| 摘要: | 亨丁頓氏症（Huntington′s disease, HD） ，也常被稱作為亨丁頓氏舞蹈症是一種由基
因突變引起的神經退化性疾病，導致有毒且錯誤摺疊的突變型亨丁頓蛋白（mutant
huntingtin, mHTT）生成。此類疾病目前無法完全治癒且現有治療手段有限。本研究團隊
先前已開發以胜肽為基礎的藥物用於治療 HD，但胜肽在生理環境中易受到酵素降解，
限制其治療效果。為克服此限制，我們選用中孔洞矽奈米材料（Mesoporous silica
nanoparticles, MSNs）作為藥物載體，其具有改善藥物動力學特性（如能夠延長體內循環
時間與可控的釋放能力）、富含高表面積與可調控表面化學性質等優點。
在本研究中，我們在功能化的 MSNs 上，以化學方法修飾了具雙親性質的 E6Q10 胜
肽。該胜肽序列同時結合了能與 mHTT 結合的聚麩醯胺，以及帶負電的親水序列，用以
解聚 mHTT 聚集體。透過傅立葉轉換紅外光譜，我們驗證了 E6Q10 胜肽能成功修飾於
MSNs 表面（MSN-TA-E6Q10）；為提升奈米材料在溶液中的分散性，我們進一步以 Pluronic F127 修飾於 MSN-TA-E6Q10 表面，並藉由動態光散射儀驗證；穿透式電子顯微鏡顯示MSN-胜肽複合物型態為球形，平均粒徑約為 25 奈米。此外，為了改善藥物的藥物動力學特性，本研究透過將治療性胜肽（E6Q10）以雙硫鍵鍵結於 MSN 表面，使其對細胞質
環境中的高還原性條件（如高濃度的穀胱甘肽）具敏感性。進一步透過體外釋放實驗驗
證此設計，在模擬生理環境中能保持封裝在載體中，且在高還原性環境能下有效觸發藥
物釋放，達成控制釋放效果。
進一步的生物應用研究證實， MSN-TA-E6Q10-F127 在多種濃度下均表現良好生物
相容性。在疾病模型中，使用表達 HTT(Q)109-eYFP 的神經母細胞瘤細胞株細胞進行實
驗，並以 MSN-TA-E6Q10-F127 進行實驗。結果顯示 MSN-TA-E6Q10-F127 可有效降低
mHTT 聚集並緩解其所引發的細胞毒性。此外，以 GFP 與磷酸化 H2AX 為指標進行
分析，觀察到兩者表現量皆顯著下降，顯示 mHTT 聚集體減少，並伴隨 DNA 損傷程
度降低。
本研究透過將胜肽（例如 E6Q10）包覆於中孔洞矽奈米材料中，有效提升其在生理
環境中的穩定性與治療潛力，為針對 mHTT 之治療策略提供一新方向，亦展現了奈米醫
學於神經退化性疾病治療中的應用潛力。;Huntington′s disease (HD) is a neurodegenerative disorder caused by a genetic mutation that leads to the production of the toxic, misfolded mutant huntingtin protein (mHTT). This devastating condition remains incurable, with limited therapeutic options. Previously, our group developed several peptide-based drugs to treat HD. However, these peptides were found to be highly susceptible to enzymatic degradation in biological systems, limiting their therapeutic potential. To address this challenge, mesoporous silica nanoparticles (MSNs) were employed
as drug carriers due to their favorable pharmacokinetic properties, including prolonged circulation time and controlled drug release, as well as their high surface area and versatile surface chemistry.
In this study, MSNs were functionalized with the amphiphilic peptide E6Q10, which
integrates polyglutamine-binding sequences with negatively charged sequences to disaggregate mHTT aggregates. Fourier-transform infrared spectroscopy (FT-IR) confirmed the successful conjugation of E6Q10 to MSNs (as MSN-TA-E6Q10). To enhance dispersion, MSN-TA-E6Q10 nanoparticles were coated with either Pluronic F127, as verified by dynamic light scattering (DLS). Transmission electron microscopy (TEM) revealed that the spherical MSN-peptide complexes with an average size of approximately 25 nm. Furthermore, to refine properties of the peptide drug, E6Q10 was conjugated to the MSN surface via disulfide linkages, making it responsive to the highly reducing intracellular environment (such as elevated glutathione levels). In vitro release studies confirmed that nanoparticles retained the peptide under physiological-mimicking conditions, while a reducing environment effectively triggered peptide release, thereby achieving controlled release.
Further bio application, we confirmed that MSN-TA-E6Q10-F127 exhibited low
cytotoxicity and good biocompatibility across various concentrations. For the disease model, neuroblastoma cell line Neuro-2a (N2a) expressing HTT(Q)109-eYFP were used and treated with the aforementioned MSNs. MSN-TA-E6Q10-F127 effectively reduced mHTT aggregation and toxicity. Further, we demonstrated decreased levels of green fluorescence protein (GFP) and H2A histone family member X (p-H2AX), indicating a reduction in mHTT aggregates and associated DNA damage.
This research incorporated peptide drugs (i.e., E6Q10) into MSNs to address the challenges of delivering therapeutic agents for treating HD, offering another strategy for targeting mHTT and advancing nanomedicine for neurodegenerative disease. |
