PO.CL07.04 · 临床研究
MDM2抑制剂与亲脂性吉西他滨的联合：在肺癌中呈强协同作用，而在脑癌中呈拮抗作用
Combination of MDM2 inhibitor and lipophilic gemcitabine: Strong synergism in lung cancer, while antagonism in brain cancer
作者与单位 Authors & Affiliations
摘要 Abstract
中文摘要
即使在后COVID-19时代，癌症仍是全球首要的死亡原因。肺癌和脑癌占这些死亡的相当大比例。尽管有多种抗癌药物可用，但单药疗法常因疗效有限或耐药产生而无法实现持久反应。因此，联合治疗被认为是增强治疗疗效并延缓耐药的有力策略。在本研究中，我们探讨了第二代MDM2抑制剂Idasanutlin(Ida)与吉西他滨亲脂性前药Gemcitabine Elaidate(Gem Eli)联合治疗非小细胞肺癌(NSCLC)和胶质母细胞瘤(GBM)的协同潜力。这两种分子均表现出疏水特性，促使其被掺入脂质体载体的脂质双分子层中。采用改良的水化方法，我们开发了共载Ida和Gem Eli的脂质体纳米制剂(IG)。一个主要挑战是Ida(一种"砖粉"分子)的快速不溶性沉淀。为解决此问题，我们引入了阳离子可电离脂质DLin-DMA，以促进阴离子性Ida与阳离子性DLin-DMA之间形成电荷可调复合物。可电离脂质在生理pH下保持中性，但在酸性条件下(如肿瘤微环境，pH约5.5)带正电荷，从而促进肿瘤靶向递送并最大限度减少全身毒性。IG制剂在NSCLC细胞系(A549和H460)和一个GBM细胞系(U-87)中进行了评估，这些细胞均携带野生型p53。在NSCLC中观察到强协同作用，联合指数值为0.07(A549)和0.3(H460)。相比之下，U-87细胞未表现出协同作用；相反，Gem Eli表现出拮抗行为，在1:1至1:20的各种比例下与Ida联用时IC50增加约7–10倍。3D球体实验进一步证实了GBM中的拮抗作用，经过10天隔日治疗后，IG、Gem Eli和Ida分别使球体面积相比对照减少了81%、61.9%和81%。NSCLC中的协同作用通过Combenefit分析得到进一步验证，其在3D协同图中显示出强协同作用。优化后的IG脂质体表现出131.1 ± 0.68 nm的粒径、0.172 ± 0.13的PDI，以及pH响应性zeta电位-2.06 ± 0.06(pH 7.4)和+8.88 ± 0.31(pH 5.5)，支持其适用于肿瘤靶向递送。在小鼠红细胞中的血液相容性检测显示，在浓度高达100 μM时溶血率<1%。冷冻电子显微镜证实了均一的颗粒形态，无聚集迹象。正在进行的研究着重于阐明GBM中所观察到的拮抗作用背后的细胞机制。
查看英文原文 English abstract
Cancer remains the leading cause of mortality worldwide, even in the post-COVID-19 era. Lung and brain cancers account for a substantial proportion of these deaths. Although multiple anticancer agents are available, single-agent therapies often fail to achieve durable responses due to limited efficacy or the development of drug resistance. Combination therapy is therefore recognized as a powerful strategy to enhance therapeutic efficacy and delay resistance.In this study, we investigated the synergistic potential of Idasanutlin (Ida), a second-generation MDM2 inhibitor, and Gemcitabine Elaidate (Gem Eli), a lipophilic prodrug of Gemcitabine, for the treatment of non-small cell lung cancer (NSCLC) and glioblastoma (GBM). Both molecules exhibit hydrophobic characteristics, prompting their incorporation into the lipid bilayer of liposomal carriers. Using a modified hydration method, we developed a liposomal nanoformulation co-loaded with Ida and Gem Eli (IG). A major challenge was the rapid insoluble precipitation of Ida, a brick-dust molecule. To address this, we introduced the cationic ionizable lipid DLin-DMA to enable the formation of a charge-tunable complex between anionic Ida and cationic DLin-DMA. Ionizable lipids remain neutral at physiological pH but acquire a positive charge in acidic conditions (e.g., the tumor microenvironment, pH ~5.5), which facilitates tumor-targeted delivery while minimizing systemic toxicity. The IG formulation was evaluated in NSCLC cell lines (A549 and H460) and a GBM cell line (U-87), all harboring wild-type p53. Strong synergism was observed in NSCLC, with combination index values of 0.07 (A549) and 0.3 (H460). In contrast, U-87 cells demonstrated no synergism; instead, Gem Eli displayed antagonistic behavior, with a ~7-10-fold increase in IC₅₀ when combined with Ida across ratios ranging from 1:1 to 1:20. A 3D spheroid assay further confirmed antagonism in GBM, after 10 days of alternate-day treatment, IG, Gem Eli, and Ida reduced spheroid area by 81%, 61.9%, and 81%, respectively, compared to controls. Synergism in NSCLC was further validated using Combenefit analysis, which demonstrated robust synergy in 3D synergy maps. The optimized IG liposomes exhibited a particle size of 131.1 ± 0.68 nm, a PDI of 0.172 ± 0.13, and pH-responsive zeta potentials of -2.06 ± 0.06 (pH 7.4) and +8.88 ± 0.31 (pH 5.5), supporting their suitability for tumor-targeted delivery. Hemocompatibility testing in mouse red blood cells showed <1% hemolysis at concentrations up to 100 µM. Cryo-electron microscopy confirmed uniform particle morphology with no evidence of aggregation. Ongoing studies are focused on elucidating the cellular mechanisms underlying the observed antagonism in GBM.
利益披露 Disclosure
B. M. Dholariya, None.