帶有多功能性與正交性觸發基團的噁唑烷籠型結構用於控制肽醛活性

Abstract

肽醛類化合物因其能與目標蛋白形成可逆共價鍵，被廣泛用作高效的抑制劑。然而，其高反應性容易導致脫靶效應，而且代謝不穩定性導致快速降解，限制了其更廣泛的應用。為了解決這些問題，我們開發了以噁唑烷保護醛基的策略，該策略結合了多種刺激響應性觸發基團來降低醛基的反應性，並能透過特定條件來釋放出醛基。 我們以MG132和Ac-LVK-CHO兩個肽醛為基礎合成的前驅藥物在微酸性和生理條件下皆有很好的水解穩定性。酵素抑制實驗也證實，此保護策略顯著降低了肽醛的活性；且在特定條件下觸發去保護反應恢復了與原藥相當的酵素抑制活性。在正交性實驗顯示，MG132-2NB和Ac-LVK-DiCN表現出部分正交性而MG132-pinBB和Ac-LVK-NIm分別對透過H2O2和nitroreductase去保護，表現出幾乎完全的正交性，在western blot實驗也證明可調控特定藥物被去保護影響細胞內蛋白表達。 因此本研究證明了以噁唑烷保護醛基的策略可以調控肽醛的活性，並且可選擇性和可控性地讓特定肽醛恢復活性，有助於增加肽醛的專一性。

Peptide aldehydes are widely utilized as potent inhibitors due to their ability to form reversible covalent bonds with target proteins. However, their high reactivity leads to off-target effects, and their metabolic instability results in rapid degradation, posing significant challenges for their broader application. To address these issues, we developed an oxazolidine-based caging system that incorporates diverse stimuli-responsive trigger groups, enabling precise spatiotemporal control of inhibitor activation. The oxazolidine cage allowed the integration of trigger groups such as photolabile (2-nitrobenzyl and coumarin derivatives), ROS-sensitive (aryl pinacol boronate), and hypoxia-responsive (nitroimidazole) groups, offering flexibility in prodrug design. The synthesized prodrugs demonstrated excellent hydrolytic stability under mildly acidic (pH 5.0) and physiological (pH 7.4) conditions. Enzyme inhibition assays confirmed that caging significantly reduced inhibitor activity, with IC50 values increasing up to 153-fold for MG132 derivatives and 92-fold for Ac-LVK-CHO derivatives. Controlled decaging using specific stimuli, such as UV light (365 nm), visible light (520 nm), reactive oxygen species (H2O2), or nitroreductase, restored enzymatic inhibition activity to level comparable to those of the uncaged parent drugs. Orthogonality experiments revealed that MG132-2NB and Ac-LVK-DiCN exhibit partial orthogonality. Photodecaging at 365 nm activated both MG132-2NB and Ac-LVK-DiCN, while 520 nm selectively triggered Ac-LVK-DiCN decaging. Similarly, MG132-pinBB and Ac-LVK-NIm responded selectively to H₂O₂ and nitroreductase, respectively, showing nearly complete orthogonality. This study establishes the oxazolidine-based caging system as a robust platform for improving the stability, selectivity, and controllability of peptide aldehyde inhibitors, providing a versatile solution to challenges in reactivity and specificity.