本研究旨在开发一种纳米结构脂质载体 (NLC)，能够共同递送紫杉醇 (PTX) 和程序性死亡配体 1 (PD-L1) 小干扰 RNA (siRNA)，以增强 PTX 的生物利用度并通过 PD-L1 敲低增强免疫力。我们制备了负载 PTX 的 NLC (P-NLC)，并用带正电荷的壳聚糖 (Chi) 涂覆它以创建 P-NLC-Chi，随后将其与 siRNA (P-NLC-Chi-siRNA) 缀合。 P-NLC-Chi 配方使用 Box-Behnken 设计进行了优化。 P-NLC-Chi 测量为 123.8 ± 0.52 nm（zeta 电位，22.71 ± 0.49 mV）。通过验证凝胶阻滞实验并观察zeta电位的变化，确定NLC​​与PD-L1 siRNA的最佳结合比例为50:1。 P-NLC-Chi-siRNA 粒径为 181.97 ± 0.67 nm，zeta 电位为 18.66 ± 0.23 mV。在 24 小时内观察血清中 siRNA 的稳定性。该复合物的细胞毒性和细胞内摄取在乳腺癌细胞和乳腺癌抗性细胞（分别为MCF-7和MCF-7/ADR细胞）中明显增强。对 P-糖蛋白介导的流出的评估表明，NLC 减轻了 MCF-7/ADR 细胞中的药物流出。将 P-NLC-Chi-siRNA 皮下注射到注射了 MCF-7/ADR 细胞的荷瘤 BALB/c 裸鼠中，结果显示肿瘤尺寸减小。体外和体内实验表明 PD-L1 mRNA 表达水平显着降低。此外，一项体内研究揭示了注射 P-NLC-Chi-siRNA 后肿瘤组织内出现肿瘤特异性 CD4 和 CD8 T 细胞反应。我们的研究结果表明，用于共同递送 PTX 和 PD-L1 siRNA 的 Chi 涂层 NLC 作为化学免疫疗法的创新递送系统具有巨大潜力。版权所有 © 2024 Elsevier B.V. 保留所有权利。

This study aimed to develop a nanostructured lipid carrier (NLC) capable of co-delivering paclitaxel (PTX) and programmed death-ligand 1 (PD-L1) small interfering RNA (siRNA) to enhance PTX bioavailability and bolster immunity through PD-L1 knockdown. We prepared a PTX-loaded NLC (P-NLC) and coated it with positively charged chitosan (Chi) to create P-NLC-Chi, which was subsequently conjugated to siRNA (P-NLC-Chi-siRNA). The P-NLC-Chi formulation was optimized using the Box-Behnken design. P-NLC-Chi measured 123.8 ± 0.52 nm (zeta potential, 22.71 ± 0.49 mV). By verifying the gel retardation assay and observing changes in the zeta potential, the optimal binding ratio of NLC to PD-L1 siRNA was identified as 50:1. The P-NLC-Chi-siRNA particle size was 181.97 ± 0.67 nm, with a zeta potential of 18.66 ± 0.23 mV. siRNA stability was observed in serum over a 24-h period. Enhanced cytotoxicity and intracellular uptake of the complex were evident in breast cancer cells and breast cancer-resistant cells (MCF-7 and MCF-7/ADR cells, respectively). Evaluation of P-glycoprotein-mediated efflux demonstrated that NLC mitigated drug efflux in MCF-7/ADR cells. Subcutaneous injection of P-NLC-Chi-siRNA into tumor-bearing BALB/c nude mice injected with MCF-7/ADR cells revealed a reduction in tumor size. In vitro and in vivo experiments indicated a significant reduction in PD-L1 mRNA expression levels. Additionally, an in vivo study revealed tumor-specific CD4 + and CD8 + T cell responses within the tumor tissue following the injection of P-NLC-Chi-siRNA. Our findings suggest that Chi-coated NLC for the co-delivery of PTX and PD-L1 siRNA has great potential as an innovative delivery system for chemoimmunotherapy.Copyright © 2024 Elsevier B.V. All rights reserved.