Fighting AMR of hard-to-reach microbial pathogens by repurposing antibiotics using a targeted liposomal delivery strategy: A Helicobacter pilot study

背景：抗菌素耐药性(AMR)是对全球健康的主要威胁，特别是在低收入国家，那里获得抗生素的限制较少，导致经常过度使用。这反过来又导致以前有效的抗生素越来越多地无法根除感染。幽门螺杆菌(Hp)是一种细菌，感染越南80%的成年人和70%的儿童。相比之下，英国许多地区的患病率为15%。AMR菌株的日益流行是控制幽门螺杆菌失败的关键原因，导致医疗费用增加，相关疾病的死亡率和发病率增加。越南2015年的一项研究发现，42.4%的菌株对克拉霉素耐药，41.3%对左氧氟沙星耐药，76.1%对甲硝唑耐药。Oda依从性：幽门螺杆菌是胃肠道溃疡和胃癌的主要原因。越南的高发病率和高再感染率被认为是由于其较低的社会经济地位，例如拥挤的生活和恶劣的卫生条件。现在，低收入国家和富裕国家之间在幽门螺杆菌和相关疾病(如胃癌)的患病率方面存在巨大差异。后者在越南最常见的癌症中排名第四，但在英国仅排名第16。幽门螺杆菌对人口中较脆弱的部分的影响尤其强烈；癌症(例如，由幽门螺杆菌引起的胃癌)是造成贫困的一个重要原因，与其在越南人口脆弱部分的治疗费用有关，将许多家庭推向贫困。因此，我们的项目旨在培训越南科学家和临床医生在尖端先进的药物输送方面，使越南卫生系统能够开发并最终向普通民众提供这种新的治疗方法。因此，根据官方的发展援助原则，这可能会给越南较贫穷的人群带来长期的社会经济利益。抗菌治疗未能根除HP感染的一个原因是，药物在厚厚的粘液下和隐窝内对细菌的可及性很低。进一步的因素是药物在胃内的滞留时间很短，以及一些抗生素对胃酸的敏感性。因此，一些在体外对幽门螺杆菌高效的抗生素在体内失败了。解决这两个问题，通过增加胃滞留时间和保护抗生素免受胃酸的影响，将提高根除Hp感染的治疗效果，同时允许使用目前无法使用的现有抗生素。改变抗生素的用途是加快药物开发进程的一种非常具有成本效益的战略。我们的长期目标是通过降低胃肠道溃疡和胃癌的发病率，为改善越南人民的健康做出贡献。我们建议使用功能化脂质体来解决胃滞留时间短和胃酸失活的问题。由于表面添加了两种幽门螺杆菌粘附素(BABA和LABA)，这些粘附素将附着在胃上皮和粘液上。包裹在功能化脂质体中的药物的胃滞留增加将使用NanoSPECT-CT成像在体内得到验证，我们已经在小鼠身上成功地进行了测试。功能化脂质体与上皮细胞的结合将通过测量稳定转染人MUC5AC的人胃细胞株AGS的结合来评估，MUC5AC是一种粘蛋白，含有用于脂质体功能化的BABA和LABA的生理配体。对来自英国和越南的约600株Hp临床分离株进行筛选，将使我们能够识别不同水平的AMR，并使我们能够在体外测试我们功能化脂质体的有效性和安全性(例如细胞毒性)。最后，将利用建立的小鼠感染模型在体内评估该制剂根除幽门螺杆菌的能力。

Background: Antimicrobial resistance (AMR) is a major threat to global health, particularly in low-income countries, where access to antibiotics is less restricted, leading to frequent overuse. This in turn leads to increasing failure of previously efficacious antibiotics to eradicate infection. H. pylori (Hp) is a bacterium infecting 80% of adults and 70% of children In Vietnam. In comparison, prevalence is <15% in many parts of the UK. The increasing prevalence of AMR strains is the crucial cause for failure in controlling Hp, leading to higher medical costs, and increased mortality and morbidity from related diseases. A 2015 study in Vietnam found 42.4% strains resistant to clarithromycin, 41.3% to levofloxacin, and 76.1% to metronidazole. ODA compliance: Hp is the main cause of gastrointestinal ulcers and stomach cancer. The high prevalence of disease and a high rate of reinfection in Vietnam is thought to depend on the low socioeconomic status, e.g. crowded living and poor hygienic conditions. There is now a huge discrepancy between low income and affluent countries in terms of Hp prevalence and related diseases, such as gastric cancer. The latter is the 4th most common type of cancer in Vietnam, but only the 16th in the UK. The impact of Hp is particularly strong on the more vulnerable sections of the population; cancer (e.g. gastric cancer caused by Hp) is a significant cause of impoverishment associated with the costs of its treatment in vulnerable sections of the Vietnamese population, pushing many households into poverty. Therefore, our project, which is designed to train Vietnamese scientists and clinicians in cutting-edge advanced drug delivery, empowering the Vietnamese health system to develop and ultimately make available this new treatment to the general population. Hence it is likely to lead to long term socioeconomic benefits to poorer sections of the Vietnamese population, in line with official development assistance principles.One cause underlying the failure of antimicrobial treatment to eradicate Hp infection is the low accessibility of the drug to the bacterium, underneath the thick mucus and in crypts. Further factors are the short retention times of drugs in the stomach, and the susceptibility of some antibiotics to stomach acid. As a result, some antibiotics which are highly effective against Hp in vitro have failed in vivo. Solving these two problems, by increasing the gastric retention time and protecting the antibiotics from the stomach acid, would increase efficacy of treatment in eradicating Hp infection while allowing the use of existing antibiotics which currently cannot be used. Repurposing of antibiotics is a very cost-effective strategy to fast-track the drug development process. Our long term aim is to contribute to improve the health of the population in Vietnam by reducing the prevalence of GI ulcers and gastric cancer. We suggest solving the problem of short gastric retention times and gastric acid inactivation by using functionalized liposomes. These will adhere to the gastric epithelium and mucus due to the addition of two Hp adhesins (BabA and LabA) on their surface. The increased gastric retention of drugs encapsulated in the functionalized liposomes will be verified in vivo using NanoSPECT-CT imaging, which we have successfully tested in mice. Binding of functionalized liposomes to epithelial cells will be assessed in vitro by measuring binding to human gastric cell line AGS stably transfected with human MUC5AC, a mucin which contains the physiological ligands of BabA and LabA used for liposome functionalization. Screening of ~600 clinical Hp isolates from both the UK and Vietnam will allow us to identify various levels of AMR and allow us to test the efficacy and safety (e.g. cell toxicity) of our functionalized liposomes in vitro. Finally, the ability of this formulation to eradicate Hp will be assessed in vivo using am established murine infection model.