Development of Gallium-Based Therapies for Pulmonary Mycobacterial Infections

描述（由申请人提供）： 由结核分枝杆菌（Mycobacterium tuberculosis，M.tb）引起的结核病（tuberculosis，TB）是一个全球性的健康问题。对药物敏感的结核病的治疗需要使用多种潜在毒性抗生素进行数月的治疗。由于耐多药结核分枝杆菌的出现和日益流行，结核病的治疗变得更加困难。这项提案的重点是开发一种新的治疗方法，靶向结核分枝杆菌铁（Fe）的收购，这一过程是至关重要的能力，造成结核病。 疾病Ga（NO3）3是FDA批准的治疗恶性肿瘤高钙血症的药物。它通过与Fe竞争吸收/使用来干扰细胞Fe代谢。我们发现，镓还减少结核分枝杆菌铁的收购，抑制药物敏感和耐药结核分枝杆菌菌株的生长在体外细胞外和人类巨噬细胞内。Ga还在结核分枝杆菌感染的鼠模型中证明了功效。间歇性全身递送Ga（NO3）3几天在人体中显示出最小的毒性。然而，结核病的治疗可能需要更持久的治疗方案。改善Ga靶向感染的巨噬细胞、延长给药间隔或避免全身给药的方法将改善Ga的治疗指数。近年来，在将药物气溶胶递送至肺部以及实现这些目标的纳米颗粒的细胞靶向方面已经取得了相当大的进展。因此，我们假设通过气溶胶或纳米颗粒向肺递送Ga可以证明是新颖的、耐受性良好的且有效的TB疗法，其将通过破坏结核分枝杆菌Fe代谢而起作用。为了检验上述假设，我们提出实现以下具体目标：1。鉴定对细胞外和细胞内M.tb表现出最大体外效力的Ga形式，包括MDR-和XDR-TB，目的是获得先导化合物。2.开发两种铅镓化合物的制剂，使其能够通过吸入给药治疗肺结核，并确定其在肺结核小鼠模型中的生物分布、药代动力学、毒性和疗效。3.开发两种铅镓化合物的纳米颗粒制剂，使其能够用于治疗肺结核，并确定其在肺结核小鼠模型中的生物分布、药代动力学、毒性和疗效。4.确定镓铅制剂对结核分枝杆菌的作用机制、出现耐药性的可能性以及其他结核药物对镓抗分枝杆菌活性的影响。研究设计和方法：人单核细胞衍生的巨噬细胞和TB的鼠模型将与完全毒性的M. tb菌株一起沿着使用。这项工作将强调：结核分枝杆菌对铁和镓的获取：镓的生物分布、药代动力学、毒性;以及镓抗微生物活性的潜在细菌靶标。所采用的实验方法包括：开发适合于气溶胶或纳米颗粒介导的递送的Ga制剂;放射性标记的Fe和Ga的细菌摄取、结核分枝杆菌在体外和体内的生长、肺TB的鼠模型、组织中Ga的定量。对兽医保健的潜在影响：退伍军人患肺结核的风险很高。这项工作可能会导致开发新的结核病治疗方法，使退伍军人受益。

DESCRIPTION (provided by applicant): Tuberculosis (TB), caused by Mycobacterium tuberculosis (M.tb), is a global health problem. Treatment of drug-susceptible TB involves months of therapy with multiple potentially toxic antibiotics. Treatment of TB has become even more difficult by the emergence and increasing prevalence of multi-drug resistant M.tb. This proposal focuses on the development of a novel therapy that targets M.tb iron (Fe) acquisition, a process that is critical to its ability to cause disease. Ga(NO3)3 is a FDA- approved drug for treating hypercalcemia of malignancy. It interferes with cellular Fe metabolism by competing with Fe for uptake/use. We find that Ga also decreases M.tb Fe acquisition, inhibits growth of both drug-susceptible and drug-resistant M.tb strains growing in vitro extracellularly and within human macrophages. Ga also demonstrated efficacy in murine models of M.tb infection. Intermittent systemic delivery of Ga(NO3)3 for a few days has shown minimal toxicity in humans. However, treatment of TB would likely require more sustained treatment regimens. Ways to improve targeting of Ga to infected macrophages, extend dosing intervals, or avoid systemic administration would improve Ga's therapeutic index. There have been considerable advances in recent years in the aerosol delivery of drugs to the lung, as well as cellular targeting of nanoparticles to accomplish these goals. Therefore, we hypothesize that Ga delivery by aerosol or nanoparticle to the lung could prove to be novel, well-tolerated, and effective therapies for TB that would work via disruption of M.tb Fe metabolism. In order to test the above hypothesis we propose to accomplish the following specific aims: 1. Identify forms of Ga that exhibit maximal in vitro potency against extracellular and intracellular M.tb, including MDR- and XDR-TB, with the goal of arriving at lead compounds. 2. Develop formulations of two lead Ga compounds that will allow them to be administered by inhalation for the treatment of pulmonary TB and determine their biodistribution, pharmacokinetics, toxicity and efficacy in murine models of pulmonary TB. 3. Develop nanoparticle formulations of two lead Ga compounds that will allow them to be administered for the treatment of pulmonary TB and determine their biodistribution, pharmacokinetics, toxicity and efficacy in murine models of pulmonary TB. 4. Determine mechanisms of action of the lead Ga formulations against M.tb, potential for emergence of resistance, and effect of the presence of other TB drugs on Ga anti-mycobacterial activity. RESEARCH DESIGN AND METHODOLOGY: Human monocyte-derived macrophages and murine models of TB will be employed along with fully virulent strains of M.tb. The work will emphasize: Fe and Ga acquisition by M.tb: Ga biodistribution, pharmacokinetics, toxicity; and potential bacterial target for Ga antimicrobial activity. Experimental methods employed include: development of Ga formulations suitable for aerosol or nanoparticle-mediated delivery; bacterial uptake of radiolabeled Fe and Ga, growth of M.tb in vitro and in vivo, murine models of pulmonary TB, quantitation of Ga in tissues. POTENTIAL IMPACT ON VETERAN HEALTH CARE: Veterans are at high risk for pulmonary TB. This work could lead to development of novel treatments for TB that would benefit veterans.