Echogenic Polymersomes for Triggered Contents Release

 描述：聚合体是由合成的两亲性嵌段共聚物制备的囊泡。与脂质体相比，它们有几个优点，包括增强的稳定性，更长的循环时间，机械稳定性，以及能够携带大量疏水和亲水药物分子。共聚物的亲水嵌段通常是聚乙二醇酯(PEG)，这赋予了所得聚合体长循环的性质。由于稳健性，多聚体需要刺激来充分干扰致密的双层，并释放包裹的内容物。尽管脂质体被广泛研究用于药物递送应用，但聚合体在靶向、触发内容物释放和同时成像方面的潜力在很大程度上仍未被探索和开发。在这一应用中，我们建议合成两亲性嵌段共聚物，并制备长循环聚合物。聚合体会主动靶向肿瘤组织，并在癌细胞内化后被触发释放被包裹的内容物。此外，我们将在聚合体中封装气泡，使其具有回声特性。这将允许我们同时进行超声成像，并使用诊断频率超声对内容物释放施加附加控制。建议的多模式多聚体的有效性将通过球体培养和耐去势前列腺癌的原位小鼠模型进行验证。拟议研究的目的概述如下。(1)两亲性嵌段共聚物的合成，以及回声、可触发释放的聚合体的制备。我们将合成含有聚乙二醇亲水部分的嵌段共聚物。疏水链段的结构和相对分子质量将被系统地改变，以优化聚合体的形成过程。为了赋予刺激响应特性，我们将通过还原敏感(用于细胞内释放)或低氧敏感连接(用于细胞外释放)连接合成的亲水和疏水聚合物块。为了主动靶向和肿瘤穿透，我们将把已报道的环状IRGD多肽化学连接到聚乙二醇端。这些聚合物将制备回声聚合体，包裹血管生成抑制剂suntinib(用于细胞外释放)或抗癌药物多西他赛，以及组蛋白去乙酰酶抑制剂莫西替坦(用于细胞内释放)。(2)回声多聚体的声学特性。我们将使用与生理相关的细胞还原剂浓度、谷胱甘肽和低氧条件来演示还原和低氧介导的包膜内容物的释放。将使用详细的声学特性、测量衰减、线性和非线性散射来研究聚合体的回声特性。我们将确定共振特性。我们将通过测量在逐渐增强的声激励下的时间依赖衰减来确定破坏这些聚合体的阈值激发水平。超声波对含量变化的影响 将研究聚合体的释放情况。将确定线性和非线性(谐波和次谐波)成像方式的最佳声学参数(激励频率、脉冲波形和强度)。(3)应用球形细胞培养和抗去势前列腺癌原位小鼠模型验证药物传递系统的有效性。我们将生成抗去势的前列腺癌细胞的三维球体，以评估我们的聚合体输送微囊化药物的有效性。在细胞内给药方面，我们将探讨多西紫杉醇和莫西替坦克服耐药性的协同作用。我们将使用肿瘤球体来确定聚合体的穿透性、细胞毒性和癌细胞生长的减少。随后，我们将建立耐去势前列腺癌的原位小鼠模型，以评估我们的多聚体制剂传递其包裹药物的有效性。将确定应用诊断频率超声的效果，并对小鼠的球体和肿瘤进行成像。我们将进一步确定释放的药物在减少肿瘤干细胞方面的效果，治疗方案的配方，肿瘤的缓解，以及药物在不同器官的潜在毒性。

 DESCRIPTION: Polymersomes are vesicles prepared from synthetic, amphiphilic-block copolymers. They have several advantages over liposomes, including enhanced stability, longer circulation times, mechanical robustness, and the ability to carry large quantities of hydrophobic and hydrophilic drug molecules. The hydrophilic block of the copolymers is usually polyethyleneglycol (PEG), imparting the long-circulating property to the resultant polymersomes. Because of robustness, the polymersomes require a stimulus to sufficiently disturb the compact bilayer, and to release the encapsulated contents. Although liposomes are extensively studied for drug delivery applications, the potential of polymersomes for targeting, triggered contents release, and simultaneous imaging remains largely unexplored and undeveloped. In this application, we are proposing to synthesize amphiphilic block copolymers, and prepare long-circulating polymersomes. The polymersomes will be actively targeted to tumor tissues, and triggered to release the encapsulated contents upon internalization in the cancer cells. In addition, we will encapsulate gas bubbles in the polymersomes to render them echogenic. This will allow us to perform simultaneous ultrasound imaging, and exert an addition control over contents release employing diagnostic frequency ultrasound. The effectiveness of the proposed multimodal polymersomes will be demonstrated using spheroid cultures and orthotopic mouse model for castration resistant prostate cancer. The Aims of the proposed studies are summarized below. (1) Synthesis of amphiphilic block copolymers, and preparation of echogenic, trigger-releasable polymersomes. We will synthesize bock copolymers containing PEG as the hydrophilic part. The structure as well as the molecular weight of the hydrophobic block will be systematically varied to optimize the polymersome formation process. In order to impart stimuli-responsive property, we will connect the synthesized hydrophilic and hydrophobic polymer blocks via either a reduction-sensitive (for intracellular release) or a hypoxia-sensitive link (for extracellular release). For active targeting and tumor penetration, we will chemically attach the reported cyclic iRGD peptide to the PEG terminus. Echogenic polymersomes will be prepared from these polymers, encapsulating the angiogenesis inhibitor suntinib (for extracellular release) or the anticancer drug docetaxel, and the histone deacetylase inhibitor mocetinostat (for intracellular release). (2) Acoustic characterization of the echogenic polymersomes. We will demonstrate reduction and hypoxia- mediated release of encapsulated contents employing physiologically-relevant concentrations of the cellular reducing agent, glutathione and hypoxic conditions. Echogenicity of the polymersomes will be investigated using detailed acoustic characterization, measuring attenuation, linear, and nonlinear scattering. We will determine the resonance characteristics. We will determine the threshold excitation level for destruction of these polymersomes by measuring time dependent attenuation under progressively higher acoustic excitations. The effect of ultrasound on altering the rate of content release from the polymersomes will be investigated. Optimum acoustic parameters (excitation frequency, pulse waveform and strength) will be determined for linear and nonlinear (harmonic and subharmonic) imaging modalities. (3) Demonstration of effectiveness of the drug delivery system employing spheroid cell cultures and orthotopic mouse model of castration resistant prostate cancer. We will generate the three dimensional spheroids of the castration resistant, prostate cancer cells to evaluate the effectiveness of our polymersomes to deliver the encapsulated drugs. For intracellular delivery, we will probe the synergistic effects of docetaxel and mocetinostat to overcome drug resistance. We will determine the penetration of the polymersomes, cytotoxicity, and reduction in growth of the cancer cells using the tumor spheroids. Subsequently, we will generate orthotopic mouse models of castration resistant prostate cancer to evaluate the effectiveness of our polymersome formulation to deliver their encapsulated drugs. The effect of applied diagnostic frequency ultrasound will be determined, and the spheroids and the tumors in mice will be imaged. We will further determine the effects of the released drugs in reducing the cancer stem cells, the therapeutic regimen for the formulations, remission of the tumors, and potential toxicity of the drugs in different organs.