Development of a high-throughput molecular delivery method using microbubbles and ultrasound and its application to cancer gene therapy

非侵入性的组织特异性分子递送对于广泛治疗的效率和减少副作用至关重要。结合微泡（MB）和超声（US）的物理方法已被认为是少数几种能够非侵入性地将基因递送到靶位点的方法之一。应用于癌症基因治疗，这种方法可以有效地靶向癌症，并且可能比免疫基因治疗更有效，因为患者通常免疫功能低下。然而，仍有几个重要方面有待探讨。首先，空化气泡被认为是分子递送的主要原因;然而，波特性与随后的空化气泡的产生和破裂之间的关系尚未阐明，因此基因转移尚未优化。此外，分子递送和随后的基因表达尚未在体内定量。本研究对这些参数进行了体内外优化，并探讨了其作用机制。 ...更多信息 通过理论分析和计算机分析，阐明了理论传递。假设自由空化气泡是由破碎的MB壳产生的空化核产生的，我们估计了从空化气泡中心引起细胞膜损伤的冲击波传播距离。接下来，我们用分子动力学模拟研究了冲击波作用下磷脂双分子层在水中的结构变化。所产生的崩溃和反弹的双分子层，然后由水分子的渗透到疏水区域的双分子层诱导后的冲击波相互作用被证明。最后，我们评估了该技术在单纯疱疹病毒胸苷激酶介导的自杀基因治疗中的应用。我们观察到通过US/MB介导的转移的肿瘤大小的显著减小。这些数据表明US/MB作为一种新的物理基因递送方法用于癌症基因治疗的潜力。少

Non-invasive, tissue-specific molecular delivery is crucial for the efficiency and reduced side effects of a wide range of treatments. A physical method that combines microbubbles (MB) with ultrasound (US) has been regarded as one of the few methods capable of delivering genes into target sites non-invasively. Applied to cancer gene therapy, this method could efficiently target cancer, and may be more efficient than immune gene therapy, as patients are often immunocompromised. However, several essential aspects remain unexplored. First, cavitation bubbles are believed to be a major cause for molecular delivery; however, the relation between wave characteristics and subsequent generation and collapse of cavitation bubbles has not been elucidated, therefore gene transfer has not been optimized. In addition, molecular delivery and subsequent gene expression have not been quantified in vivo. In the present study, these parameters in vitro and in vivo were optimized, and the mechanism of mo … More lecular delivery with theoretical and computer analysis were elucidated. Hypothesizing that free cavitation bubbles were generated from cavitation nuclei created by fragmented MB shells, we estimated the shock wave propagation distance that would induce cell membrane damage from the center of the cavitation bubble. Next, we investigated the structural change of a phospholipid bilayer in water under the action of a shock wave with molecular dynamics simulations. The resulting collapse and rebound of the bilayer followed by the penetration of water molecules into the hydrophobic region of the bilayer induced after the shock wave interaction were demonstrated. Finally, we evaluated the application of this technology to cancer gene therapy using herpes simplex virus thymidine kinase-mediated suicide gene therapy. We observed dramatic reductions of the tumor size by US/MB-mediated transfer. These data demonstrate the potential of US/MB as a new physical gene delivery method for cancer gene therapy. Less