Engineering Polymers For Gene Therapy of Head Cancer

描述(由申请人提供)： 这项研究的长期目标是设计聚合物递送系统，以提高头颈癌基因治疗的疗效并降低毒性。本项目的目的是设计丝状弹性聚合物(SELP)基质，用于携带RB94肿瘤抑制基因(Ad-RB94)的腺病毒构建物的微创可控递送。其基本原理是，通过SELP的基因工程，可以量身定制室温下液态的递送系统，在温和的条件下与腺病毒抑癌基因治疗载体混合，在单次肿瘤内注射后，在体温下形成载体载水凝胶，在所需的时间内在肿瘤部位释放活载体，以最低的系统毒性和最高的疗效杀死肿瘤细胞。将致力于以下具体目标：1)合成和表征SELP水凝胶，用于局部和受控腺病毒基因传递。含有不同序列和长度的丝状和弹性蛋白样重复单元的线性SEP共聚类似物将被生物合成并使用重组技术进行表征。水凝胶将由线型聚合物形成。水凝胶的溶胀度将作为聚合物结构和生理温度下的初始聚合物浓度、pH和离子强度的函数进行检测。2)考察聚合物和腺病毒组成对体外溶胀度、腺病毒颗粒释放和生物活性的影响。模型腺病毒将被掺入水凝胶中。肿胀程度将在腺病毒颗粒存在的情况下进行评估。释放的量将随着时间的推移作为聚合物结构和浓度的函数进行评估。在相关的体外模型中，释放将与颗粒的生物活性相关。根据模型腺病毒释放和体外基因转移的结果，将使用合适的聚合物和Ad-RB94组合物来检测释放的治疗性抑癌基因的生物活性。3)评价聚合物组成对腺病毒SELP水凝胶的体内转导效率、转基因表达持续时间、生物分布、治疗效果和毒性的影响。在头颈部肿瘤裸鼠移植瘤模型中，通过瘤内注射含Ad-GFP(腺病毒携带绿色荧光蛋白基因作为基因转移标志)和Ad-RB94的SELP水凝胶来评估这些参数。在未来阶段，所提出的使用基因工程聚合物的基质介导腺病毒基因治疗方法可用于开发临床可接受的头颈癌基因治疗系统。

DESCRIPTION (provided by applicant): The long-term goal of this research is to engineer polymeric delivery systems that improve the efficacy and reduce the toxicity of head and neck cancer gene therapy. The purpose of this project is to engineer silk-elastinlike polymeric (SELP) matrices for minimally invasive controlled delivery of adenovirus constructs carrying the RB94 tumor suppressor gene (Ad-RB94). The rationale is that by genetic engineering of SELPs, it is possible to tailor-make delivery systems that are liquid at room temperature, mixed under mild conditions with adenoviral tumor suppressor gene therapy vectors, form vector-laden hydrogels at body temperature after a single intratumoral injection, release viable vectors at the site of tumor over a desired period of time, and kill the tumor cells with minimum systemic toxicity and maximum efficacy. The following Specific Aims will be addressed: 1) To synthesize and characterize SELP hydrogels for localized and controlled adenoviral gene delivery. Linear SELP copolymer analogs containing silk-like and elastin-like repeating units with various sequences and lengths will be biosynthesized and characterized using recombinant techniques. Hydrogels will be formed from the linear polymers. The degree of swelling of the hydrogels will be examined as a function of polymer structure and initial polymer concentration at physiological temperature, pH, and ionic strength. 2) To examine the influence of polymer and adenoviral composition on the degree of swelling, adenoviral particle release and bioactivity in vitro. Model adenoviruses will be incorporated in the hydrogels. The degree of swelling will be evaluated in the presence of adenoviral particles. The amount released will be evaluated over time as a function of polymer structure and concentration. Release will be correlated with the bioactivity of the particles in relevant in vitro models. From the results of model adenoviral release and in vitro gene transfer, appropriate polymer and Ad-RB94 compositions will be used to examine the bioactivity of the released therapeutic tumor suppressor gene. 3) To evaluate the influence of polymer composition on transduction efficiency, duration of transgene expression, biodistribution, therapeutic efficacy, and toxicity of adenoviral-containing SELP hydrogels in vivo. A nude mouse tumor xenograft model of head and neck cancer will be used to evaluate these parameters by intratumoral administration of SELP hydrogels containing Ad-GFP (Adenoviruses containing green fluorescent protein gene as markers of gene transfer) and Ad-RB94. In the future phases, the proposed matrix-mediated adenoviral .gene therapy approach using genetically engineered polymers can be used for the development of clinically acceptable systems for gene therapy of head and neck cancer.