南京大學吳稚偉課題組嘗試使用外泌體靶向清除HIV體內儲存庫

導 讀

雖然高效抗逆轉錄病毒療法能夠有效控制艾滋病病毒複製，可以將外周血中的病毒複製控制在極低的水平。然而，HIV體內潛伏感染存儲庫（reservoir）的存在為進一步清除病毒帶來了巨大的障礙。特別是，腸道淋巴結等實體組織作為病毒體內重要的存儲庫，由於抗逆轉錄病毒藥物無法穿透實體組織投遞藥物，因而無法達到抑制實體組織存儲庫中病毒的效果。

細胞外泌體（exosome）是哺乳類動物細胞分泌的具有磷脂膜結構的納米顆粒。作為藥物投遞載體，外泌體具有免疫原性低，組織細胞親和力高，導入效率高等優點。吳稚偉課題組在研究外泌體作為細胞間信息交流載體和免疫信號傳遞的過程中發現外泌體具有良好載貨和投遞能力（Fu, et al., PloS Pathogen 2017），在最新的研究中，吳稚偉課題組嘗試使用外泌體作為藥物載體將藥物投遞到表達HIV-1標記的細胞和組織，研究清除艾滋病病毒體內存儲庫的可行性。

圖1：表面表達10E8ScFv抗體靶向分子的外泌體的構建過程

在該項研究中，吳稚偉團隊將HIV-1包膜蛋白envelope特異的人源單鏈可變區抗體（10E8ScFv）表達在外泌體表面上作為載體的靶向分子。該研究詳細分析了抗體靶向分子對提高外泌體對HIV-1感染的宿主細胞的靶向性和導入效率的作用。利用表達Env三聚體的CHO細胞模型，該研究發現10E8ScFv可以大幅提高外泌體載體的靶向性；進一步地，HIV潛伏細胞模型，假病毒模型和HIV-1病人外周淋巴細胞都證明，10E8ScFv外泌體能夠將藥物或誘導細胞凋亡的miRNA-143特異性投遞進入細胞，殺死並誘導凋亡。通過使用表達Env三聚體的CHO細胞在NCG小鼠上誘導表達Env三聚體的實體腫瘤，該研究發現10E8ScFv外泌體能夠將curcumin特異性投遞到固體組織上，抑制腫瘤的生長，說明10E8ScFv外泌體藥物可以特異性有效清除實體組織中的病毒。

圖2：假病毒模型（A-D）、ACH2潛伏感染細胞（E-G）模型和慢性感染者PBMC（H）的研究都證明了HIV-1感染靶向性外泌體10E8ScFv外泌體可以殺死HIV感染細胞並抑制病毒複製。

圖3：HIV-1感染靶向性外泌體10E8ScFv外泌體可以靶向Env 腫瘤組織，顯示了清除實體組織中病毒存儲庫的潛力

結語

該研究首次嘗試利用外泌體作為藥物載體，使用細胞和實體組織模型，清除HIV體內病毒存儲庫，證明了該策略的可行性。該研究以「Extracellular vesicles expressing a single-chain variable fragment of an HIV-1 specific antibody selectively target Env tissues」為題發表於醫學期刊Theranostics 。

該研究工作得到了復旦大學黃競荷教授，中山大學鄧凱教授的大力支持。論文的第一作者為博士研究生鄒雪，通訊作者為南京大學醫學院公共健康醫學中心吳稚偉教授和鄭楠博士。

Abstract

Rationale:Antiretroviral therapy can effectively suppress HIV-1 replication in the peripheral blood to an undetectable level. However, elimination of the latent virus in reservoirs remains a challenge and is a major obstacle in the treatment of HIV-1-infected patients. Exosomes exhibit huge promise as an endogenous drug delivery nanosystem for delivering drugs to solid tissues given their unique properties, including low immunogenicity, innate stability, high delivery efficiency, and most importantly the ability to penetrate solid tissues due to their lipophilic properties.

Methods:We engineered and expressed the scFv of a high affinity HIV-1-specific monoclonal antibody, 10E8, on the exosomal surface (10E8scFv-exos). Subsequently, the 10E8scFv-exos were loaded with curcumin (Cur), a chemical that kills HIV-1-infected cells, or miR-143, an apoptosis-inducing miRNA. We tested the ability of 10E8scFv-exos to deliver cargo to Env target cells and tissues, as well as their ability to suppress HIV-1 infection.

Results:10E8scFv-exos efficiently targeted CHO cells expressing a trimeric gp140 on their surface (Env cells)in vitro, as demonstrated by confocal imaging and flow cytometry. 10E8scFv-exos loaded with Cur or miR-143 showed specific killing of Env cells. In addition, 10E8scFv-exos loaded with Cur or miR-143 could suppress p24 expression in an HIV-1 latency cell line ACH2 and in PBMCs from an ART-treated HIV-1-infected patient. In an NCG mouse model grafted with tumorigenic Env CHO cells and which had developed solid tissue tumors, intravenously injected 10E8scFv-exos targeted the Env-expressing tissues and delivered Cur to induce a strong suppression of the Env tumor growth with low toxicity.

Conclusion:In principle, engineered exosomes can deliver anti-HIV agents to solid tissues by specifically targeting cells expressing viral envelop proteins and inducing cell killing, suggesting that such an approach could be developed for eradicating virus-infected cells in tissue reservoirs.

本期編輯：vetjamie

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