背景：癌症是威胁人类健康的重大疾病之一，化疗普遍存在生物利用度低、毒 副作用大、易产生耐药性以及靶向性差等问题，许多与肿瘤相关的药物递送系 统出现，用于提高疗效、增加安全性。目前，药物递送系统研究面临的技术瓶 颈问题是如何将药物精准定向输送到病灶部位(靶向性)，实现可控的药物释放， 从而提高药物的体内生物利用度和降低药物的副作用。脂质体(Liposome， Lipo)作为一种广泛使用的纳米药物递送系统能够在一定程度上提高化疗药物 的利用度，但依旧存在靶向性不足且容易被机体免疫系统快速清除的问题，红 细胞(Red blood cells，RBCs)具有很长的体内循环时间，它凭借着没有免疫原 性、易获取以及无需体外培养等诸多优点，成为了很好的药物天然递送载体。 鉴于 Lipo 的可改造型(根据需求负载不同药物)与 RBCs 的优点(长循环时间)， 本研究构建的 RBCs-lipo 联用药物递送系统能有效提升肿瘤靶向性、延长其在体 内的循环时间且减少肝脾对其的清除，从而提高其抗肿瘤效果。 方法:基于肿瘤微环境呈酸性(pH<7)的特点，本研究通过薄膜旋蒸法制备 的两种 Lipo，DPPC 脂质体(Lipo-DPPC)以及含有 pH 响应磷脂的 DOPE 脂质 体(Lipo-DOPE)。通过 Lipo 与 RBCs 膜结合方式的差异来调控递送药物的靶向 性和半衰期，即将 Lipo 粘附在 RBCs 膜表面或与 RBCs 膜融合。本研究还构建 了血小板膜(Platelet membrane，PLM)修饰的 Lipo(PLM-lipo)药物递送系统， 将其与 RBC-lipo 递送系统联用可以提高 PLM-lipo 的血液循环时间及其在肺部的 富集，从而实现肺癌的靶向治疗，有效地预防或减少 CTCs 的肺转移，延长肿 瘤患者的生存期。

结果:将载药 Lipo 与 RBCs 结合，可有效增强血液循环寿命，降低肝肾清除， 提高 Lipo 抗肿瘤治疗效果。研究结果证明血液长循环时间和肿瘤靶向能力都是 提高抗肿瘤疗效的关键。粘附在 RBCs 表面的 Lipo 可以通过红细胞搭便车 (RBCs-hitchhiking，RH)技术的原理将 Lipo 富集于肺部(被动靶向)，使得药 物在肺部富集;而与 RBCs 膜融合的 Lipo 会随着 RBCs 在体内进行长时间循环 并在血液中持续缓释药物，且在48 h内不会富集于任何脏器。因此Lipo与RBCs的联用可以提高药物在体内的生物利用度，提高肿瘤疗效。为了能够更好的增 加肺癌的靶向性并且进一步减少肝肾对 Lipo 的清除效率，本研究使用 PLM 修 饰 Lipo，PLM 与 Lipo 融合能够显著提高肿瘤的靶向性，通过 RH 技术使 PLM- lipo 粘附至 RBCs 膜上构建 RBC-PLM-lipo，可通过血液循环至肺部时因毛细血 管外力的挤压而将 PLM-lipo 转移至肺部，从而提高肺部的靶向性，同时还能延 长在血液中的循环时间，减少肝脾的清除以降低对机体的毒副作用。 结论:PLM 与 RH 技术的联用能够提升现有的 Lipo 载药系统的生物利用度，将 RBCs 和 PLM 的优势赋予 Lipo 后改善其靶向性不足和半衰期短的缺陷，有望成 为肺癌治疗的一种新方式，也为自体血细胞载药技术在肿瘤临床治疗方面的发 展打下前期基础。

Background: Cancer is one of the major diseases that threaten human health. Chemotherapy generally has problems such as low bioavailability, large toxic and side effects, easy to produce drug resistance, and poor targeting. Many tumor-related drug delivery systems appear to improve efficacy and increase safety. At present, the technical bottleneck of drug delivery system research is how to accurately deliver drugs to the lesion site to achieve controlled drug release, thereby improving the in vivo bioavailability of drugs and reducing the side effects of drugs. Liposome (Lipo), as a widely used nano drug delivery system, can improve the utilization of chemotherapeutic drugs to a certain extent, but it still has the problem of insufficient targeting and easy to be quickly cleared by the immune system. Red blood cells (RBCs) have long circulation time in vivo. It has become a good natural drug delivery carrier with many advantages such as no immunogenicity, easy access and no need for in vitro culture. In view of the advantages of Lipo 's modifiable type (loading different drugs according to demand) and RBCs (long circulation time), the RBCs-lipo combined drug delivery system constructed in this study can effectively improve tumor targeting, prolong its circulation time in vivo and reduce its clearance by liver and spleen, thereby improving its anti-tumor effect.

Methods: Based on the acidic nature of the tumor microenvironment (pH < 7), two kinds of Lipo, DPPC liposomes (Lipo-DPPC) and DOPE liposomes containing pH- responsive phospholipids (Lipo-DOPE) were prepared by thin film spin evaporation. The targeting and half-life of the delivery drug are regulated by the difference in the binding mode between Lipo and RBCs membrane, that is, Lipo adheres to the surface of RBCs membrane or fuses with RBCs membrane. In this study, a platelet membrane (PLM) modified Lipo (PLM-lipo) drug delivery system was also constructed. The combination of PLM-lipo and RBC-lipo delivery system can improve the blood circulation time of PLM-lipo and its enrichment in the lung, so as to achieve targeted treatment of lung cancer, effectively prevent or reduce the lung metastasis of CTCs, and prolong the survival time of tumor patients.

Results: The combination of drug-loaded Lipo and RBCs can effectively enhance the life of blood circulation, reduce liver and kidney clearance, and improve the anti-tumor effect of Lipo. The results show that long blood circulation time and tumor targeting ability are the key to improving anti-tumor efficacy. Liposomes adhered to the surface of RBCs can enrich Liposomes in the lungs (passive targeting) through the principle of RBCs-hitchhiking (RH) technology, so that drugs can be enriched in the lungs; the Lipo fused with RBCs membrane will circulate in the body for a long time with RBCs and continuously release the drug in the blood, and will not be enriched in any organ within 48 h. Therefore, the combination of Lipo and RBCs can improve the bioavailability of drugs in vivo and improve tumor efficacy. In order to better increase the targeting of lung cancer and further reduce the clearance efficiency of liver and kidney to Lipo, PLM was used to modify Lipo in this study. The fusion of PLM and Lipo can significantly improve the targeting of tumors. PLM-lipo was adhered to RBCs membrane by RH technology to construct RBC-PLM-lipo, which can transfer PLM- lipo to the lung due to the extrusion of capillary force when blood circulation to the lung, thereby improving the targeting of the lung. At the same time, it can also prolong the circulation time in the blood and reduce the clearance of liver and spleen to reduce the toxic and side effects on the body.

Conclusion: The combination of PLM and RH technology can improve the bioavailability of the existing Lipo drug delivery system. The advantages of RBCs and PLM are given to Lipo to improve its shortcomings of insufficient targeting and short half-life. It is expected to become a new way for the treatment of lung cancer and lay a foundation for the development of autologous blood cell drug delivery technology in the clinical treatment of tumors.