Targeted expression of factor VIII in liver sinusoidal endothelial cells for gene therapy for hemophilia A

肝窦内皮细胞中因子 VIII 的靶向表达用于血友病 A 的基因治疗

• 批准号: 10242722
• 负责人: DENISE E SABATINO
• 金额: $ 64.11万
• 依托单位: CHILDREN'S HOSP OF PHILADELPHIA
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-09-01 至 2024-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10242722
• 关键词: Address; Bar Codes; Biochemical; Biological; Biological Assay; Biological Response Modifier Therapy; Biology; Blood Circulation; Blood Coagulation Disorders; Blood Coagulation Factor; Capsid; Cells; Cellular Stress; Cellular Stress Response; Characteristics; Clinical; Clinical Data; Clinical Research; Clinical Trials; Data; Disease; Dose; Elements; Endothelial Cells; Engineering; F8 gene; Factor VIII; Gene Delivery; Gene Transfer; Genes; Goals; Hemophilia A; Hemorrhage; Hepatocyte; Immune response; Libraries; Link; Liver; Pathway interactions; Patient-Focused Outcomes; Patients; Plasma; Post-Translational Protein Processing; Proteins; Risk; Role; Safety; Serotyping; Site; System; Therapeutic; Time; Tissues; Transgenes; Variant; Viral; adeno-associated viral vector; base; biological adaptation to stress; blood product; cell type; disease phenotype; efficacy study; enzyme replacement therapy; follow-up; gene therapy; genotoxicity; improved; novel; preclinical study; prevent; promoter; response; risk minimization; therapeutic development; vector

ABSTRACT Hemophilia A (HA) is an X-linked bleeding disorder caused by a deficiency in coagulation factor VIII (FVIII). Therapies aimed at even modest increases in clotting factor levels are associated with substantial improvement of the severe disease phenotype. The current treatment for HA is protein replacement therapy, however, the therapeutic landscape is rapidly changing for this disorder. Gene therapy approaches for adeno-associated viral (AAV) vector delivery of FVIII are in clinical trials. Interestingly, these clinical studies target expression of FVIII to hepatocytes using hepatocyte specific promoter elements which is based on the assumption that FVIII is synthesized in hepatocytes. However, the primary site of FVIII synthesis was identified as the liver sinusoidal endothelial cell (LSEC). While preclinical and clinical studies have demonstrated that hepatocytes can synthesize functional FVIII, it has been challenging to produce high levels of FVIII after hepatocyte targeted expression. In the ongoing AAV clinical studies for HA, there have been several unexpected observations. The first successful AAV-hFVIII trial with the most patients (>13 subjects) and the longest follow-up (>3 years, ongoing) observed a significant decrease in FVIII expression. All of the available clinical data from three trials to date shows a lack of a vector dose response with significant variability among subjects. Importantly, these findings were not observed in the AAV-FIX clinical studies suggesting that there is additional complexity to the delivery and expression of FVIII. These unanticipated findings in the clinical studies may be related to the site of FVIII synthesis. Thus, while significant hurdles have been overcome in gene therapy for HA, unexplored opportunities for improved hemophilia patient outcomes remain. The goal of this proposal is to target FVIII expression to LSECs using AAV vectors to study if there are biological differences in expressing FVIII in LSECs and hepatocytes and to study the efficacy of AAV-FVIII delivery to LSECs. FVIII expression will be targeted to LSECs using novel promoter elements (Specific Aim 1), novel AAV capsids that more specifically target AAV to these cells will be identified (Specific Aim 2) and the biological differences between hepatocyte and LSEC derived FVIII expression after AAV delivery will be investigated (Specific Aim 3). Together, these studies will provide the basis for understanding the biology and efficacy of LSEC targeted AAV-FVIII expression to support the development of this therapeutic approach for hemophilia A.

抽象的 血友病A（HA）是由凝结因子VIII（FVIII）缺乏引起的X连锁出血障碍。 旨在适度增加凝血因子水平的疗法与大幅改善有关 严重疾病表型。 HA的当前治疗方法是蛋白质替代疗法，但是 对于这种疾病，治疗景观正在迅速变化。腺相关病毒的基因治疗方法 （AAV）FVIII的矢量传递正在临床试验中。有趣的是，这些临床研究针对FVIII的表达 使用肝细胞特异性启动子元素到基于FVIII为 在肝细胞中合成。但是，FVIII合成的主要位点被确定为肝弦曲线 内皮细胞（LSEC）。尽管临床前和临床研究表明肝细胞可以合成 功能性FVIII，在肝细胞靶向表达后产生高水平的FVIII一直具有挑战性。在 正在进行的HA的AAV临床研究，有几个意外的观察结果。第一个成功 AAV-HFVIII试验大多数患者（> 13名受试者）和最长的随访（> 3年，正在进行） FVIII表达显着降低。到目前为止三个试验中的所有可用临床数据都表明缺乏 矢量剂量反应的受试者之间有显着差异的响应。重要的是，这些发现不是 在AAV固定临床研究中观察到，这表明输送和 FVIII的表达。临床研究中的这些意外发现可能与FVIII部位有关 合成。因此，尽管HA基因疗法已经克服了重大障碍，但未开发的机会 为了改善血友病患者的预后。该提议的目的是将FVIII表达靶向 LSEC使用AAV矢量研究是否存在LSEC中FVIII的生物学差异 肝细胞并研究AAV-FVIII向LSEC递送的功效。 FVIII表达将针对LSEC 使用新颖的启动子元素（特定目标1），新颖的AAV capsids更具体地针对AAV 将鉴定细胞（特定目标2）以及肝细胞和LSEC衍生的FVIII之间的生物学差异 将研究AAV输送后的表达（特定目标3）。这些研究将共同​​提供基础 为了理解LSEC的生物学和功效，以AAV-FVIII的表达来支持发展 关于血友病的治疗方法。