Evaluation of vectors based on group B adenoviruses

描述（由申请人提供）： 十多年来，基于腺病毒（Ad）血清型5的载体已用于基因治疗，但成功有限。基于Ad 5的载体的主要缺点似乎是在大多数人中预先存在的免疫力、重要基因治疗靶细胞的低转导（由于初级Ad 5受体CAR的低表达）和在血管内应用时诱导的先天毒性（部分由于网状内皮系统的细胞，特别是枯否细胞的摄取）。最近的数据表明，含有来自Ad组B血清型11和35的纤维的载体识别不同于CAR的细胞受体，并有效地识别相对难治Ad 5感染的人细胞类型，包括恶性肿瘤细胞、骨髓来源的造血干细胞、间充质细胞和树突状细胞。此外，我们发现，静脉注射的载体含有B组Ad 35或Ad 1纤维只能无效地抑制枯否细胞，因此引起显着降低的先天性毒性相比，Ad 5载体。然而，关于B组腺病毒载体在体内外感染的机制和作用知之甚少。在考虑这些新载体的临床应用之前，必须填补这一知识空白。因此，本提案的中心目标是研究B组Ad载体感染在细胞和生物体水平上的效率、特异性和安全性。我们将首先描绘参与B组Ad感染的细胞受体，并构建一组包含来自代表性B组Ad血清型的纤维的报告载体，所述纤维在细胞受体的使用方面不同。为了研究Ad感染对细胞生物学（包括信号传导和基因表达的变化）的影响，我们将关注表达B组Ad受体并且是离体基因治疗的相关靶点的人细胞培养物。然后，在体内应用后，我们将研究B组Ad载体的生物分布并监测血液参数、组织病理学、细胞因子产生/释放和抗载体免疫应答。这些研究将在表达B组Ad受体的转基因小鼠中进行。在此基础上，我们将选择代表性载体和参数，并在狒狒中重复生物分布和安全性研究。

DESCRIPTION (provided by applicant): For more than a decade, adenovirus (Ad) serotype 5-based vectors have been used in gene therapy, with limited success. The main disadvantages of Ad5-based vectors appear to be preexisting immunity in the majority of humans, low transduction of important gene therapy target cells (due to low expression of the primary Ad5 receptor, CAR), and innate toxicity induced upon intravascular application (in part due to uptake by cells of the reticuloendothelial system, particularly Kupffer cells). Recent data indicate that vectors containing fibers from Ad group B serotypes 11 and 35 recognize cellular receptors different from CAR and efficiently transduce human cell types that are relatively refractory to Ad5 infection, including malignant tumor cells, bone marrow derived hematopoietic stem cells, mesenchymal cells, and dendritic cells. Furthermore, we found that intravenously injected vectors containing B-group Ad35 or Ad 1 fibers only inefficiently transduce Kupffer cells and therefore elicit significantly reduced innate toxicity as compared to Ad5 vectors. However, little is known about mechanisms and effects of B-group Ad vector infection in vitro and in vivo. Before a clinical application of these new vectors can be considered, this knowledge gap has to be filled. Therefore, the central goal of this proposal is to study the efficiency, specificity, and safety of B-group Ad vector infection on the cellular and organism levels. We will first delineate the cellular receptors involved in B-group Ad infection and construct a set of reporter vectors containing fibers from representative B-group Ad serotypes that differ in the usage of cellular receptors. To study the effects of Ad infection on cell biology (including signaling and changes in gene expression), we will focus on human cell cultures that express B- group Ad receptors and are relevant targets for ex vivo gene therapy. Then, upon in vivo application, we will study the biodistribution of B-group Ad vectors and monitor blood parameters, histopathology, cytokine production/release, and anti-vector immune responses. These studies will be performed in transgenic mice that express B-group Ad receptors. Based on this, we will select representative vectors and parameters and repeat biodistribution and safety studies in baboons.