A Bioengineering Approach to Gene Therapy for Peripheral Arterial Disease

外周动脉疾病基因治疗的生物工程方法

• 批准号: 9249087
• 负责人: BRIAN H ANNEX
• 金额: $ 69.13万
• 依托单位: UNIVERSITY OF VIRGINIA
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-04-01 至 2019-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9249087
• 关键词: 20 year old; Adenovirus Vector; Adenoviruses; Age; Angiotensin-Converting Enzyme Inhibitors; Animal Experiments; Animal Model; Aorta; Arteries; Asialoglycoproteins; Aspirin; Atherosclerosis; Back; Biodistribution; Biomedical Engineering; Biopsy Specimen; Blood Vessels; Blood flow; Body Weight; Clinical; Clinical Investigator; Clinical Trials; Collaborations; Data; Dependovirus; Distal; Dose; Failure; Family suidae; Foundations; Galactose; Gene Delivery; Gene Expression; Gene Transfer; Glycoproteins; Growth; Health; Heart; Hindlimb; Human; Hypoxia; Immune response; Impairment; Inbred BALB C Mice; Individual; Intramuscular; Intramuscular Injections; Intravenous; Investigation; Ischemia; Isolated limb perfusion; Laboratories; Leg; Limb structure; Link; Liver; Lower Extremity; Lung; Mediating; Medical; Membrane Glycoproteins; Modeling; Mus; Muscle; Myocardial Ischemia; Necrosis; Obstruction; Pathway interactions; Patients; Perfusion; Peripheral arterial disease; Plasmid Cloning Vector; Plasmids; Prevalence; Recovery; Reporter Genes; Response Elements; Scientist; Serotyping; Skeletal Muscle; Symptoms; System; T cell response; Therapeutic; Tissues; Toxic effect; Transduction Gene; Translating; Translations; Universities; Virginia; Walking; Weight; bench to bedside; clinical application; clinical practice; clinically relevant; clopidogrel; disorder control; gene delivery system; gene therapy; improved; man; particle; pre-clinical; programs; public health relevance; receptor; skeletal; success; sugar; therapeutic angiogenesis; therapeutic gene; vector; vector genome

DESCRIPTION (provided by applicant): Peripheral arterial disease (PAD) is caused by atherosclerosis that results in obstructions in the arteries to the lower extremities. The prevalence of PAD is nearly equal to that of ischemic heart disease. In a sizeable fraction of patients with severe forms of PAD, there is a complete occlusion in one or more major arteries, and thus blood flow to the distal lower limb becomes dependent on the extent, number and function of collateral blood vessels. There are no medical therapies available for patients with PAD that favorably modulate collateral number or function and perfusion to the lower extremity. The concept of stimulating new blood vessel growth ('therapeutic angiogenesis') to improve limb perfusion in patients with PAD is now a 20 year old investigational strategy. The vast majority of human trials use gene transfer by intra-muscular (IM) delivery with plasmid or adenoviral vectors. After dozens of trials and thousands of subjects, success in this field has been meager. Human studies have failed to convincingly show therapeutic gene expression following IM injection, and this may well account for the failure of clinical trials in therapeutic angiogenesis conducted to date. Adeno-associated virus can be transformational in providing the link from mouse to man for therapeutic angiogenesis in PAD. Following systemic (intravenous) delivery, several AAV serotypes efficiently transduce skeletal muscle. While PAD could potentially limit the access of AAV9 to ischemic muscle, we found enhanced gene expression in ischemic compared to non-ischemic muscle following systemic delivery and we have examined potential mechanisms. Building on this foundation, the central hypothesis of this revised proposal is that bioengineered AAV9 vectors are uniquely suited to accomplish therapeutic angiogenesis in PAD via local delivery using isolated limb perfusion. The three complementary but independent Aims are to: Specific Aim 1: Optimize AAV9-mediated gene delivery to ischemic muscle using systemic vs. isolated limb perfusion in small and large animal models of PAD as a link to clinical application. We will use reporter genes and quantitative biodistribution studies of gene expression to compare systemic versus local delivery in ischemic muscle relative to off-target tissues in mice and pigs. Further, we will incorporate hypoxia-response elements and examine immune responses in both murine and porcine hind-limb ischemia models of PAD. Specific Aim 2: Establish that ischemia-induced desialylation, which results in a difference in the ratio of sialylated to desialylated glycoproteins, is a mechanism for enhanced gene expression in ischemic muscle in pre-clinical and clinical PAD subjects vs. non-PAD controls. Specific Aim 3: Determine the efficacy of AAV-mediated gene therapy delivered via isolated limb perfusion in both mouse and porcine hind-limb ischemia models of PAD. In this aim, we will compare ADAM12 vs. EcSOD gene therapy in the mouse hind-limb ischemia model of PAD, then confirm the efficacy of the most effective gene therapy in the porcine hind-limb model of PAD in collaboration with Dr. Albert Sinusas (Yale).

描述（由申请人提供）：外周动脉疾病（PAD）是由导致下肢动脉阻塞的动脉粥样硬化引起的。 PAD 的患病率几乎与缺血性心脏病的患病率相同。在相当一部分患有严重 PAD 的患者中，一根或多根主动脉完全闭塞，因此流向下肢远端的血流取决于侧支血管的范围、数量和功能。对于 PAD 患者，尚无可有效调节下肢侧支循环数量或功能以及灌注的药物疗法。刺激新血管生长（“治疗性血管生成”）以改善 PAD 患者肢体灌注的概念现已成为一项已有 20 年历史的研究策略。绝大多数人体试验都使用质粒或腺病毒载体通过肌内 (IM) 递送进行基因转移。经过数十次试验和数千个受试者之后，该领域的成功甚微。人体研究未能令人信服地显示肌内注射后治疗基因的表达，这很可能解释治疗性血管生成临床试验的失败 迄今为止进行的。 腺相关病毒可以为 PAD 治疗性血管生成提供从小鼠到人类的转化联系。全身（静脉）递送后，几种 AAV 血清型可有效转导骨骼肌。虽然 PAD 可能会限制 AAV9 进入缺血肌肉，但我们发现全身递送后，与非缺血肌肉相比，缺血肌肉中的基因表达增强，并且我们已经检查了潜在的机制。在此基础上，本修订提案的中心假设是生物工程 AAV9 载体特别适合通过使用隔离肢体灌注进行局部递送来完成 PAD 中的治疗性血管生成。这三个互补但独立的目标是： 具体目标 1：在 PAD 的小型和大型动物模型中使用全身与孤立肢体灌注来优化 AAV9 介导的基因递送至缺血肌肉，作为临床应用的链接。我们将使用报告基因和基因表达的定量生物分布研究来比较小鼠和猪的缺血肌肉中相对于脱靶组织的全身递送和局部递送。此外，我们将纳入缺氧反应元件并检查 PAD 小鼠和猪后肢缺血模型中的免疫反应。 具体目标 2：确定缺血诱导的去唾液酸化（导致唾液酸化与去唾液酸化糖蛋白的比例存在差异）是临床前和临床 PAD 受试者与非 PAD 对照者缺血肌肉中基因表达增强的机制。 具体目标 3：确定通过离体肢体灌注 AAV 介导的基因治疗在小鼠和猪 PAD 后肢缺血模型中的疗效。为此，我们将在小鼠 PAD 后肢缺血模型中比较 ADAM12 与 EcSOD 基因疗法，然后与 Albert Sinusas 博士合作，确认最有效的基因疗法在猪 PAD 后肢模型中的疗效（耶鲁大学）。

项目成果

BAG3 (Bcl-2-Associated Athanogene-3) Coding Variant in Mice Determines Susceptibility to Ischemic Limb Muscle Myopathy by Directing Autophagy.

小鼠中的 BAG3（Bcl-2 相关 Athanogene-3）编码变体通过指导自噬确定对缺血性肢体肌肉肌病的易感性。

• 发表时间: 2017-07-18
• 影响因子: 37.8
• 作者: McClung, Joseph M;McCord, Timothy J;Ryan, Terence E;Schmidt, Cameron A;Green, Tom D;Southerland, Kevin W;Reinardy, Jessica L;Mueller, Sarah B;Venkatraman, Talaignair N;Lascola, Christopher D;Keum, Sehoon;Marchuk, Douglas A;Spangenburg, Espen E
• 通讯作者: Spangenburg, Espen E

Loss of interleukin-21 receptor activation in hypoxic endothelial cells impairs perfusion recovery after hindlimb ischemia.

缺氧内皮细胞中白细胞介素 21 受体激活的丧失会损害后肢缺血后的灌注恢复。

• 发表时间: 2015-05
• 作者: Wang, Tao;Cunningham, Alexis;Dokun, Ayotunde O;Hazarika, Surovi;Houston, Kevin;Chen, Lingdan;Lye, R John;Spolski, Rosanne;Leonard, Warren J;Annex, Brian H
• 通讯作者: Annex, Brian H

Extensive skeletal muscle cell mitochondriopathy distinguishes critical limb ischemia patients from claudicants.

广泛的骨骼肌细胞线粒体病将严重肢体缺血患者与跛行患者区分开来。

• DOI: 10.1172/jci.insight.123235
• 发表时间: 2018-11-02
• 期刊: JCI insight
• 影响因子: 8
• 作者: Terence E. Ryan;D. Yamaguchi;Cameron A. Schmidt;Tonya N Zeczycki;S. Shaikh;Patricia M. Brophy;T. D. Green;Michael D. Tarpey;R. Karnekar;Emma J. Goldberg;G. Sparagna;Maria.J. Torres;B. Annex;P. Neufer;E. Spangenburg;J. McClung
• 通讯作者: J. McClung

Endothelial interleukin-21 receptor up-regulation in peripheral artery disease.

外周动脉疾病中内皮白细胞介素 21 受体上调。

• 发表时间: 2016-04
• 作者: Wang, Tao;Cunningham, Alexis;Houston, Kevin;Sharma, Aditya M;Chen, Lingdan;Dokun, Ayotunde O;Lye, R John;Spolski, Rosanne;Leonard, Warren J;Annex, Brian H
• 通讯作者: Annex, Brian H

Neuraminidase-mediated desialylation augments AAV9-mediated gene expression in skeletal muscle.

神经氨酸酶介导的去唾液酸化增强了骨骼肌中 AAV9 介导的基因表达。

• 发表时间: 2018-09
• 作者: Zhu, Hongling;Wang, Tao;John Lye, Robert;French, Brent A;Annex, Brian H
• 通讯作者: Annex, Brian H