药物， POA而不是肠道微生物群。前药将进行生物化学和微生物学表征 一组β-内酰胺酶和微生物在目标2中，我们将测量前药在血清中的稳定性， 刺激胃液和Caco-2细胞的渗透性。我们接下来将确定完整的药代动力学 参数和肠道微生物组释放POA的程度。最后，我们计划评估一种化合物， 在小鼠TB感染模型中。