Pre-clinical Validation of A Novel Protein Drug Candidate for ASH and NASH Treatment

摘要 酒精性脂肪性肝炎(ASH)和非酒精性脂肪性肝炎(NASH)影响很大 美国和世界各地的人口。目前，未得到满足的主要医疗需求包括缺乏方法或 特异地耗尽活化的HSC和毛细血管的LSEC以及非侵入性的试剂 可视化胶原堆积、HSC激活和LSEC的方法和试剂 肝纤维化中的毛细血管形成。我们已经开发出一种蛋白质药物候选药物(称为 靶向整合素αvβ3的新位点诱导表达整合素αvβ3的细胞凋亡 通过一种新的机制。ProAgio特异性诱导整合素v3表达的细胞凋亡 通过在的胞浆区域招募和激活caspase8而获得高效的细胞。我们 我们的初步研究表明，对携带肝纤维化/肝硬变的小鼠的治疗 TAA/乙醇、CCl_4和高脂饲料诱导的NASH小鼠ProAgio翻转肝模型 纤维化/肝硬变。此外，我们还开发了新型蛋白质磁共振造影剂(PROCAS)， 使我们能够评估纤维化肝脏中的胶原含量和整合素αvβ3阳性的肝星状细胞和肝内皮细胞。 肝纤维化小鼠的磁共振成像显示我们开发的造影剂具有优越的性能 对胶原蛋白和整合素v3阳性细胞进行评估。对健康人群的初步毒性分析 小鼠表明，ProAgio和我们开发的MRI造影剂对小鼠在 高剂量。该项目的目标是大力进行ProAgio作为药物的临床前验证 ASH/NASH患者治疗的候选人。我们将通过三个具体目标来实现我们的目标。 目的1是研究ProAgio在高脂饮食下逆转肝纤维化的有效性。 再加上多次酗酒和慢性酗酒诱发的ASH模型。对这一事件的调查 ProAgio在ASH小鼠模型中的作用将进一步在临床前验证ProAgio作为一种 ASH/NASH治疗药物。目的2是监测和验证ProAgio对胶原的影响 用我们研制的磁共振造影剂对纤维化的肝脏进行磁共振成像和肝星状细胞。我们的先生 影像辅助验证不仅将验证ProAgio作为ASH/NASH的有效性 治疗剂，也验证了药物作用的靶点和作用机制。目标3处于临床前阶段 ProAgio作为ASH/NASH治疗药物候选药物的毒理学(TOX)和 药代动力学(PK)分析。我们的研究将为ASH/NASH的治疗开辟一条新的途径 通过蛋白质设计进行诊断/预后。我们研究的成功不仅将开发出一种新的蛋白质 治疗肝纤维化/肝硬化的药物，但也测试高效的MRI造影剂 使我们能够准确和非侵入性地监测纤维化的进展和消退 治疗效果评估。这样的发展有望填补主要的医学空白 并便于制定逆转纤维化的治疗策略和跟踪高危患者。

Summary Alcoholic Steatohepatitis (ASH) and Nonalcoholic Steatohepatitis (NASH) affects a large population in US and worldwide. Currently, major unmet medical needs include lack of method or agent to specifically deplete activated HSC and capillarized LSEC as well as noninvasive methodology and reagents to visualize collagen build-up, HSC activation, and LSEC capillarization in fibrotic liver. We have developed a protein drug candidate (referred to as “ProAgio”) that targets integrin αvβ3 at a novel site to induce apoptosis of integrin αvβ3 expressing cells by a novel mechanism. ProAgio specifically induces apoptosis of integrin v3 expressing cells with a high efficacy by recruiting & activating caspase 8 at cytoplasmic domain of. We demonstrated in our preliminary studies that treatment of mice that carries liver fibrosis/cirrhosis induced by TAA/alcohol CCl4 and the high-fat diet induce NASH mice with ProAgio reversed liver fibrosis/cirrhosis. In addition, we have developed novel protein MRI contrast agents (ProCAs) that allow us to assess collagen contents and integrin αvβ3 positive HSCs & LSECs in fibrotic liver. MR imaging of fibrotic mice demonstrated superior properties of our developed contrast agents for collagen and integrin v3 positive cell assessment. Preliminary toxicity analyses with healthy mice indicate that ProAgio and our developed MRI contrast agents are not toxic to mice at very high dose. The goal of this project is to vigorously pre-clinical validation of ProAgio as a drug candidate for ASH/NASH patient treatment. We will achieve our objective by three specific aims. Aim 1 is to examine the effectiveness of ProAgio in reversal of liver fibrosis using high-fat diet plus multiple binge alcohol and chronic alcohol binge induced ASH models. Investigation of the effects of ProAgio in ASH mouse models will further pre-clinical validation of ProAgio as an ASH/NASH treatment drug. Aim 2 is to monitor and validate the effects of ProAgio on collagen and HSC in fibrotic liver by MR imaging using our developed MRI contrast agents. Our MR imaging aided validation will not only validate the effectiveness of ProAgio as an ASH/NASH treatment agent, but also validate the target and mechanism of drug action. Aim 3 is Pre-clinical validations of ProAgio as an ASH/NASH treatment drug candidate via toxicology (TOX) and pharmacokinetic (PK) analyses. Our study will open a new avenue for ASH/NASH treatment and diagnosis/prognosis by protein design. Success in our studies will not only develop a new protein drug for liver fibrosis/cirrhosis treatment but also test highly effective MRI contrast agents that allow us to accurately and non-invasively monitor fibrosis progression and regression for assessment of treatment effects. Such development is expected to fill in the major medical gaps and to facilitate to devise treatment strategy to reverse fibrosis and follow high risk patients.