婁筱叮、夏帆教授課題組Angew. Chem. Int. Ed.: 多功能性多肽類聚集誘導發光探針實現基因藥物高效可控的細胞核靶向性運輸和實時追蹤

tt 4年前 (2019-02-28) 7681瀏覽

【引言】

基因干擾療法（也稱為基于寡核苷酸的療法）可以在哺乳動物細胞中穩定地沉默或抑制序列特異性基因的表達。其在病原體、癌癥和代謝疾病的治療中獲得了顯著的成果。在基于寡核苷酸的療法中，反義單鏈DNA寡核苷酸（ASO）療法和RNA干擾（RNAi）在過去幾十年中在胞質內和內核干擾方面取得了顯著成果。然而，由于負電荷和核苷酸易于酶促降解以及生物體的多種生物屏障的限制，基于寡核苷酸的療效受到嚴重阻礙。

為了解決基于寡核苷酸療法的局限性，多種載體被進行了深入研究。其中，多肽連接的載體由于其有利的生物相容性和多功能組份而被認為是優良的基因遞送體系之一。這些復合物通常通過靜電相互作用，分子自組裝或共價結合形成穩定的納米顆粒。此外，通過利用各種合成肽，已經開發出用于亞細胞器的有效基因傳送系統。Tseng及其同事使用具有多步修飾的自組裝超分子系統，包含與DNA質粒復合的轉錄因子，細胞穿透肽，整合型肽，聚乙烯亞胺，聚乙二醇，聚酰胺胺樹枝狀大分子，能夠將DNA質粒以精確和高效的方式傳遞到細胞核。此外，已經證實具有復雜和多功能肽的多結構域設計載體可以通過合理排列保留其成分的個體功能。Hilvert及其同事開發了一種簡單有效的模塊化系統，用于體內包裹RNA。然而為了實現高效和準確的治療，寡核苷酸療法應該將亞細胞器靶向、模塊化設計和實時長期跟蹤一起考慮。因此，可實現有效基因傳遞和實時長期追蹤的多功能多肽連接的載體在治療中很有需求。

【成果簡介】

近日，中國地質大學（武漢）婁筱叮、夏帆教授課題組報導了一種多功能的基因傳遞策略，可以以高效和可視化的方式將治療基因傳遞到靶向的細胞核中。作者開發了一種整合素靶向、細胞滲透性和核質體運輸肽連接的AIEgen（TDNCP），用于有效和順序靶向傳遞反義單鏈DNA寡核苷酸（ASO），并進行實時追蹤。與TDNCP/siRNA-NPs相比，TDNCP/ASO-NPs表現出更好地干擾效應，進一步證明TDNCP是細胞核靶向載體。此外，TDNCP/ASO-NPs在體內顯示出良好的腫瘤抑制作用，證明多功能肽連接的AIEgen可用于順序靶向基因傳遞，并實現有效的基因干擾治療。該成果以題為”A Multifunctional Peptide-Conjugated AIEgen for Efficient and Sequential Targeted Gene Delivery into the Nucleus”發表在Angew. Chem. Int. Ed.上。

華中科技大學博士后程勇和孫春麗以及中國地質大學（武漢）博士后劉瑞為論文的共同第一作者，中國地質大學（武漢）婁筱叮教授和夏帆教授為論文的共同通訊作者。該研究工作得到了國家重點研發計劃（2017YFA0208000, 2016YFF0100800）國家自然科學基金（21525523, 21722507, 21574048, 21605053）中國博士后科學基金（2017M620309, 2017M610492）等項目的資助。?

【圖文導讀】

Scheme?1.TNCP和TNCP/ASO自組裝的納米粒子的示意圖

(a).TNCP分子結構

(b).TNCP/ASO-NPs用于實時追蹤和靶向傳遞

Figure 1.TNCP及其衍生物的表征

(a).多肽序列

(b).高效液相色譜表征

(c,e).DGRKRRRR和RGDKRRRR對整合素的分子結合能力與氫鍵作用

(d,f).不同探針與整合素結合的熒光變化

(g,h).用TNCP處理細胞后的細胞熒光強度

Figure 2.細胞實驗

(a-c).TNCP的ASO負載能力

(d).細胞成像

(e).細胞的平均熒光強度和Pearson相關系數

(f).western blot分析

(g).Bio-TEM圖像

圖3.動物實驗

(a).小鼠體重變化

(b).小鼠腫瘤體積變化

(c).Bcl-2基因干擾效率

【小結】

在這個工作中，作者利用多功能肽和AIEgen來實現有效和順序靶向遞送以及跟蹤細胞核。通過整合素靶向肽、細胞滲透性肽和核定位序列的組合，TDNCP比其他衍生物顯示出更優異的生物相容性、核選擇性成像和光穩定性。與TDNCP/siRNA-NPs，lipo/ASONPs和lipo/siRNA-NPs相比，TDNCP/ASO-NPs在整合素過表達的細胞系中顯示出優異的基因干擾效應。由于順序和靶向細胞穿透，TDNCP/ASO-NPs通過瘤內注射顯示出比體內靜脈注射更好的干擾效果。

A Multifunctional Peptide-Conjugated AIEgen for Efficient and Sequential Targeted Gene Delivery into the Nucleus

(Angew. Chem. Int. Ed., 2019, DOI: 10.1002/anie.201901527)

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