基于“药物-递送-药物”策略的黄芩素-pDNA靶向纳米系统的构建及其抗肺动脉高压研究

Pulmonary arterial hypertension (PAH) that occurs as a result of dysregulated smooth-muscle cells (SMCs) and the subsequent blockade of pulmonary vessels is an inflammatory disease with pulmonary-vascular hemodynamic abnormality. Because of the specificity of the diseased location and the inherent deficiency of traditional drug delivery system, no efficient treatment strategy of PAH is reported until now. Baicalein (BCL) is a drug that is able to potently resist inflammation; and while, protein 53 (p53), a plasmid DNA (pDNA), can efficiently inhibit the proliferation of diseased cells. Therefore, their combined use has a promising potential to cure PAH. Our previous findings demonstrated that a rod-shaped drug nanocrystal allowed for potent intracellular delivery of biopharmaceuticals by bypassing the lysosomal route, consequently establishing a “drug-delivering-drug (DDD)” platform. In this study, a BCL-p53 targeted nanoplatform will be constructed via the DDD strategy, aiming to robustly deliver the gene drug, p53, to the nucleus as well as achieve a synergistic treatment of PAH. The platform will be prepared as follows: using BCL rod-like nanocrystals (BRNs) as carriers, the p53-Sis1p complex is condensed on the BRNs followed by gluconic acid (GA) coating, in which the coating of GA is to target to the glucose transporter-1 (GLUT-1) highly expressed on SMCs and Sis1p, a shuttled protein between the cytosol and nucleus, specifically binds with the nuclear pore complex (NPC) and thus transport p53 to the nucleus. Overall, via targeting to the pulmonary vessel-SMC-organelle axis, the platform is promising to deliver nucleic acid to the nucleus and achieve combined treatment of PAH as well.

肺动脉高压（PAH）系肺血管血液动力学异常的炎症型疾病，源于平滑肌细胞（SMCs）过快增殖导致的肺血管堵塞等。由于其病灶的特殊性与传统递药系统的内在缺陷，至今尚无有效的治疗策略。黄芩素（BCL）能有效抵抗炎症，p53（一种pDNA）能抑制病变细胞的增殖。因此，二者联用有望高效治疗PAH。前期发现：难溶性药物棒状纳米晶能绕开溶酶体，递送大分子药物入胞；据此构建了“药物-递送-药物（drug-delivering-drug，DDD）”技术平台。本项目，拟构建基于DDD技术的BCL-p53靶向纳米系统，以高效递送p53入核并联合治疗PAH：以BCL棒状纳米结晶为载体，负载p53-Sis1p复合物并包裹葡萄糖醛酸（GA）；其中，GA靶向SMCs的GLUT-1受体促进胞吞；Sis1p靶向核孔复合体，转运p53入核。通过靶向肺血管-SMCs-细胞器，该体系有望高效递送核酸药物入核、联合治疗PAH。

肺动脉高压是一种肺血管血液动力学异常的炎症型心肺疾病。肺血管重构导致的右心重构和心衰是病人死亡的根本原因。临床药物只能缓解症状，无法根本治疗。本项目利用“药物晶体-药物”复合物的策略，构建了抗炎黄芩素晶体-促凋亡质粒p53共递送制剂，并评价了其野百合碱（MCT）-诱导肺高压模型的治疗作用，最后探索了制剂的潜在作用机制。研究结果发现：（1）构建的药物晶体-大分子药物复合物共递送制剂，载药量提高到了80% VS. 传统载体（5%）。（2）150-200 nm棒状制剂能高效靶向肺-肺血管-肺血管平滑肌细胞，在肺部的分布远大于其他器官（肝除外）。（3）制剂能以非溶酶体内吞方式进入细胞：主要通过小窝蛋白途径入胞（75%），避免了溶酶体对基因药物的破坏，极大地提高了药物的胞内递送效率。（4）递送p53至PASMCs能有效抑制其增殖，且共递送黄芩素晶体-p53能抑制增殖和抑制炎症。（5）该共递送制剂能有效抑制肺血管重构，改善血流动力学，改善右心功能等。（6）协同机制研究表明，制剂主要通过激活p53/Bax/Bcl-2/ Cas-3信号通路，促进PASMCs凋亡。申请人的这些工作表明，该制剂能有效抑制肺血管重构也能高效治疗肺高压，有望为肺高压的治疗提供新的思路。棒状晶体制剂能靶向肺血管，这些发现对于肺血管靶向制剂的设计与开发，具有良好的借鉴意义。本课题只验证了制剂对MCT-模型（突发性肺高压）的治疗作用，设计的制剂对其他模型如低氧、肺血管纤维化导致的肺高压是否有效，尚待进一步验证。