Injectable Hydrogels to Deliver Gene Therapy for Myocardial Infarct

心肌梗死(MI)后，局部组织重构导致慢性心功能恶化，即 这是美国的一个主要死因。几项临床前研究表明，局部注射生长因子或 编码生长因子的基因可以显著改善心脏功能。不幸的是，有效的交付 心脏跳动的治疗仍然是一个巨大的挑战，阻碍了新药的临床翻译 治疗学。理想的心肌梗死给药系统应该是导管注射的，可以防止从导管中挤出 可收缩心肌，并将提供有效的治疗剂量的持续输送。不幸的是， 大多数导管可注射的生物材料都是从收缩的心脏组织中迅速挤出的弱水凝胶。 为了克服这一临床挑战，我们提出了通过动态交联的可注射凝胶的设计 共价化学键(DCC)，强度很强，但可逆。因此，这些DCC水凝胶结合了临床上 可注射性和在打浆中保持所需的机械完整性所需的特性 心。具体地说，我们的凝胶是通过化学修饰的透明质酸之间的DCC糖苷键形成的 酸和一种重组的类弹性蛋白。得到的凝胶是可酶生物降解的，完全是化学的。 在FDA的研究中定义了未来的潜力。在目标1中，20个凝胶家族具有独特的粘弹性机械性能 性能将被合成和表征，以便于导管注射和在收缩中保持 心。我们将通过改变透明质酸的相对分子质量和产率来调节凝胶的粘度 通过改变DCC交联物的浓度对凝胶施加应力并在体外和体内执行 可注射性的量化。同时，在目标2中，我们评估了一种假设，即 再生的有效载荷可以通过组合混合具有不同切割的药物系链来实现 运动学。具体地说，我们的有效载荷是编码基质细胞衍生因子-1α(sdf-1α)的微环基因，它是 已知可诱导血管生成并改善心肌梗死后的心功能。该有效载荷被系在可注射的 通过与多肽核酸(PNA)-多肽的DNA杂交进行凝胶。在目标3中，目标1的凝胶配方与 最佳的体内保留性能和来自Aim 2的具有持续基因释放的药物系链设计将是 合并成可注射的心肌梗死疗法。将在临床前大鼠心肌梗死模型中评估功能表现 使用携带sdf-1α和萤火虫荧光素酶报告基因的微环基因。诱发心肌梗死后 通过结扎左前降支(LAD)动脉，动物将随机分为两组 或治疗组。治疗动物将接受60-μL心肌内注射的生理盐水、水凝胶 只有生理盐水中的非系留基因、凝胶中的非系留基因或凝胶中的系留基因。生物发光成像 (第0、1、4、7、21、42、60和90天)将用于监测基因表达。心肌梗塞后的功能恢复将是 使用超声心动图(第7天、第21天)和血流动力学测量(第90天)进行评估。最后，心 将分析外植体的坏死、炎症、血管生成和组织再生的证据(第90天)。

Following myocardial infarction (MI), local tissue remodeling leads to chronically worsening heart function that is a major cause of death in the US. Several preclinical studies have shown that local delivery of growth factors or growth factor-encoding genes can significantly improve cardiac function. Unfortunately, effective delivery of therapeutics to the beating heart remains a formidable challenge, impeding clinical translation of novel drug therapeutics. The ideal MI drug-delivery system would be catheter injectable, would prevent extrusion out of the contractile myocardium, and would provide sustained delivery of an effective therapeutic dosage. Unfortunately, most catheter-injectable biomaterials are weak hydrogels that are rapidly extruded out of contractile heart tissue. To overcome this clinical challenge, we propose the design of injectable gels that are crosslinked by dynamic covalent chemistry (DCC) bonds that are strong yet reversible. Thus, these DCC hydrogels combine the clinically desired properties of being injectable and having the mechanical integrity required for retention in the beating heart. Specifically, our gels are formed through DCC hydrazone bonds between a chemically modified hyaluronic acid and a recombinant, elastin-like protein. The resulting gel is enzymatically biodegradable and fully chemically defined for future potential in FDA studies. In Aim 1, a family of 20 gels with distinct viscoelastic mechanical properties will be synthesized and characterized for ease of catheter injection and retention in the contracting heart. We will modulate the viscosity of the gels by altering the molecular weight of hyaluronic acid and the yield stress of the gel by varying the concentration of a DCC crosslink competitor and perform in vitro and in vivo quantifications of injectability. In parallel in Aim 2, we evaluate the hypothesis that sustained release of a regenerative payload can be achieved through combinatorial mixing of drug tethers with distinct cleavage kinetics. Specifically, our payload is minicircle genes encoding stromal cell-derived factor-1α (SDF-1α), which is known to induce angiogenesis and improved heart function following MI. This payload is tethered to the injectable gel via DNA hybridization with peptide nucleic acid (PNA)-peptides. In Aim 3, the gel formulation from Aim 1 with optimal in vivo retention properties and the drug tether design from Aim 2 with sustained gene release will be combined into an injectable MI therapy. Functional performance will be evaluated in a preclinical rat MI model using minicircle genes carrying both SDF-1α and a firefly luciferase reporter gene. Following induction of MI through ligation of the left anterior descending (LAD) artery, animals will be randomly assigned into either sham or treatment groups. Treatment animals will receive a 60-μL intramyocardial injection of saline only, hydrogel only, untethered genes in saline, untethered genes in gel, or tethered genes in gel. Bioluminescence imaging (days 0, 1, 4, 7, 21, 42, 60, and 90) will be used to monitor gene expression. Functional recovery after MI will be assessed using echocardiography (days 7, 21) and hemodynamic measurements (day 90). Finally, heart explants will be analyzed for evidence of necrosis, inflammation, angiogenesis, and tissue regeneration (day 90).