Phosphodiesterase 4B Inhibition as a Therapeutic Target for Alcohol-associated Liver Disease

项目总结 酒精相关性肝病(ALD)是由过量饮酒引起的严重健康问题。 ALD通常被发现与脂肪肝、脂肪变性或纤维化进展有关，从而导致肝硬化。 不幸的是，目前还没有批准的治疗ALD的药物，目前的治疗方法依赖于戒酒、免疫 抑制剂，如皮质类固醇，或肝移植，如果是晚期肝硬变。研究表明， 炎症在ALD的发生和发展中起着重要作用。库普弗细胞(KCs)，居住在 肝脏中的巨噬细胞，主要参与肝脏的炎症反应。KCS，在接触伤者时 肝细胞或肠道细菌内毒素，如脂多糖(LPS)，被激活并分泌 炎症细胞因子，包括肿瘤坏死因子-α和启动炎症。Small对KCs的药理抑制作用 分子，如氯化钯或减少肠道细菌负荷的抗生素被证明可以预防 肝损伤和酒精性肝病进展。环磷酸腺苷(CAMP)信号在血管紧张素转换酶中的重要作用 抑制肝脏损伤和脂肪变性。已知酒精可增加cAMP降解酶的表达 磷酸二酯酶4(PDE4)。上调的PDE4降低cAMP水平并诱导肝脏 炎症/损伤。PDE4抑制剂已被研究为抑制巨噬细胞募集和激活 然而，目前的PDE4抑制剂如罗利普兰和罗氟司特与严重的副作用有关 影响，包括恶心、呕吐、腹泻和消化不良。本研究旨在抑制PDE4B的活性 抑制酒精性脂肪变性的KVA-D-88。我们的初步数据显示，KVA-D-88是一种强大的 具有合适药代动力学特征的选择性PDE4B抑制剂。高选择性PDE4B的研制 像KVA-D-88这样的抑制剂直接应用于人的ALD。然而，PDE4B在其他器官中的抑制，如 作为大脑，众所周知会产生严重的副作用。因此，需要KC特异性药物递送来 增强药物疗效和特异性。我们将开发肝脏特异性PDE4B的KVA-D-88纳米粒 抑制力。我们的具体目标是：1建立PDE4B抑制剂KVA-D-88在体外ALD中的作用； 在活体内。为此，我们将确定与PDE4D相比，KVA-D-88抑制PDE4B同型的特异性 酒精诱导的原代KCs、肝细胞和肝星状细胞(HSCs)的同型。接下来，我们将评估 KVA-D-88介导的PDE4B抑制对cAMP信号转导、肿瘤坏死因子-α诱导的内质网应激和乙醇损伤的影响 在改良的NIAAA小鼠模型中诱导毒性。目标2.制定和表征以KVA为靶点的KCs- 负载D-88的聚乙二醇修饰固体脂质纳米粒(SLN)。为了实现这一目标，我们将 以KVA-D-88为靶向的聚乙二醇脂微球的研制和表征。接下来，我们将对治疗方案进行评估 KVA-D-88负载SLN在NIAAA模型中的作用这个项目意义重大，因为它可以为我们提供 ALD治疗和治疗非酒精性脂肪性肝病(NAFLD)和胆汁淤积症的潜在平台 肝纤维化。

PROJECT SUMMARY Alcohol-associated liver disease (ALD) is a significant health problem caused by excessive alcohol consumption. ALD is generally discovered in association with fatty liver, steatosis, or fibrosis progression, leading to cirrhosis. Unfortunately, there is no approved drug for ALD, and the current therapies rely on abstinence, immune suppressants like corticosteroids, or liver transplantation in case of advanced cirrhosis. Studies suggest that inflammation plays a significant role in ALD initiation and progression. Kupffer cells (KCs), the residing macrophages in the liver, are mainly involved in hepatic inflammatory reactions. KCs, upon contact with injured hepatocytes or gut bacterial endotoxins such as lipopolysaccharides (LPS), get activated and secrete inflammatory cytokines, including TNF-α and start the inflammation. Pharmacological inhibition of KCs by small molecules such as gadolinium chloride or decreasing intestinal bacterial load with antibiotics is shown to prevent liver injury and ALD progression. Cyclic adenosine monophosphate (cAMP) signaling plays a significant role in dampening hepatic injury and steatosis. Alcohol is known to increase the expression of cAMP degrading enzyme phosphodiesterase 4 (PDE4). The upregulated PDE4 decreases the cAMP levels and induces liver inflammation/injury. PDE4 inhibitors have been investigated to inhibit macrophage recruitment and activation in ALD; however, the current PDE4 inhibitors such as rolipram and roflumilast are associated with severe side effects, including nausea, vomiting, diarrhea, and dyspepsia. This study aims to inhibit PDE4B activity by novel inhibitor KVA-D-88 in alcohol-induced steatosis. Our preliminary data show that KVA-D-88 is a potent and selective PDE4B inhibitor with a suitable pharmacokinetic profile. The development of highly selective PDE4B inhibitors such as KVA-D-88 directly applies to human ALD. However, PDE4B inhibition in other organs, such as the brain, is known to produce severe side effects. Therefore, there is a need for KC-specific drug delivery to enhance the drug efficacy and specificity. We will develop KVA-D-88 nanoparticles for liver-specific PDE4B inhibition. Our specific aims are Aim 1 to establish the role of PDE4B inhibitor KVA-D-88 in ALD in vitro and in vivo. In this aim, we will determine the specificity of KVA-D-88 to inhibit PDE4B isotype compared to PDE4D isotype in alcohol-induced primary KCs, hepatocytes, and hepatic stellate cells (HSCs). Next, we will evaluate the effect of KVA-D-88 mediated PDE4B inhibition on cAMP signaling, TNF-α induced ER stress, and ethanol- induced toxicity in the modified NIAAA mouse model. Aim 2. Formulate and characterize KCs targeted KVA- D-88 loaded polyethylene glycol (PEG) decorated solid lipid nanoparticles (SLNs). In this aim, we will develop and characterize KCs targeted PEG-SLNs loaded with KVA-D-88. Next, we will evaluate the therapeutic efficacy of KVA-D-88 loaded SLNs in the NIAAA model. This project is highly significant as it could provide us with ALD therapy and a potential platform for nonalcoholic fatty liver disease (NAFLD) and cholestasis-induced liver fibrosis.