Neutral pressure-sensitive nanoemulsion for localized siRNA delivery

描述（由申请人提供）：应用小干扰RNA（siRNA）进行癌症治疗的最大障碍是将位点特异性siRNA递送至肿瘤部位的可靠方法。病毒载体具有不可接受的毒性水平，并且阳离子脂质体和聚合物胶束遭受从循环和非特异性递送的快速清除。因此，仍然需要能够有效且特异性地将siRNA递送至靶细胞的长循环非病毒载体。为了满足这些要求，我们提出了一个中立的 超声可降解纳米乳剂，其可以全身给药并以位点特异性方式递送siRNA。该纳米乳剂由涂覆有阳离子亲脂性肽、辅助脂质以及聚乙二醇化脂质和磷脂的混合物的全氟化碳纳米液滴组成。siRNA复合后，加入乙酰化剂以中和残留的表面电荷，这使复合物的非特异性细胞摄取最小化。液态全氟化碳核可以用聚焦超声脉冲蒸发，产生气泡并释放siRNA。额外的声脉冲用于驱动空化（即气泡振荡），其通过气泡/细胞相互作用暂时增加肿瘤细胞的渗透性并促进癌细胞的递送。siRNA.to使用这种创新的方法，我们绕过了内吞途径，这是siRNA进入细胞质的常规途径。此外，siRNA仅在换能器聚焦区内被递送至细胞，从而解决了困扰常规非病毒siRNA递送方法的特异性缺乏的问题。在初步研究中，我们已经证实，这种方法可以成功地将荧光标记的siRNA递送到培养的细胞中，从而成为第一组结合联合收割机压力敏感纳米乳液和超声进行局部siRNA递送的方法。所提出的递送方案有可能解决与当前siRNA递送技术相关的局限性，从而允许siRNA充分发挥其作为癌症治疗剂的潜力。

DESCRIPTION (provided by applicant): The greatest barrier to the application of small interfering RNA (siRNA) for cancer therapy is a reliable method for site-specific siRNA delivery to the tumor site. Viral vectors have an unacceptable level of toxicity, and cationic liposomes and polymeric micelles suffer from rapid clearance from circulation and non-specific delivery. Thus, there remains a need for long- circulating non-viral carriers that can deliver siRNA efficiently and specifically to targeted cells. To fulfill these requirements, we propose a neutral ultrasound-triggerable nanoemulsion that can be administered systemically and deliver siRNA in a site-specific manner. The nanoemulsion consists of perfluorocarbon nanodroplets coated with a mixture of cationic lipophilic peptides, helper lipids, and PEGylated lipids and phospholipids. After siRNA complexation, an acetylating agent is added to neutralize the residual surface charge, which minimizes non- specific cellular uptake of the complex. The liquid perfluorocarbon core can be vaporized with focused ultrasound pulses, creating bubbles and releasing the siRNA. Additional acoustic pulses are used to drive cavitation (i.e. bubble oscilations), which temporarily increases the permeability of tumor cell through bubble/cell interactions and facilitates the delivery of siRNA.to cancer cells. Using this innovative approach, we bypass the endocytic pathway, which is the conventional pathway for siRNA to gain access to the cell cytoplasm. Furthermore, siRNA is delivered to cells only within the transducer focal zone, thus addressing the lack of specificity that plagues conventional non-viral siRNA delivery methods. In preliminary studies, we have confirmed that this approach can successfully deliver fluorescently-labeled siRNA to cultured cells, thus becoming the first group to combine pressure-sensitive nanoemulsions and ultrasound for localized siRNA delivery. The proposed delivery scheme has potential to resolve the limitations associated with current siRNA delivery techniques, thus allowing siRNA to reach its full potential as an agent for cancer therapy.