| 摘要: | 在正常細胞的生長及組織器官的發育中，蛋白質酪胺酸激酶原本即扮演著相當重要的角色，然而近年來的研究指出，蛋白質酪胺酸激酶過度地表現活性與許多細胞過度增殖疾病有關，如癌症、血癌、牛皮癬等疾病，蛋白質酪胺酸激酶因而成為癌症治療領域中相當熱門的研究領域。在細胞訊息傳遞路徑中，生長因子與其相對應的受器型蛋白質酪胺酸激酶的活性對於腫瘤細胞的生長具有舉足輕重的影響，有鑑於此，生化學家們便企圖尋找出蛋白質酪胺酸激酶抑制劑，並希望由此找尋出治療癌症的曙光。 本文中，我們測試由中大李文仁教授所合成的一系列環形類??化合物：2,5-dioxopiperazine衍生物，意欲模擬蛋白質酪胺酸激酶的受質結合位置，進而阻礙蛋白質酪胺酸激酶的活性，而達到抑制癌細胞生長的目的；同時我們也企圖找尋出2,5-dioxopi- perazine衍生物的結構與生長抑制活性間的關係，初步發現2,5- dioxopiperazine衍生物的R1及R2取代基對於抑制癌細胞生長的活性較為重要。在測試的2,5- dioxopiperazine衍生物中，p194 (IC50 = 110-130mM)，p185 (IC50 = 90 mM)，C1 (IC50 = 100 mM)，p206 (IC50 = 5-7 mM)及p236-3 (IC50 = 45-55 mM)具有明顯抑制癌細胞生長的活性，然而p194、p185及C1對於細胞中蛋白質酪胺酸的磷酸化並無明顯的影響。相反地，p206明顯會影響蛋白質酪胺酸的磷酸化，並且造成有絲分裂期的停滯；而p236- 3雖然對細胞週期沒有明顯影響，但具有明顯抑制多種人類癌細胞的能力，且對人類正常的纖維母細胞無生長抑制的作用；然而 p236-3對於蛋白質酪胺酸激酶的作用卻與當初藥物設計的目的恰好相反，p236-3的處理會促使肺癌細胞中60及70 kDa蛋白質酪胺酸部位磷酸化的增加，此結果似乎說明p236-3應為去磷酸酶的抑制劑。此磷酸化的修飾作用與生長抑制活性間的關係值得進一步的研究。 Protein Tyrosine Kinases (PTKs) play a key role in normal cell and tissue development. However, enhanced PTK activity is correlated with proliferative diseases, such as cancers, leukemias, psoriasis, and restenosis. Recently, PTKs have emerged as crucial targets for therapeutic intervention in cancers. In the signal transduction pathways, growth factor ligands and their respective receptor tyrosine kinases (RTKs) have been shown to be required for tumor cell growth. This realization have prompted bioorganic chemists to systematically synthesize tyrosine kin- ase inhibitors. In this study, we have applied a panel of cyclic-peptidomimetic compounds 2,5-dioxopiperazine derivatives(provided by Dr. Lee), and attempted to screen their growth inhibitory effect on tumor cells. Furthermore, we will also take a shot at organizing the relationship of cyclic-peptidomimetic compounds structure and inhibitory effect. Among these, p194, p185, C1, p206 and p236-3 showed promising inhibitory effect on tumor cell growth with IC50= 110-130 mM, 90 mM, 100 mM, 5-7 mM and 45-55 mM respectively. p194, p185 and C1 had no significantly effect on protein phosphorylation profile on tyrosine residues. In contrast, p206 treatment on H460 cells did alter several proteins phosphorylation and resulted in mitosis-arrest. The degradation of spindle protein Mad2 was also significantly inhibited by p206 treatment in H460 cells. Although, p236-3 had no effect on cell cycle progress, it showed differential cytotoxicity between tumor and normal fibroblast cells. p236-3 almost had no inhibitory effect on human normal fibroblast cells. The Western blotting results with phosphotyrosine antibody showed that this compound dose-dependently enhanced the phosphorylation level of two proteins around 70 and 60 kDa. These results suggest that p236-3 may be an inhibitor of phosphatase, instead of tyrosine kinase. The inhibitory mechanism of p236-3 and the two proteins (70 and 60 kDa) deserves for further investigation. |
