Targeting Cyclic Dinucleotide Signaling Pathways to Interrupt the Nature Cycle of Borrelia burgdorferi

莱姆病已成为美国主要的公共卫生威胁。目前，还没有可用的人类疫苗。 虽然用化学杀螨剂瞄准硬壳类动物是降低感染风险的有效方法，但 使用杀螨剂会对健康造成不利影响，并引起环境问题。以主机为目标的方法 现场投放载多西环素的毒饵可显著降低两种啮齿动物的感染率。 莱姆病病原伯氏疏螺旋体寄生于水库和肩部硬蜱。然而，抗生素如 因为多西环素不适合用于这种目的，因为它们用于治疗病人。然而，这样的证据证明 概念研究表明，使用替代化合物的宿主靶向方法是一种很有前途的方法 消灭鼠类宿主和壁虱媒介中的螺旋体。细菌第二信使c-di-GMP和 C-di-AMP，已成为细菌生理的中心调节剂，也是潜在的药物靶点。许多细菌 为了合成c-di-GMP和c-di-AMP，编码多个拷贝的环化酶，这使得靶向变得困难 那些小路。伯氏疏螺旋体只有一个二鸟苷环化酶Rrp1和一个二腺苷环化酶 CDAA用于c-二-AMP的合成，这使它们成为有吸引力的药物靶点。如初步数据所示，我们发现 这两个环二核苷酸，一个控制伯氏杆菌的两个宿主中的螺旋体的生存。 地方性循环，壁虱和哺乳动物：虽然c-di-GMP对于壁虱的定植是必不可少的，但c-di-AMP对于 哺乳动物感染。这一提议的假设是，靶向c-di-GMP和c-di-GMP的小分子抑制剂 AMP环化酶，Rrp1和CDAA，可以消除扁虱和哺乳动物宿主中的伯氏杆菌，这可能是 用来减少莱姆病的发病率。这项提议的联合作者赫尔曼·辛蒂姆博士，药物发现 化学教授，率先开发细菌二鸟苷环化酶和二腺苷环化酶抑制剂， 他的团队已经报道了几种有效的抑制其他细菌环化酶的药物。因此， 我们建议开发一种针对伯氏杆菌CDAA的策略，以消灭鼠类水库中的螺旋体(AIM) 1)，以及以伯氏杆菌Rrp1为靶点的战略，以消除硬蜱中的螺旋体(目标2)。我们还将测试一个 针对这两条途径的抑制剂组合，在伯氏杆菌的地方性循环中消灭它。潜在的 伯氏杆菌如何利用c-di-AMP和c-di-GMP在哺乳动物和壁虱体内生存的机制 也在调查中。该提案涉及本RFA中列出的特定重点领域之一：以水库为目标 中断感染自然历史的方法。在完成拟议的工作后，进一步的领域 实验将使用含有两种抑制剂组合的动物诱饵来评估它们的效果。 关于减少自然界中的扁虱和动物的螺旋体负担。这种以宿主为目标战略将产生重大影响 与莱姆病作斗争。

Lyme disease has emerged as a major public health threat in the US. Currently, no human vaccine is available. Although targeting tick populations with chemical acaricides is an effective way of reducing the risk of infection, the use of acaricides has adverse health effects and raises environmental concerns. A host-targeted approach by deploying doxycycline hyclate-laden baits to the field showed dramatic reduction of infection rates in both rodent reservoirs and Ixodes scapularis ticks by Borrelia burgdorferi, the agent of Lyme disease. However, antibiotics such as doxycycline are not suitable for such purpose as they are used for treating patients. Nevertheless, such proof of concept study demonstrates that a host-targeted approach using alternative compounds is a promising approach to eliminate spirochetes in rodent reservoir hosts and tick vectors. The bacterial second messengers, c-di-GMP and c-di-AMP, have emerged as central regulators for bacterial physiology and are potential drug targets. Many bacteria encode multiple copies of cyclases for the synthesis of c-di-GMP and c-di-AMP, which make it difficult to target the pathways. Borrelia burgdorferi only has a single diguanylate cyclase Rrp1 and a single diadenylate cyclase CdaA for c-di-AMP synthesis, which makes them attractive drug targets. As shown in the preliminary data, we found that these two cyclic dinucleotide, one controls spirochetes’ survival in each of the two hosts in B. burgdorferi enzootic cycle, ticks and mammals: while c-di-GMP is essential for tick colonization, c-di-AMP is indispensable for mammalian infection. The hypothesis of this proposal is that small molecule inhibitors targeting c-di-GMP and c-di- AMP cyclases, Rrp1 and CdaA, would eliminate B. burgdorferi in ticks and mammalian reservoirs, which can be exploited to reduce Lyme disease incidence. The co-PI of this proposal, Dr. Herman Sintim, a Drug Discovery Professor of Chemistry, pioneered developing inhibitors for bacterial diguanylate cyclase and diadenylate cyclase, and his group has already reported several potent inhibitors against these cyclases of other bacteria. Accordingly, we propose to develop a strategy to target CdaA of B. burgdorferi to eliminate in spirochetes rodent reservoirs (Aim 1), and a strategy to target Rrp1 of B. burgdorferi to eliminate spirochetes in ticks (Aim 2). We will also test a combination of inhibitors targeting both pathways to eliminate B. burgdorferi in its enzootic cycle. The underlying mechanisms of how c-di-AMP and c-di-GMP are employed by B. burgdorferi to survive in mammals and ticks will also be investigate. This proposal address one of the specific Focus Areas listed in this RFA: reservoir-targeted approaches to interrupt the natural history of infection. Upon accomplishing the proposed work, further field experiments will be conducted using animal baits containing a combination of both inhibitors to evaluate their effects on reducing spirochete burden in ticks and animals in nature. Such host-targeted strategy will have significant impact on combating Lyme disease.