Development of endochin-like quinolones for babesiosis therapy

人类巴贝斯虫病是一种类似疟疾的多系统疾病，主要由一种新出现的尖端复合体引起。 微小巴贝斯虫，在人类红细胞内感染和发育。寄生虫被传播到 人类通过扁虱传播，也可以通过输血传入肩部硬蜱。巴贝斯虫病 治疗选择包括两种抗疟疾药物的组合，阿奇霉素+阿托瓦酮或 克林霉素+奎宁。然而，即使有了这些药物和其他支持措施，如交换 输血，这种疾病在老年人或免疫功能低下的患者中可能是致命的。为了发展 对于人类巴贝斯虫病的更好的治疗和诊断测试，我们开发了一种短期的体外增殖 测试目前临床上使用的四种药物以及已知具有 对其他原生动物的活动。我们的研究表明内分泌素样喹诺酮类药物(ELQs)具有很强的活性。 和阿托瓦酮对微小巴贝斯虫的作用，并表明用于治疗巴贝斯虫病的其他药物的活性有限。在……里面 体内药效研究确定了2个化合物ELQ-271和ELQ-316对微小白僵菌有较强的抑制作用 LabS1株，灌胃给药10 mg/kg。然而，我们发现单一疗法包括以下两种 ELQ或阿托瓦酮在停药后9~16天常伴有复发， 强调需要确定这些化合物的更好的类似物或适当的治疗策略 导致根治。值得注意的是，我们发现由阿托瓦酮和前药组成的联合疗法 ELQ-316和ELQ-334在2.5+2.5的剂量下可根治小鼠的巴贝斯虫病 Mg/kg。生化分析表明，对B.microti细胞色素Bc1复合体的活性有抑制作用 ELQ，以及对ELQs或阿托瓦酮耐药的寄生虫的分析确定了B。 微细胞色素b(BmCytb)基因。在这些数据的基础上，我们提出以下三个具体目标。 在目标1中，我们将表征ELQ-316和30个结构不同的ELQ的活性和选择性。 生化分析将被用来表征它们对B.microti和人类Bc1复合体的活性， 细胞培养试验将被用来评估它们对寄生虫的疗效，并评估它们对 哺乳动物细胞。具有最大治疗指数的化合物将在#年进一步检查其效力。 小鼠和清除感染而不复发的能力。在目标2中，我们将合成这两个最主要的前药 有效的ELQ选自AIM 1。产生最大口服生物利用度的ELQ前药策略将是 与ELQ-334+阿托瓦酮联合应用进行对比试验。在目标3中，我们 将进行生化和遗传分析，以阐明ELQs的作用机制，确定可能的 药物失败的途径，并设计最佳治疗策略，以实现根治的理想配方 解药。拟议研究的成功将为未来的临床试验创造一种治疗方法奠定基础 人类巴贝斯虫病的治疗方案是安全的，并实现了根治。

Human babesiosis is a malaria-like multisystem disease caused primarily by an emerging apicomplexan parasite Babesia microti that infects and develops within human erythrocytes. The parasite is transmitted to humans by the tick vector, Ixodes scapularis and can also be introduced through blood transfusion. Babesiosis treatment options consist of two combinations of antimalarials, azithromycin+atovaquone or clindamycin+quinine. However, even with these drugs and other supportive measures such as exchange transfusion, the disease can be fatal in elderly individuals or immunocompromised patients. In order to develop better therapies and diagnostic tests for human babesiosis, we developed a short-term ex vivo proliferation assay to test the efficacy of the four currently clinically used drugs as well as compounds known to have activity against other protozoa. Our studies demonstrated potent activity of endochin-like quinolones (ELQs) and atovaquone against B. microti and indicated limited activity of other drugs used in babesiosis therapy. In vivo efficacy studies identified 2 compounds ELQ-271 and ELQ-316 with potent activity against the B. microti LabS1 strain following oral administration at 10mg/kg. However, we found that monotherapy involving either ELQs or atovaquone was always accompanied by recrudescence 9 to 16 days post-drug removal, emphasizing the need to identify better analogs of these compounds or a suitable therapeutic strategy that can lead to radical cure. Remarkably, we found that a combination therapy consisting of atovaquone and a prodrug of ELQ-316, ELQ-334, results in radical cure of experimental babesiosis in mice at doses as low as 2.5+2.5 mg/kg. Biochemical assays demonstrated inhibition of the activity of the B. microti cytochrome bc1 complex by ELQ, and analysis of parasites resistant to either ELQs or atovaquone identified single mutations in the B. microti Cytochrome b (BmCytb) gene. Building upon these data, we propose the following three specific aims. In Aim 1, we will characterize the activity and selectivity of ELQ-316 and 30 structurally diverse ELQs. Biochemical assays will be used to characterize their activity against the B. microti and human bc1 complex, and cell culture assays will be used to assess their efficacy against the parasite and evaluate their toxicity on mammalian cells. Compounds with the greatest therapeutic indices will be further examined for their potency in mice and ability to clear infection without recrudescence. In Aim 2, we will synthesize prodrugs of the 2 most potent ELQs selected from aim 1. The ELQ-prodrug strategy that yields the greatest oral bioavailability will be tested in combination with atovaquone in comparison to the ELQ-334+Atovaquone combination. In Aim 3, we will perform biochemical and genetic assays to elucidate the mechanism of action of ELQs, identify possible routes for drug failure and design the best therapeutic strategy for an ideal formulation that achieves radical cure. The success of the proposed studies will set the stage for future clinical trials to create a treatment regimen for human babesiosis that is safe and achieves radical cure.