Target based discovery of next generation pyrazinamide

基于目标的下一代吡嗪酰胺的发现

• 批准号: 10404533
• 负责人: Thomas Dick
• 金额: $ 79.17万
• 依托单位: HACKENSACK UNIVERSITY MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-08-01 至 2023-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10404533
• 关键词: Acquired Immunodeficiency Syndrome; Affinity; Anabolism; Animal Model; Anti-Bacterial Agents; Antibiotics; Antitubercular Agents; Antitubercular Antibiotics; Aspartate; Award; Bacillus; Bacteria; Binding; Biochemical; Biological Assay; Biophysics; Carboxy-Lyases; Cause of Death; Cells; Chemistry; Chimeric Proteins; Clinical; Dependence; Disease; Drug Kinetics; Drug Targeting; Drug Tolerance; Enzyme Inhibition; Frequencies; Generations; Genetic; Goals; Granuloma; Growth; HIV; HIV Seropositivity; Human; Immunity; Immunocompetent; In Vitro; Inbred BALB C Mice; Individual; Infection; Knowledge; Lead; Lesion; Mediating; Medical; Medicine; Minor; Modeling; Mutation; Mycobacterium tuberculosis; Necrosis; Necrotic Lesion; Oryctolagus cuniculus; Outcome; Patients; Penetration; Peptide Hydrolases; Persons; Pharmaceutical Preparations; Pharmacodynamics; Pharmacology; Pharmacotherapy; Play; Population; Property; Pyrazinamide; Pyrazinamide resistance; Regimen; Relapse; Reporter; Resistance; Role; Site; Sterilization; Testing; Time; Treatment Protocols; Tuberculosis; analog; bacterial fitness; base; biophysical analysis; biophysical techniques; chemotherapy; cofactor; enzyme activity; fight against; fitness; genetic approach; improved; in vitro activity; in vivo; inhibitor; lung lesion; metabolomics; molecular drug target; mouse model; mutant; next generation; novel; programs; protein degradation; pyrazinoic acid; rational design; treatment duration; tuberculosis drugs

Tuberculosis (TB) is a main cause of death for people living with AIDS. In immune-competent populations, drug therapy and immunity join forces to win the fight against Mycobacterium tuberculosis (Mtb). In HIV-positive individuals on the other hand, chemotherapy must fully sterilize all infection sites. There is an urgent medical need to develop more potent `sterilizing' drugs. The inclusion of pyrazinamide (PZA) into the TB regimen allowed reduction of treatment time to six months, while maintaining low relapse rates. The pharmacological basis for PZA's remarkable sterilizing activity in patients remains obscure, considering the drug's poor in vitro potency (MIC = 30-100 µg/mL). In the rabbit model of active TB, an animal model that recapitulates the various lung lesion types observed in human TB disease, we showed that PZA not only penetrates caseous necrotic lesions but also sterilizes them. In an ex vivo assay using caseum from infected rabbits, we showed that PZA kills non-growing, drug tolerant Mtb. These findings provide an explanation for the clinically observed treatment shortening effect of PZA: the drug reaches difficult-to-penetrate TB lesions and kills recalcitrant `persister' Mtb. However, consistent with the modest in vitro potency of PZA, onset of lesion sterilization is slow and concentrations required to kill Mtb in ex vivo caseum are high. Based on these findings, the logical way forward is to improve the potency of PZA while maintaining its unique lesion penetration and sterilization properties. To enable the rational optimization of PZA, we identified aspartate decarboxylase PanD, required for coenzymeA biosynthesis, as a the first genetically, biochemically and biophysically validated target of PZA. Consistent with poor whole cell activities of PZA, affinity of the drug (more specifically its bioactive component pyrazinoic acid, POA) for PanD was in the µM range, confirming room for improvement. Interestingly, mechanism of action studies revealed a novel antibacterial on-target mechanism whereby, rather than inhibiting PanD's catalytic activity, binding of POA to PanD appears to trigger degradation of the protein by the caseinolytic protease ClpC1. Here, we propose to build on our discoveries and 1. fully characterize the novel on-target mechanism by which the drug induces degradation of its target, 2. exploit in vitro pharmacological `sterilizing' models and PanD-based assays for the discovery of novel PZA analogs with improved potency and sterilizing activity, and 3. characterize lesion specific growth and replication status of PZA resistant panD mutant Mtb vs. wild type Mtb in the TB rabbit model with and without PZA treatment. In summary, we have identified key pharmacological properties responsible for the treatment shortening activity of PZA in clinical settings, as well as the molecular target of the drug, and we propose a `first-in-class to best-in- class' program to exploit these findings and rationally design the `next generation' PZA with improved sterilizing activity.

结核病（TB）是艾滋病患者的主要死亡原因。在免疫能力人群中，药物 治疗和免疫学联手赢得了与结核分枝杆菌（MTB）的战斗。在HIV阳性中 另一方面，化学疗法必须完全立体化所有感染部位。有紧急医疗 需要开发更多潜在的“消毒”药物。 将吡嗪酰胺（PZA）纳入结核病方案允许将治疗时间减少到六个月，而 保持低继电器率。 PZA在患者中显着的灭菌活性的药物基础 考虑到该药物的体外效力较差（MIC = 30-100 µg/ml），仍然保持晦涩难懂。在活动的兔子模型中 TB是一种概括人类结核病中观察到的各种肺部病变类型的动物模型，我们表明 PZA不仅穿透了案例坏死病变，还可以立体声。在使用caseum的离体测定中 从感染的兔子中，我们表明PZA杀死了非生长的耐药MTB。这些发现提供了 PZA临床观察到的治疗缩短效果的解释：该药物达到难以渗透率 结核病病变并杀死顽固的``毅力''mtb。但是，与PZA的体外效力相一致， 病变灭菌的发作缓慢，杀死MTB的浓度很高。基于 这些发现，前进的逻辑方法是提高PZA的效力，同时保持其独特的病变 穿透性和灭菌特性。为了实现PZA的合理优化，我们确定了天冬氨酸 脱羧酶PAND，是辅酶生物合成所需的，作为第一个遗传学，生化和 PZA的生物物理验证靶标。与PZA的全细胞活性不佳，药物的亲和力一致（更多 它的生物活性成分吡唑酸，poa），用于pand，在µm范围内，确认了空间的空间 改进。有趣的是，作用研究机制揭示了一种新型的抗菌攻击机制 因此，POA与PAND的结合似乎触发了降解，而不是抑制PAND的催化活性 酪蛋白蛋白质CLPC1的蛋白质蛋白质。 在这里，我们建议建立在我们的发现和1的基础上。充分表征了新颖的目标机制 该药物诱导其靶标降解，2。利用体外药理“灭菌”模型和基于PAND的模型 发现具有提高效力和消毒活性的新型PZA类似物的测定法，3。 PZA耐PZA PAND突变体MTB与野生型MTB的病变特异性生长和复制状态 进行有或没有PZA处理的模型。 总之，我们已经确定了负责治疗缩短活性的关键药物特性 PZA在临床环境中以及该药物的分子靶标的，我们提出了一流至最佳的一流 探索这些发现并合理设计的“下一代” PZA的课程“计划” 活动。

项目成果

A Ginger Root or Plum Model for the Tuberculosis "Granuloma"?

• DOI: 10.1164/rccm.202104-1052ed
• 发表时间: 2021-09-01
• 期刊: American journal of respiratory and critical care medicine
• 影响因子: 24.7
• 作者: Dartois V;Dick T
• 通讯作者: Dick T

PROTAC antibiotics: the time is now.

• DOI: 10.1080/17460441.2023.2178413
• 发表时间: 2023-04
• 期刊: EXPERT OPINION ON DRUG DISCOVERY
• 影响因子: 6.3
• 作者: Sarathy, Jickky Palmae;Aldrich, Courtney C.;Go, Mei-Lin;Dick, Thomas
• 通讯作者: Dick, Thomas

Targeted protein degradation in antibacterial drug discovery?

• DOI: 10.1016/j.pbiomolbio.2019.11.005
• 发表时间: 2020-05-01
• 期刊: PROGRESS IN BIOPHYSICS & MOLECULAR BIOLOGY
• 影响因子: 3.8
• 作者: Gopal, Pooja;Dick, Thomas
• 通讯作者: Dick, Thomas

Pyrazinamide Resistance Is Caused by Two Distinct Mechanisms: Prevention of Coenzyme A Depletion and Loss of Virulence Factor Synthesis.

• DOI: 10.1021/acsinfecdis.6b00070
• 发表时间: 2016-09-09
• 期刊: ACS infectious diseases
• 影响因子: 5.3
• 作者: Gopal P;Yee M;Sarathy J;Low JL;Sarathy JP;Kaya F;Dartois V;Gengenbacher M;Dick T
• 通讯作者: Dick T

Mycobacterium tuberculosis PanD Structure-Function Analysis and Identification of a Potent Pyrazinoic Acid-Derived Enzyme Inhibitor.

• DOI: 10.1021/acschembio.1c00131
• 发表时间: 2021-06-18
• 期刊: ACS chemical biology
• 影响因子: 4
• 作者: Ragunathan P;Cole M;Latka C;Aragaw WW;Hegde P;Shin J;Subramanian Manimekalai MS;Rishikesan S;Aldrich CC;Dick T;Grüber G
• 通讯作者: Grüber G