Clinical Application of Cell Cycle Control Therapy to Rheumatoid Arthritis

在滑膜组织中强制表达细胞周期蛋白依赖性激酶抑制因子(CDKI)基因p16^&lt；INK4a&gt；或p21^&lt；Cip1&gt；可有效治疗类风湿关节炎(RA)动物模型。1)CDKI基因治疗的抗炎作用：CDKI基因治疗通过CDK抑制依赖途径和CDK抑制非依赖途径抑制炎症介质和蛋白水解酶的表达。2)改良腺病毒提高基因转移效率：由于用腺病毒载体进行基因治疗发生致命事故，对腺病毒载体的安全性提出了更高的要求。通过修饰表达Arg-Gly-Asp(RGD)基序的纤维外壳蛋白或携带部分35型腺病毒纤维分子的Ad5，提高了5型腺病毒(Ad5)载体向类风湿滑膜成纤维细胞(RSF)的基因转移效率。与非…相比修饰的Ad5越多，用于治疗RA动物模型滑膜炎症所需的纤维修饰Ad5载体的量越少。3)CDKI-‘生物制品的开发：CDKI治疗无病毒载体的研究。如果HIV跨膜结构域(TAT)被结合，蛋白质可以在细胞内传递。4)合成小分子(Sm)-CDKI对RA动物模型的治疗作用：一些合成的sm-CDKI化合物对RA动物模型的滑膜炎有效，我们已经申请了两种sm-CDKI化合物用于治疗RA的专利。5)选择性p16^&lt；INK4a&gt；p21^&lt；Cip1&gt；诱导RSF中p21^&lt；Cip1&GT；在RSF诱导的p16^&lt；INK4a&GT；表达。在其他成纤维细胞中没有观察到这种p16^&lt；INK4a&gt；诱导。使用荧光差异显示或基因芯片(Affimetrix)技术，我们筛选了一组可能参与p21^&lt；Cip1&gt；-p16^&lt；INK4a&gt；途径的分子。其中一个与怀孕有关的候选分子增加了p16；；INK4a；启动子活性较少

Forced expression of a cyclin-dependent kinase inhibitor (CDKI) gene, p16^<INK4a> or p21^<Cip1> in the synovial tissues was effective in treating animal models of rheumatoid arthritis (RA). Subsequently, we have studied molecular mechanism and application of cell cycle control for treatment of rheumatoid arthritis (RA).1) Anti-inflammatory effects of CDKI gene therapy : CDKI gene therapy suppressed expression of inflammatory mediators and proteinases via both CDK inhibition dependent- and CDK inhibition independent-pathways.2) Improvement of gene transfer efficacy by modified adenovirses : Because of a fatal accident by gene therapy using adenovirus vector, safety of adenovirus vector is strongly demanded. Efficacies of gene transfer to rheumatoid synovial fibroblasts (RSF) by type 5 adenoviruses (Ad5) vector were improved by modification fiber-coat proteins expressing Arg-Gly-Asp (RGD) motif or by Ad5 carrying a part of the fiber molecule of adenovirus serotype 35. Comparing with non- … More modified Ad5, fewer quantities of these fiber-modified Ad5 vectors were required to treat synovitis of RA animal model.3) Development of CDKI-'biologics : CDKI therapy without viral vector was studied. Protein can be delivered intracellularly if it HIV transmembrane domain (TAT) is conjugated. TAT-conjugated p16^<INK4a> fusion protein inhibited RSF proliferation, and therapeutic effects for RA animal model is investigated.4) Synthetic small molecule (sm)-CDKI for treatment of RA : Some synthetic sm-CDKI compounds were effective for synovitis of RA animal model, and we have applied patents of two sm-CDKI compounds for RA therapy.5) Selective p16^<INK4a> induction in RSF by p21^<Cip1> : Forced expression of p21^<Cip1> in RSF induced p16^<INK4a> expression. This p16^<INK4a> induction was not observed in the other fibroblasts. Using fluorescent differential display or GeneChip (Affimetrix) techniques, we screened a group of molecules that might be involved in the p21^<Cip1>-p16^<INK4a> pathway. One of the candidate molecules, that is related to pregnancy, increased p16^<INK4a> promoter activity Less