Engineered “muco-trapping” antibodies for inhaled therapy of parainfluenza and human metapneumovirus infections

人副流感病毒（PIV）和偏肺病毒（MPV）是下呼吸道感染的常见原因。 婴儿和幼儿的呼吸道感染，以及免疫受损儿童呼吸道疾病的主要原因 成年人和老年人。不幸的是，目前还没有针对这两种病毒的治疗方法或疫苗， 提供医疗服务。有趣的是，PIV和MPV有许多共同的病理和临床表现， 呼吸道合胞病毒（RSV）。事实上，PIV和MPV都不知道会导致血液中的病毒血症， 感染的患者，这表明这两种感染都严格局限于呼吸道，类似于RSV， 病毒只从受感染细胞的顶端表面传播。这种独特的病理生理学意味着后代 病毒在扩散到邻近细胞之前必须穿过气道粘液（AM）。这反过来又促使我们 开发一种病原体特异性的抗病毒药物，可以在物理上限制呼吸道内感染的传播。 我们最近在粘液中发现了一种新的抗体效应子功能--在粘液中捕获单个病原体 基于IgG-Fc和粘蛋白之间的精心调节的亲和力-并开发了一个平台，用于增强mAb 在粘膜表面起作用。我们假设，递送到气道的“粘膜捕获”mAb可以直接 通过在AM中拦截和诱捕脱落的子代病毒来干预病毒的生命周期， 通过自然的粘膜纤毛清除将病毒从气道中捕获，并能够在体内进行有效的治疗。在 为了支持这一策略，我们设计了在AM中有效捕获RSV的mAb，并表明雾化吸入的 从感染后第3天开始向RSV感染的新生羔羊有效递送“粘膜捕获”mAb 在3天内将肺组织中的感染性病毒载量降低到不可检测的水平， 与溶剂对照相比，支气管组织中的细胞数减少了11倍。这些结果促使我们去探索 类似的方法是否可以有效治疗PIV和MPV感染。为了支持这项申请， 我们已经设计了对两种病毒的不同毒株具有皮摩尔亲和力的抗体，并证明， 我们可以有效地将病毒捕获在人AM中，并限制感染在高分化人类中的体外传播， 气道上皮（WD HAE）生长在气液界面。 在这个建议中，我们将继续我们的工作与亲和力成熟使用哺乳动物和酵母展示 以产生广泛结合和中和PIV和MPV的不同毒株的高亲和力mAb（目的1）。我们将 验证这些mAb是否可以捕获新鲜未稀释的人AM中的PIV和MPV，以及它们是否可以抑制 预先建立的PIV和MPV感染在WD-HAE培养物中的传播（目的2）。然后我们将推进 用于仓鼠鼻感染模型评价的两种病毒的先导mAb（目的3）。通过增强 mAb在粘液分泌物中的功能，我们希望我们将有助于为改善分子靶向 治疗和预防所有主要粘膜表面的广谱病原体， 这是一个解决目前呼吸道感染药物干预差距的有力选择。

Human parainfluenza virus (PIV) and metapneumovirus (MPV) are common causes of lower respiratory tract infections in infants and young children, and major causes of respiratory illness in immune compromised adults and the elderly. Unfortunately, there are no therapy or vaccine for either viruses, and only supportive medical care is available. Interestingly, PIV and MPV share many pathological and clinical manifestations as Respiratory Syncytial Virus (RSV). Indeed, neither PIV nor MPV are known to cause viremia in the blood of infected patients, indicating both infections are strictly localized in the airways similar to RSV that shed progeny viruses exclusively from the apical surface of infected cells. Such unique pathophysiology implies progeny viruses must traverse airway mucus (AM) before spreading to neighboring cells. This in turn motivated us to develop a pathogen-specific antiviral that could physically limit the spread of the infections within the airways. We recently discovered a novel Ab effector function in mucus – trapping individual pathogens in mucus based on carefully-tuned affinity between IgG-Fc and mucins – and developed a platform for enhancing mAb function at mucosal surfaces. We hypothesize that “muco-trapping” mAb delivered to the airways can directly intervene with the viral life cycle by intercepting and trapping shed progeny viruses in AM, rapidly eliminate trapped viruses from the airways by natural mucociliary clearance, and enable effective therapy in vivo. In support of this strategy, we engineered mAbs that potently trap RSV in AM, and showed that nebulized delivery of “muco-trapping” mAb to RSV-infected neonatal lambs beginning on Day 3 post infection effectively reduced the infectious viral load in lung tissues to non-detectible levels within 3 days, with viral RNA in bronchial tissues reduced by 11-fold compared to vehicle control. These results motivated us to explore whether a similar approach may be effective in treating PIV and MPV infections. In support of this application, we have engineered antibodies with picomolar affinity to diverse strains of both viruses, and demonstrated that we could effectively trap viruses in human AM and limit spread of infection in vitro in well-differentiated human airway epithelium (WD-HAE) grown at the air-liquid interface. In this proposal, we will continue our work with affinity maturation using mammalian and yeast display to produce high affinity mAb that broadly bind and neutralize diverse strains of PIV and MPV (Aim 1). We will validate whether these mAbs can trap PIV and MPV in fresh undiluted human AM, and whether they can inhibit the spread of pre-established PIV and MPV infections in WD-HAE cultures (Aim 2). We will then advance the lead mAb for both viruses for evaluation in a hamster nasal infection model (Aim 3). By enabling enhanced mAb function in mucus secretions, we expect we will help pave the way for improved, molecularly-targeted therapies and prophylaxis against a broad spectrum of pathogens across all major mucosal surfaces, providing a powerful option addressing the current gap in pharmacological interventions for respiratory infections.