Discovery of antivirals against vaccinia and smallpox

描述（由申请人提供）：人们越来越担心恐怖组织可能拥有或能够获得天花病毒（天花感染的病原体）。为了应对这一具体威胁，美国必须拥有可供支配的牛痘病毒疫苗和抗天花病毒化合物。在疫苗禁忌的情况下需要抗病毒化合物，如免疫抑制。此外，需要抗病毒化合物来减弱已知与疫苗施用相关的不良并发症。理想的抗病毒药物还应该能够在疫苗接种延迟的情况下直接预防天花感染。为此，针对牛痘病毒DNA聚合酶（V AC-POL）和持续合成因子（V AC-PF）的抗病毒药物具有很大的潜力。通常，PF充当沿着DNA沿着滑动并结合到其各自的POL的夹子。通过这种POL/PF相互作用，DNA合成将是进行性的，产生数千个核苷酸长的新DNA链。具有突变的PF的病毒在复制中是有缺陷的，因为单独的POL只能合成短的DNA产物（少至三个核苷酸）。PF的一个非常有利的特征是它们与它们自己的POL一起起作用，因此靶向病毒PF的抗病毒药物应该是非常特异的，并且不干扰细胞复制。VAC-POL和VAC-PF相互作用最近已经被表征，并且这两种蛋白质可以被纯化并组合以在进行性DNA合成中起作用。该项目的目标是发现特异性阻断V AC-POL和V AC-PF进行性DNA合成的抗病毒药物。新开发的机械快速平板试验将用于筛选数千种潜在的抑制化合物。高通量筛选之后将进行评价抑制性化合物的程序，并最终测试其阻断牛痘病毒感染的能力。一种阻止牛痘进行性DNA合成的药物将有助于减少牛痘疫苗的并发症。相同的药物也可以直接阻断天花感染，因为牛痘和天花的POL和PF分别具有97%的直接同源性。这种方法可能有助于提供“及时”的试剂需求，以应对天花生物恐怖主义威胁。

DESCRIPTION (provided by applicant): There is a mounting concern that terrorist groups may possess or can obtain their own stocks of variola virus, the etiological agent of smallpox infection. To respond to this specific threat, the United States must have at its disposal supplies of both vaccinia virus vaccine and antiviral compounds directed against smallpox infection. The antiviral compounds are needed in situations in which the vaccine is contraindicated, such as immunosuppression. In addition, the antiviral compounds are needed to blunt adverse complications that are known to be associated with vaccine administration. An ideal antiviral should also be able to directly prevent smallpox infection in situations where vaccine delivery is delayed. Toward this end, antivirals directed against the vaccinia virus DNA polymerase (V AC-POL) and processivity factor (V AC-PF) have great potential. In general, PFs act as clamps that slide along the DNA and bind to their respective POLs. Through this POL/PF interaction, DNA synthesis will be processive, generating new DNA strands that are thousands of nucleotides long. Viruses that have mutated PFs are defective in replication since POL alone can synthesize only short DNA products (as few as three nucleotides). A strongly favorable feature of PFs is that they function with their own POLs, so that an antiviral that targets a viral PF should be very specific and not interfere with cellular replication. V AC-POL and V AC-PF interaction has been recently characterized and both proteins can be purified and combined to function in processive DNA synthesis. The goal of this project is to discover antivirals that will specifically block processive DNA synthesis by V AC-POL and V AC-PF. A newly developed mechanistic rapid plate assay will be used to screen thousands of potentially inhibitory compounds. This high throughput screening will be followed by procedures to evaluate the inhibitory compounds and to ultimately test their ability to block vaccinia virus infection. A drug that prevents vaccinia processive DNA synthesis will be useful in curtailing vaccinia vaccine complications. The same drug may also directly block variola infection since the POLs and PFs, respectively, of vaccinia and variola have 97% direct homology. The approach may help deliver the 'just-in-time' need for reagents to combat a smallpox bioterrorism threat.