Onivyde于2015年获得食品药品监督管理局（FDA）批准，用于治疗包括转移性胰腺癌在内的实性肿瘤。它的设计目的是将伊立替康浓缩包裹在内，延长其在血液中的循环寿命，并将其运送到细胞中，然后通过酶的作用将其转化为具有100至1000倍更高抗癌活性的SN-38代谢物。尽管在临床途径方面表现出积极的效果，但对Onivyde中伊立替康的物理状态以及该合成物在从制造过程到细胞内处理过程中的变化知之甚少。本研究利用伊立替康的固有荧光和荧光寿命成像显微镜（FLIM）选择性地探测药物的超分子组织结构。我们对制造商的配方进行FLIM分析，揭示了Onivyde脂质体内两种共存的物理状态：（i）凝胶/沉淀型伊立替康和（ii）与脂质体膜相关的伊立替康，这两种状态并不可从制造商的指示中推测出来。通过融合高效液相色谱法（HPLC）和伊立替康的难透过细胞膜的动态猝灭剂，我们发现在将Onivyde稀释至标准细胞培养培养基中时，凝胶/沉淀相迅速（在几分钟内）完全溶解，并导致伊立替康前药从脂质体中大量泄露。事实上，共聚焦成像和细胞增殖实验证明，封装和非封装伊立替康制剂在细胞摄取机制和细胞分裂抑制方面是相似的。最后，2通道FLIM分析将伊立替康和其红移的SN-38代谢物的特征区分开来，证明了后者作为Onivyde细胞内加工的结果出现。本研究的发现为Onivyde的合成特性及其从生产到体外实验中的转变提供了新的见解。此外，这些结果进一步验证了利用FLIM分析来阐明封装荧光药物的超分子组织结构的有效性。这项研究强调了利用先进的成像技术加深对药物配方的理解并优化其在递送应用中的性能的重要性。

Onivyde was approved by the Food and Drug Administration (FDA) in 2015 for the treatment of solid tumors, including metastatic pancreatic cancer. It is designed to encapsulate irinotecan at high concentration, increase its blood-circulation lifetime, and deliver it to cells where it is enzymatically converted into SN-38, a metabolite with 100- to 1000-fold higher anticancer activity. Despite a rewarding clinical path, little is known about the physical state of encapsulated irinotecan within Onivyde and how this synthetic identity changes throughout the process from manufacturing to intracellular processing. Herein, we exploit irinotecan intrinsic fluorescence and fluorescence lifetime imaging microscopy (FLIM) to selectively probe the supramolecular organization of the drug. FLIM analysis on the manufacturer's formulation reveals the presence of two coexisting physical states within Onivyde liposomes: (i) gelated/precipitated irinotecan and (ii) liposome-membrane-associated irinotecan, the presence of which is not inferable from the manufacturer's indications. FLIM in combination with high-performance liquid chromatography (HPLC) and a membrane-impermeable dynamic quencher of irinotecan reveals rapid (within minutes) and complete chemical dissolution of the gelated/precipitated phase upon Onivyde dilution in standard cell-culturing medium with extensive leakage of the prodrug from liposomes. Indeed, confocal imaging and cell-proliferation assays show that encapsulated and nonencapsulated irinotecan formulations are similar in terms of cell-uptake mechanism and cell-division inhibition. Finally, 2-channel FLIM analysis discriminates the signature of irinotecan from that of its red-shifted SN-38 metabolite, demonstrating the appearance of the latter as a result of Onivyde intracellular processing. The findings presented in this study offer fresh insights into the synthetic identity of Onivyde and its transformation from production to in vitro administration. Moreover, these results serve as another validation of the effectiveness of FLIM analysis in elucidating the supramolecular organization of encapsulated fluorescent drugs. This research underscores the importance of leveraging advanced imaging techniques to deepen our understanding of drug formulations and optimize their performance in delivery applications.