The use of mass spectrometry for rapid detection of carbapenemase-producing bacte

描述(申请人提供)：现代医学的实践在很大程度上依赖于有效的抗生素治疗来对抗细菌感染。如果没有它们，许多用于抗击癌症和抑制慢性炎症的手术和化疗方案将根本不可能实现。多药耐药(MDR)细菌菌株的发展有可能破坏这些进展，并代表着一个重大的全球卫生挑战。“革兰氏阴性杆菌”(GNR)细菌的一种常见耐药机制是分泌一种β-内酰胺酶，该酶能分解青霉素类所有成员共有的β-内酰胺环。针对多种β-内酰胺类抗生素感染的治疗，已进化出数百种不同的β-内酰胺酶，它们的催化机制和底物专一性各不相同。碳青霉烯类抗生素是对抗耐多药核糖核酸酶的最后一道有效防线，该酶表达超广谱β-内酰胺酶(ESBLS)，能够降解所有非碳青霉烯类β-内酰胺类抗生素。然而，它们的有效性已经受到最近出现的质粒编码碳青霉烯酶的影响。为了有效地控制和/或预防碳青霉烯类耐药细菌感染的暴发，并提供最佳的患者护理，快速检测临床标本中的碳青霉烯酶活性势在必行。我们已经开创了一种依赖于质谱学的快速表型方法来检测碳青霉烯酶活性，方法是选择性地监测碳青霉烯酶解产物的外观。在这项应用的第一个目标中，我们将改进和优化我们目前基于MS的方法，以检测碳青霉烯类抗生素的β-内酰胺环水解。我们将确定必要的试剂盒组件和检测细菌分离株中碳青霉烯类抗生素水解酶活性的检测方法。在这项应用的第二个目的中，我们将使用从耶鲁-纽黑文医院挑选的回顾和预期的临床标本进行盲法临床研究，以建立整个方案的生物学敏感性和特异性。未来的开发阶段将把这项技术应用于初级样本，随后将进行多中心临床试验，以争取FDA 510(K)批准最终产品。

DESCRIPTION (provided by applicant): The practice of modern medicine relies heavily on effective antibiotic therapy to combat bacterial infections. In their absence, many surgeries and chemotherapeutic regimens used to combat cancer and suppress chronic inflammatory conditions would simply not be possible. Development of multidrug-resistant (MDR) bacterial strains threatens to undermine these advances and represents a major global health challenge. A common resistance mechanism in "gram negative rod" (GNR) bacteria is secretion of a beta-lactamase that hydrolyzes the beta-lactam ring common to all members of the penicillin class. Hundreds of diverse beta-lactamases, which vary in their catalytic mechanism and substrate specificity, have evolved in response to treatment of infections with multiple b-lactam antibiotics Carbapenems represent the last line of effective defense against MDR GNRs expressing extended spectrum beta-lactamases (ESBLs) capable of hydrolyzing all non-carbapenem beta-lactams. However, their effectiveness has been compromised by the recent emergence of plasmid-encoded carbapenemases. To effectively contain and/or prevent outbreaks of carbapenem resistant bacterial infections and to offer optimal patient care, rapid detection of carbapenemase activity in clinical specimens is imperative. We have pioneered a rapid, phenotypic method relying on mass spectrometry to detect carbapenemase activity, by selectively monitoring for the appearance of carbapenem hydrolysis products. In the first aim of this application, we will refine and optimize our current MS- based assay to detect ¿-lactam ring hydrolysis of carbapenem antibiotics. We will define necessary kit components and assay protocols for detection of carbapenem hydrolysis activity in bacterial isolates. In the second aim of this application, we will perform a blinded clinical study using selected retrospective and prospective clinical specimens from Yale-New Haven Hospital in order to establish biologic sensitivity and specificity of the overall protocol. Future phases of development will apply the technology to primary specimens, which will be followed by multicenter clinical trials in pursuit o FDA 510(k) clearance of the final product.