CNC-Center for Neuroscience and Cell Biology, University of Coimbra, Largo Marquês de Pombal, 3004-517 Coimbra, Portugal; FFUC-Faculty of Pharmacy, University of Coimbra, Pólo das Ciências da Saúde, Azinhaga de Santa Comba, 3000-548 Coimbra, Portugal.
CNC-Center for Neuroscience and Cell Biology, University of Coimbra, Largo Marquês de Pombal, 3004-517 Coimbra, Portugal.
J Control Release. 2014 Dec 28;196:122-31. doi: 10.1016/j.jconrel.2014.09.024. Epub 2014 Oct 11.
Drug resistance remains the Achilles tendon undermining the success of chemotherapy. It has been recognized that success requires the identification of compounds that, when combined, lead to synergistic tumor inhibition while simultaneously minimizing systemic toxicity. However, in vivo application of such protocols is dependent on the ability to deliver the appropriate drug ratio at the tumor level. In this respect, nanotechnology-based delivery platforms, like liposomes, offer an elegant solution for the in vivo translation of such strategy. In this work, we propose the active intracellular delivery of combinations of doxorubicin and the pro-apoptotic sphingolipid, C6-ceramide, using our previously described cytosolic triggered release-enabling liposomes, targeting nucleolin with the F3 peptide. Combination of doxorubicin (DXR):C6-ceramide (C6-Cer) at 1:2 molar ratio interacted synergistically against drug resistant/triple negative MDA-MB-231 breast cancer cells, as well as drug sensitive MDA-MB-435S melanoma cells. Cell viability studies indicated that F3-targeted liposomes encapsulating DXR:C6-Cer 1:2 molar ratio (p[F3]DC12) performed similarly as targeted liposomal DXR (p[F3]SL), encapsulating twice the amount of DXR, at the IC50, for an incubation time of 24 h. Importantly, F3-targeted liposomes encapsulating DXR:C6-Cer 1:2 molar ratio (p[F3]DC12) enabled a cell death above 90% at 24 h of treatment against both DXR-resistant and sensitive cells, unattainable by the F3-targeted liposomal doxorubicin. Furthermore, a F3-targeted formulation encapsulating a mildly additive/antagonistic DXR:C6-Cer 1:1 molar ratio (p[F3]DC11) enabled an effect above 90% for an incubation period as short as 4 h, suggesting that the delivery route at the cell level may shift the nature of drug interaction. Such activity, including the one for p[F3]DC12, induced a marked cell and nucleus swelling at similar extent, consistent with necrotic cell death. Overall, these results demonstrated that F3-targeted intracellular delivery of different DXR/C6-Cer ratios, with diversed drug interactions, enabled a highly relevant increased efficacy against chemotherapy resistant cells.
耐药性仍然是阻碍化疗成功的阿喀琉斯之踵。人们已经认识到，成功需要识别出化合物，当这些化合物组合使用时，能在同时最小化全身毒性的情况下导致协同的肿瘤抑制。然而，这种方案的体内应用取决于在肿瘤水平上输送适当药物比例的能力。在这方面，基于纳米技术的递药平台，如脂质体，为这种策略的体内转化提供了一种优雅的解决方案。在这项工作中，我们使用之前描述的细胞质触发释放使能脂质体，通过 F3 肽靶向核仁素，提出了阿霉素（DXR）和促凋亡神经酰胺（C6-神经酰胺，C6-Cer）组合的主动细胞内递药。摩尔比为 1:2 的阿霉素（DXR）：C6-神经酰胺（C6-Cer）组合对耐药/三阴性 MDA-MB-231 乳腺癌细胞以及药物敏感的 MDA-MB-435S 黑色素瘤细胞具有协同作用。细胞活力研究表明，在 24 小时孵育时间内，摩尔比为 1:2 的 F3 靶向脂质体包封 DXR：C6-Cer（p[F3]DC12）与包封两倍量 DXR 的靶向脂质体 DXR（p[F3]SL）的 IC50 相似。重要的是，摩尔比为 1:2 的 F3 靶向脂质体包封 DXR：C6-Cer（p[F3]DC12）能够在 24 小时的治疗中使 DXR 耐药和敏感细胞的细胞死亡超过 90%，而 F3 靶向脂质体阿霉素则无法达到这一效果。此外，包封摩尔比为 1:1 的轻度相加/拮抗的 DXR：C6-Cer（p[F3]DC11）的 F3 靶向制剂能够在短至 4 小时的孵育时间内产生超过 90%的效果，这表明细胞水平的递药途径可能改变药物相互作用的性质。这种活性，包括 p[F3]DC12 的活性，导致细胞和核肿胀程度相似，这与坏死性细胞死亡一致。总的来说，这些结果表明，F3 靶向的不同 DXR/C6-Cer 比例的细胞内递药，具有不同的药物相互作用，能够显著提高对化疗耐药细胞的疗效。
