Development of therapeutic antibodies for vaccinia virus

说明(申请人提供)：为了应对生物恐怖分子可能释放的天花，对高危人群接种牛痘病毒(VV)疫苗是唯一可用的预防性干预措施。然而，疫苗也有已知的并发症，特别是在免疫功能低下的宿主、孕妇和婴儿中。在过去的疫苗接种工作中，这种并发症在美国是用从VV免疫者那里获得的牛痘人类免疫球蛋白(VIG)来治疗的。尽管这种材料有疗效，但人们对免疫球蛋白的哪些成分有效知之甚少，而且疗效可能会因批次而异。此外，由于常规疫苗接种已停止多年，可供使用的VIG供应有限，人们对其安全性感到担忧。我们的长期目标是开发一种针对VV蛋白的明确和高亲和力的抗体鸡尾酒，在需要大规模VV疫苗接种的情况下取代传统VIG的使用。我们的策略是开发针对各种VV包膜蛋白的单抗，这些蛋白在小鼠模型中可引起中和和/或保护性反应。这种抗体的鸡尾酒，我们将称为VIG-R(替代)将提供一个统一和安全的VV免疫治疗试剂的来源。 在本申请中，我们提出了四个特定目标的实验。1)在杆状病毒表达系统中表达和鉴定VV包膜蛋白LIR、A33R和B5R，并制备针对这些蛋白的鼠和兔抗体；2)利用噬菌体展示技术制备抗LIR、A33R和BSR的人源性单抗；3)检测针对VV蛋白的免疫试剂保护小鼠免受VV攻击的能力。这笔赠款的目标的实现应该提供关于每种VV蛋白和抗体的新信息，以及可以组成VIG-R的试剂的来源。如果我们的方法看起来很有希望，我们将扩大这项研究，以包括其他VV蛋白。在这项拨款申请中开发的试剂也将对有关这些蛋白质在VV生命周期中的作用的基础研究很有价值，并可在未来的研究中被视为候选疫苗。

DESCRIPTION (provided by applicant): In response to the possible release of smallpox (variola) by bioterrorists, vaccinia virus (VV) vaccination of an at-risk population is the only available prophylactic intervention. However, the vaccine has known complications, especially in immunocompromised hosts, pregnant women and infants. In past vaccination efforts, such complications were treated in the U.S. with vaccinia human immune globulin (VIG) obtained from VV immunized people. Although this material had efficacy, little is known about which components of the immune globulin were effective and there could be batch-to-batch variation in efficacy. Moreover, since routine vaccination has been discontinued for many years, there is only a limited supply of VIG available, and there are concerns about its safety. Our long-term goal is to develop a cocktail of defined and high affinity antibodies to VV proteins that will replace the use of traditional VIG in the event that mass VV vaccination is needed. Our strategy is to develop monoclonal antibodies to various VV envelope proteins that are known to provoke a neutralizing and/or protective response in a mouse model. A cocktail of such antibodies, which we will call VIG-R (replacement) will provide a uniform and secure source of a VV immune therapeutic reagent. In this application, we propose experiments in four specific aims. These are: 1) to express and characterize VV envelope proteins LIR, A33R and B5R in a baculovirus expression system and to prepare mouse and rabbit antibodies to those proteins; 2) to develop and characterize human monoclonal antibodies to LIR, A33R and BSR produced using phage display; and 3) to test the ability of immune reagents against VV proteins to protect mice from challenge with VV. Fulfillment of the aims of this grant should provide new information about each of the VV proteins and antibodies, as well as a source of reagents that can comprise a VIG-R. If our approach appears promising, we will expand this study to include other VV proteins. The reagents developed in this grant application will also be valuable for basic studies concerning the role of these proteins in the VV life cycle and could themselves be considered as vaccine candidates in future investigations.