靶向lncRNA UCA1和Hedgehog通路的siRNA/药物共载体系的构建及其抗胆管癌作用及机制研究

Current chemotherapies are not effective for inhibition the progress of cholangiocarcinoma(CCA) and the overall 5-year survival rate of patients with CCA is less than 10%. Our preliminary results indicate that lncRNA UCA1 was highly expressed in CCA tissue，which led to poor prognosis and might related to chemotherapy resistance. While cancer associated fibroblasts (CAFs) could trigger CCA resist the apoptosis induced by lncRNA UCA1 knockdown. Then we hypothesized that when silencing lncRNA UCA1 induces cell apoptosis of CCA cells, CAFs secreted excess PDGF-BB, and Hedgehog signaling pathway was activated in CCA cells, thus resulting in apoptosis resistance. However, the regulation mechanism between CCA cells and CAFs is still unclear. Based on the previous study, we intend to prepare a “core-shell” structure nanoparticles with PLL-PPhe as the core and liposome as the shell, which used for simultaneously deliver lncRNA UCA1 siRNA and Sonidegib, a Hedgehog signaling pathway inhibitor. With further in vitro assays and in vivo animal experiments, this project attempt to demonstrate the synergistic effect of this codelivery system in the treatment of cholangiocarcinoma, reveal the molecular mechanism underlying the CCA cells regulate the secretion of PDGF-BB from CAFs, and identify the role of PDGF-BB on Hedgehog signaling pathway in CCA cells.

胆管癌化疗敏感性差，5年总体生存率不足10%。申请人前期研究发现胆管癌组织过表达lncRNA UCA1导致患者预后不良，可能是胆管癌化疗不敏感的原因，肿瘤相关成纤维细胞（CAFs）可促使CCA细胞抵抗lncRNA UCA1沉默诱导的凋亡。我们推测当沉默lncRNA UCA1诱导胆管癌细胞凋亡时，CAFs分泌过量的PDGF-BB细胞因子，激活胆管癌细胞Hedgehog信号通路，导致胆管癌细胞凋亡抵抗，但两种细胞之间相互调控的机制尚不明确。本研究拟在前期基础上，制备以PLL-PPhe为核心、脂质体为外壳的“核-壳”结构共载药纳米粒，用于siRNA和Hedgehog信号通路抑制剂Sonidegib的同步输送，并通过细胞学实验和动物实验，验证共输送体系在胆管癌治疗中的协同作用，揭示胆管癌细胞调控CAFs分泌PDGF-BB的分子机制，明确PDGF-BB对胆管癌细胞Hedgehog信号通路的具体作用。

胆管癌化疗敏感性差，5年总体生存率不足10%。课题组研究发现降低胆管癌细胞lncRNA UCA1表达可促进其凋亡，而肿瘤相关成纤维细胞（CAFs）分泌的PDGF-BB激活胆管癌细胞Hedgehog信号通路，导致胆管癌细胞凋亡抵抗。本项目主要研究内容是：1.纳米载药输送体系的构建及体内外抗胆管癌疗效研究2. CAFs 分泌PDGF-BB并进一步激活胆管癌细胞Hedgehog 信号通路的作用机制研究。通过本项目的研究，成功构建了稳定的质粒DNA/靶向药物的共输送体系，体外实验和小动物PDX模型研究表明，负载lncRNA UCA1质粒DNA和Hedgehog信号通路抑制剂Sonidegib共载药纳米粒可有效抑制胆管癌细胞增殖和侵袭转移能力，并明显抑制荷瘤小鼠的肿瘤生长；揭示了CAFs 分泌PDGF-BB通过改变PTCH1、SUFU表达激活胆管癌细胞Hedgehog 信号通路的作用机制。本项目制备的核酸和小分子药物共输送的纳米药物治疗体系，为临床胆管癌的治疗提供了新策略。