Preclinical development of AlaTIMP-2 as an cancer therapeutic

AlaTIMP-2 作为癌症治疗剂的临床前开发

• 批准号: 7966212
• 负责人: William Stetler-Stevenson
• 金额: $ 101.24万
• 依托单位: DIVISION OF BASIC SCIENCES - NCI
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/7966212
• 关键词: A549; Active Sites; Baculoviruses; Binding; Binding Sites; Biological Assay; Cell Surface Receptors; Cells; Development; Dose; Effectiveness; Extracellular Matrix; Family; Goals; Growth; Homeostasis; Homologous Gene; Human; Integrin alpha3; Laboratories; Lung; Maintenance; Malignant Neoplasms; Malignant neoplasm of lung; Mammalian Cell; Materials Testing; Matrix Metalloproteinases; Metalloproteases; Modeling; Mus; Neoplasm Metastasis; Normal tissue morphology; Peptides; Production; Recombinant Proteins; Recombinants; Relative (related person); Retroviral Vector; Role; System; Therapeutic; Therapeutic Agents; Tissue Inhibitor of Metalloproteinases; Transforming Growth Factor beta; Treatment Effectiveness; Treatment Protocols; Work; Xenograft procedure; analog; high throughput screening; in vivo; inhibitor/antagonist; melanoma; member; neoplastic cell; novel; peptide analog; pre-clinical; research study; small molecule; tumor; tumorigenic

Working towards our stated goal of preclinical development of TIMP-2 as a novel cancer therapeutic we have performed in vivo xenograft growth experiments. These experiments have shown that the TIMP-2 homolog lacking MMP inhibitory activity known as Ala+TIMP-2 effectively inhibits growth of the human A549 lung cancer xenograft, when overexperssed in the tumor cells using a retroviral vector system. To further the development of Ala+TIMP-2 as a therapeutic we have developed both baculovirus and mammalian cell expression systems for the production of recombinant Ala+TIMP-2 and TIMP-2. It is our plan to test these materials as therapeutic agents in both xenogfraft and syngeneic murine tumor models to demonstrate the effectiveness of treatment with exogenous recombinant proteins. These models will focus on the treatment of lung cancer (A549 and Lewis Lung) as well as melanoma (A2058 and B16F10). Various dosing regimens will be utilized to compare the relative in vivo effectiveness of Ala+TIMP-2 compared to TIMP-2. Preliminary studies indicate that Ala+TIMP-2 is more effective than TIMP-2, which is attributed to the fact that Ala+TIMP-2 does not bind to the active site of MMP like TIMP-2, therefore effectively increasing its concentration for cell binding sites. Another important aspect of this project is to determine if we can develop peptide analogs that could be utilized for in vivo therapy. Furthermore we propose to develop a high throughput screening assay to screen synthetic small molecule analogs that can compete for TIMP-2, Ala+TIMP-2, or TIMP peptide binding to the cell surface receptor integrin alpha3 beta1, that we have shown modulates the anti-angiogenic and anti-tumorigenic activity of Ala+TIMP-2.

为了实现TIMP-2作为一种新型癌症治疗药物的临床前开发目标，我们已经进行了体内异种移植物生长实验。这些实验表明，缺乏MMP抑制活性的TIMP-2同源物，即Ala+TIMP-2，当使用逆转录病毒载体系统在肿瘤细胞中过表达时，可有效抑制人类A549肺癌异种移植物的生长。为了进一步开发Ala+TIMP-2作为一种治疗方法，我们开发了杆状病毒和哺乳动物细胞表达系统，用于生产重组Ala+TIMP-2和TIMP-2。我们计划在异种移植和同基因小鼠肿瘤模型中测试这些材料作为治疗剂，以证明外源性重组蛋白治疗的有效性。这些模型将专注于肺癌（A549和Lewis lung）以及黑色素瘤（A2058和B16F10）的治疗。我们将利用不同的给药方案来比较Ala+TIMP-2与TIMP-2在体内的相对有效性。初步研究表明，Ala+TIMP-2比TIMP-2更有效，这是由于Ala+TIMP-2不像TIMP-2那样结合到MMP的活性位点，从而有效地增加了其对细胞结合位点的浓度。该项目的另一个重要方面是确定我们是否可以开发出可用于体内治疗的肽类似物。此外，我们建议开发一种高通量筛选方法来筛选合成的小分子类似物，这些小分子类似物可以竞争TIMP-2、Ala+TIMP-2或TIMP肽与细胞表面受体整合素α 3 β 1的结合，我们已经证明这些小分子类似物可以调节Ala+TIMP-2的抗血管生成和抗肿瘤活性。