Acid Adaptation Targets for Eradication of Helicobacter pylori

描述(由申请人提供)： 幽门螺杆菌感染了世界上一半人口的胃。这种感染是消化性溃疡疾病的主要原因，也是胃癌的高危因素。因此，根除这种幽门螺杆菌感染会导致溃疡愈合，并降低患胃癌的风险。在幽门螺杆菌被发现之前，人们一直认为胃酸是细菌感染的不适宜环境。然而，幽门螺杆菌独一无二地发展了在人类胃的酸性表面生存和生长的方法，这一过程被称为酸驯化。破坏这一过程会导致在酸中存活的损失，因此这一过程的组成部分将为根除治疗提供新的靶点。目前的根除疗法需要在使用抑酸药物的同时使用抗生素。由于抗生素耐药性的增加，这些疗法的成功根除正在减少，导致美国的应答率为80%。酸适应是指有机体在有酸的情况下维持中性胞质内pH动态平衡的能力，从而允许胃部感染。细菌通过两个组分的组氨酸激酶信号系统感知和响应环境刺激。在幽门螺杆菌的情况下，通过HP0165/HP0166两组分系统启动环境酸化传感。因此，要探索的第一个假设是，对酸性的分级反应受HP0165/HP0166双组分系统的调节。该系统通过酸诱导尿素酶基因簇中两个启动子pureA和pureI的转录上调，酸诱导与尿素酶和Ni2+进入所需的尿素酶辅助蛋白组装的UreI翻译后上调，以及在中性pH下sRNAs转录后下调。尿素酶/尿素酶膜的组装对于有机体的酸驯化和酸生存是必不可少的，需要了解组装机制才能对这一过程进行靶向治疗。因此，我们将探讨HP0165/HP0166 TCS在尿素酶的膜募集、组装和激活中的作用，在调节SRNA调控尿素酶表达中的作用，以及TCS对pH的感知。依赖生长的抗生素克拉霉素或阿莫西林除非细菌处于生长阶段，否则无效。改善酸抑制应能改善治疗结果。在第二个目标中，我们假设抑制酸分泌增加了生长阶段的细菌数量，使它们对生长依赖的抗生素敏感，这就要求除了成功根除抗生素外，还必须使用抑酸化合物。仅用奥美拉唑和阿莫西林，缓慢的奥美拉唑代谢物对酸的控制和有效的根除效果显著改善，这一发现表明，改善的酸抑制增加了有机体对生长依赖抗生素的敏感性。由此可见，随着胃内pH值的更一致和更大的升高，更多的微生物处于抗生素敏感生长期，而不是耐药静止期。白天和晚上保持较高的pH值应该会使几乎所有的幽门螺杆菌都处于生长阶段。因此，作为细菌细胞生长指标的酸抑制对幽门螺杆菌蛋白质合成速率的影响将在体外和体内进行测量。 公共卫生相关性： 幽门螺杆菌感染在消化性溃疡疾病中起因果作用，并增加患胃癌的风险。消化性溃疡疾病和胃癌是美国和世界各地退伍军人人口发病率的主要原因。幽门螺杆菌生活在胃中，并进化出独特的酸反应机制，使胃得以定植。使用酸阻断药物和抗生素的标准三联或四联疗法已经变得不那么有效，由于抗生素耐药性的增加，成功率降至80%以下。了解幽门螺杆菌如何在胃内定植将有助于开发新的根除疗法。取代目前疗法的一个可能的策略是将允许细菌在含有非抗生素化合物的酸性环境中生存的基因作为靶点。另一种可能的策略是改善细菌环境的pH控制，以便可以使用酸抑制和阿莫西林的双重治疗。这将涉及开发具有更好的阻酸性能的药物。

DESCRIPTION (provided by applicant): Helicobacter pylori infects the stomach of half of the world's population. This infection is a primary cause of peptic ulcer disease and a high risk factor for gastric cancer. Therefore, eradication of this H. pylori infection leads to ulcer healing and lowers the risk of gastric cancer. Until the discovery of H. pylori, it was thought that stomach acid presented an inhospitable environment for bacterial infection. However, H. pylori has uniquely developed the means of surviving and growing on the acidic surface of the human stomach, a process termed acid acclimation. Disruption of this process leads to loss of survival in acid, therefore the components of this process would provide novel targets for eradication therapy. Current eradication therapies require the use of antibiotics along with acid inhibitory drugs. The successful eradication by these therapies is decreasing due to increasing antibiotic resistance, resulting in a response rate of < 80% in the USA. Acid acclimation is the ability of the organism to maintain a neutral periplasmic pH homeostasis in the presence of acid to allow gastric infection. Bacteria sense and respond to environmental stimuli via two component histidine kinase signaling systems. In the case of H. pylori, environmental acidification sensing is initiated through the HP0165/HP0166 two component system. Accordingly, the first hypothesis to be explored is that graded responses to acidity are regulated by this HP0165/HP0166 two component system. The system responds by acid-induced transcriptional upregulation of two promoters in the urease gene cluster, pureA and pureI, acid-induced post-translational upregulation of assembly UreI with apourease and the urease accessory proteins required for Ni2+ insertion along with Ni2+ entry, and also post-transcriptional down regulation by sRNAs at neutral pH. The UreI/urease membrane assembly is essential for acid acclimation and acid survival of the organism, and an understanding of the assembly mechanism is required for targeted therapy to this process. Therefore, the role of the HP0165/HP0166 TCS in membrane recruitment, assembly and activation of urease, in regulation of sRNA control of urease expression, and pH sensing by the TCS will be explored. The growth dependent antibiotics clarithromycin or amoxicillin are not effective unless bacteria are in growth phase. Improving acid inhibition should improve therapeutic outcome. In the second aim, we hypothesize that inhibition of acid secretion increases the number of bacteria in growth phase, making them sensitive to growth dependent antibiotics necessitating the requirement for administration of acid inhibitory compounds, in addition to antibiotics for successful eradication. The finding that slow omeprazole metabolizers respond with much improved acid control and effective eradication with omeprazole and amoxicillin alone, suggests that improved acid inhibition increases the sensitivity of the organism to growth dependent antibiotics. From this it follows that, with more consistent and greater elevation of intragastric pH, more organisms are in the antibiotic sensitive growth phase rather than the resistant stationary phase. Maintenance of an elevated pH for both day and night should put almost all H. pylori in growth phase. Therefore, the effect of acid inhibition on the rate of H. pylori protein synthesis as an indicator of bacterial cell growth will be measured in vitro and in vivo. PUBLIC HEALTH RELEVANCE: H. pylori infection has a causal role in peptic ulcer disease and increases the risk of gastric cancer. Peptic ulcer disease and gastric cancer are major causes of morbidity in the Veterans population, in the United States and worldwide. H. pylori inhabits the stomach and has evolved unique acid response mechanisms to allow gastric colonization. Standard triple or quadruple therapies with acid blocking medicines and antibiotics have become less effective, falling below 80% success rate due to increasing antibiotic resistance. An understanding of how H. pylori colonizes the stomach will aid the development of novel eradication therapies. A possible strategy to replace current therapy is to target genes that allow the bacteria to survive in acid with non-antibiotic compounds. Another possible strategy is to improve control of the pH of the bacterial environment, so that dual therapy with acid inhibition and amoxicillin could be used. This would involve development of drugs with better acid blocking properties.