PKD Family Kinase Function and Signaling in Lung Fibroblasts

描述（由申请方提供）：特发性肺纤维化（IPF）是一种病因不明的慢性、进行性和通常致死性肺部疾病，其特征为肺成纤维细胞活化和增殖、细胞外基质（ECM）过度沉积和正常肺结构破坏。IPF患者的长期生存率较差，5年生存率仅为20%。从历史上看，抗炎药一直是治疗的主要手段，但已被证明在降低疾病的严重程度或进展方面无效。因此，迫切需要鉴定新的可药用靶标，以开发用于治疗IPF患者的有效新疗法。由于肺成纤维细胞是IPF中的主要效应细胞，因此鉴定控制IPF肺成纤维细胞生物学的新型药物分子或途径可能导致IPF中更特异和有效的治疗干预。在这项提议中，我们试图确定丝氨酸/苏氨酸蛋白激酶D（PKD）家族激酶是否在IPF成纤维细胞活化、ECM产生和促纤维化生长因子受体表达中发挥关键作用。PKD家族激酶包括PKD 1、PKD 2和PKD 3。在此，我们首次发现所有PKD亚型在成纤维细胞病灶（IPF的标志性病变）中容易检测到。我们进一步表明，PKD 1和PKD 2在原代IPF成纤维细胞中组成性激活，但在对照正常人肺成纤维细胞中不激活。此外，PKD 1和PKD 2可被原代IPF成纤维细胞中的促纤维化生长因子进一步激活。有趣的是，PKD抑制显著下调IPF成纤维细胞中前胶原和纤连蛋白的蛋白水平，而不显著影响其mRNA水平。然而，PKD抑制显著抑制成纤维细胞中促纤维化血小板衍生生长因子受体（PDGFR）的mRNA和蛋白水平。基于这些新发现，该重新提交R21提案旨在检验PKD家族激酶可能在IPF成纤维细胞生物学中发挥关键作用的假设，包括成纤维细胞活化、ECM产生和促纤维化生长因子受体的表达，这可能为阻止IPF进展提供新的药物靶点。本重新提交提案的具体目的是：目的1）表征原代IPF成纤维细胞中PKD家族激酶的组成性和促纤维化激动剂诱导的活化;目的2）确定PKD家族激酶的作用和原代IPF成纤维细胞中促纤维化基质产生的机制;目的3）探讨PKD家族激酶在IPF成纤维细胞表达PDGF受体中的作用及其机制。我们预计，该提案将确定PKD家族激酶是参与IPF发病机制和进展的肺成纤维细胞生物学的新型和重要调节剂，因此靶向抑制PKD家族激酶或特定PKD亚型可能为IPF患者提供新型和有效的治疗干预。 公共卫生相关性：该重新提交的R21提案旨在检验PKD家族激酶可能在肺成纤维细胞生物学中发挥关键作用的假设，包括成纤维细胞活化、细胞外基质产生和促纤维化生长因子受体表达，这可能为阻止特发性肺纤维化进展提供新的药物靶点。

DESCRIPTION (provided by applicant): Idiopathic pulmonary fibrosis (IPF) is a chronic, progressive and usually fatal lung disease of unknown etiology, characterized by lung fibroblast activation and proliferation, excessive deposition of extracellular matrix (ECM), and destruction of the normal lung architecture. Long-term survival of IPF patients is poor, with a 5-year survival rate of only 20%. Historically, anti-inflammatory agents have been the mainstay of therapy, but have been proven to be ineffective in reducing either the severity or progression of the disease. Hence, there is a profound need for the identification of novel drugable targets to develop efficacious new therapies for treating IPF patients. Because lung fibroblasts are primary effector cells in IPF, the identification of novel drugable molecules or pathways that control IPF lung fibroblast biology may lead to more specific and efficacious therapeutic intervention in IPF. In this proposal, we seek to determine whether the serine/threonine protein kinase D (PKD) family kinases play critical roles in IPF fibroblast activation, production of ECM, and expression of profibrotic growth factor receptors. PKD family kinases include PKD1, PKD2 and PKD3. Herein, we found for the first time that all the PKD isoforms were readily detected in fibroblastic foci, the hallmark lesions of IPF. We further showed that PKD1 and PKD2 were constitutively activated in primary IPF fibroblasts but not in control normal human lung fibroblasts. Moreover, PKD1 and PKD2 could be further activated by profibrotic growth factors in primary IPF fibroblasts. Interestingly, PKD inhibition markedly downregulated the protein levels of procollagen and fibronectin without significantly affecting their mRNA levels in IPF fibroblasts. Whereas, PKD inhibition markedly suppressed both mRNA and protein levels of profibrotic platelet-derived growth factor receptor-( (PDGFR() in the fibroblasts. On the basis of these novel findings, this resubmission R21 proposal is designed to test the hypothesis that PKD family kinases may play critical roles in IPF fibroblast biology, including the fibroblast activation, ECM production and expression of profibrotic growth factor receptors, which may offer novel drugable targets for halting IPF progression. The Specific Aims of this resubmission proposal are: Aim 1) To characterize the constitutive and profibrotic agonist-induced activation of PKD family kinases in primary IPF fibroblasts; Aim 2) To determine the roles of PKD family kinases and the mechanisms in the production of profibrotic matrix in primary IPF fibroblasts; Aim 3) To determine the roles of PKD family kinases and the mechanisms in IPF fibroblast expression of profibrotic PDGF receptor-(. We anticipate that this proposal will identify PKD family kinases as novel and important modulators of lung fibroblast biology involved in the pathogenesis and progression of IPF, therefore targeting to inhibit PKD family kinases or a specific PKD isoform may provide a novel and efficacious therapeutic intervention for IPF patients. PUBLIC HEALTH RELEVANCE: This resubmission R21 proposal is designed to test the hypothesis that PKD family kinases may play critical roles in lung fibroblast biology, including the fibroblast activation, production of extracellular matrix, and expression of profibrotic growth factor receptors, which may offer novel drugable targets for halting the progression of idiopathic pulmonary fibrosis.