EVOLUTION OF ANTIMETASTATIC ENZYME

描述（由申请人提供）：癌症是一种几乎无法治愈的毁灭性疾病。大多数治疗依赖于破坏细胞DNA或破坏肿瘤细胞的细胞周期。转移性黑色素瘤尤其具有毁灭性，由于其对常规化疗和放疗的相对抗性，通常会迅速致命。然而，在黑色素瘤和其他癌症的肿瘤发生和转移中，氨基酸酪氨酸激活了一个新的途径。值得注意的是，在黑色素瘤细胞系和小鼠模型中，酪氨酸的消耗抑制了肿瘤的侵袭和转移。酪氨酸的消耗可以通过饮食限制或通过酪氨酸酚裂解酶（TPL）的分解代谢来完成。Km测量的TPL活性较低，需要大量的酶进行进一步的动物研究或最终的人类治疗。“DNA洗牌”是一种能够通过基于DNA同源性的过程进化增强蛋白质的方法。然而，这种方法并不适用于TPL，因为其结构同源物在DNA水平上并不相似。IntegriGen正在开发一种非同源“洗牌”基因的方法。该方法将应用于TPL，以增强其功能的测量Km。因此，该项目有两个应用：1)进化一种具有增强活性的新型TPL变体，可随后作为抗癌治疗进行评估；2)建立一种非同源“洗牌”基因的新方法，作为一种适用于蛋白质工程的分子进化技术。

DESCRIPTION (provided by applicant): Cancer is a devastating disease with few cures. Most treatments rely on damaging cellular DNA or disrupting the cell cycle in tumor cells. Metastatic melanoma is especially devastating, as it is often rapidly fatal due to its relative resistance to conventional chemo and radiation therapy. However, a novel pathway involved in tumorigenesis and metastasis in melanoma and other cancers is activated by the amino acid tyrosine. Remarkably, tumor invasion and metastases are suppressed by depletion of tyrosine in melanoma cell lines, as well as mouse models. Tyrosine depletion can be accomplished by dietary restriction or through catabolism by the enzyme tyrosine phenol-lyase (TPL). The activity of TPL as measured by Km is low, requiring large amounts of the enzyme for further animal investigation or eventual human therapy. "DNA shuffling" is one method capable of evolving enhanced proteins through a DNA homology based process. This method, however, is not applicable to TPL because its structural homologs are not similar at the DNA level. IntegriGen is developing a method to non-homologously "shuffle" genes. This method will be applied to TPL in order to enhance its function as measured by Km. Thus, this project has two applications: 1) to evolve a novel TPL variant which has enhanced activity, that can subsequently be evaluated as an anti-cancer therapeutic, and 2) to establish a new method to non-homologously "shuffle" genes as a molecular evolution technology applicable in general to protein engineering. PROPOSED COMMERCIAL APPLICATIONS: NOT AVAILABLE