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弹性蛋白样聚合物（SELP）基质，用于携带RB 94肿瘤抑制基因（Ad-RB 94）的腺病毒构建体的微创控制递送。基本原理是通过SELP的基因工程，可以定制在室温下为液体的递送系统，在温和条件下与腺病毒肿瘤抑制基因治疗载体混合，在单次瘤内注射后在体温下形成载有载体的水凝胶，在所需的时间段内在肿瘤部位释放活载体，并以最小的全身毒性和最大的功效杀死肿瘤细胞。本论文的主要目的如下：1）合成和表征SELP水凝胶，用于腺病毒基因的局部和可控递送。将使用重组技术生物合成和表征含有具有各种序列和长度的丝样和弹性蛋白样重复单元的线性SELP共聚物类似物。水凝胶将由线性聚合物形成。水凝胶的溶胀度将作为在生理温度、pH和离子强度下聚合物结构和初始聚合物浓度的函数进行检查。2)检测聚合物和腺病毒组合物对溶胀度、腺病毒颗粒释放和体外生物活性的影响。将模型腺病毒掺入水凝胶中。将在存在腺病毒颗粒的情况下评价肿胀程度。随着时间的推移，释放量将作为聚合物结构和浓度的函数进行评价。释放将与相关体外模型中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，将使用适当的聚合物和Ad-RB 94组合物来检查释放的治疗性肿瘤抑制基因的生物活性。3)评价聚合物组成对体内含腺病毒SELP水凝胶的转导效率、转基因表达持续时间、生物分布、治疗功效和毒性的影响。将使用头颈癌的裸鼠肿瘤异种移植模型，通过瘤内给予含有Ad-GFP（含有绿色荧光蛋白基因作为基因转移标志物的腺病毒）和Ad-RB 94的SELP水凝胶来评价这些参数。在未来的阶段中，提出的基质介导的腺病毒基因治疗方法，使用基因工程聚合物可用于开发临床上可接受的系统，用于基因治疗的头颈癌。

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.