公开(公告)号：US5747251A

公开(公告)日：1998-05-05

申请号：US442141

申请日：1995-05-16

申请人： Dennis A. Carson ,  Hitoshi Kohsaka

发明人： Dennis A. Carson ,  Hitoshi Kohsaka

IPC分类号： C12Q1/68 ,  C07H21/04 ,  C12P19/34 ,  C12Q1/70

CPC分类号： C12Q1/6851

摘要： Combinations of polymerization, non-competitive hybridization and assay techniques is disclosed. In one aspect of the method, one member of a regular or anchored primer pair is modified to include a coupling agent capable of forming a tight bond (resistant to uncoupling in an alkaline denaturing environment) with a reactant. Competitive PCR is performed and the PCR products are coupled via the coupling agent to a reactant on the surface of a solid phase support. The bond between the reactant and the solid phase support in this and all embodiments is also resistant to uncoupling in an alkaline denaturing environment. In another aspect of the method, a primer is tightly coupled to the bound reactant and a polymerization of the competitor and target nucleic acids is performed on the solid phase. A third embodiment uses at least three primers, one of which is internal to the PCR templates and is bound to the solid phase support on which the entire PCR takes place. In the first two embodiments, sense strands are completely removed from solution with the bound antisense strands of the PCR products. Hybridization with sequence-specific probes is then performed in the absence of competition for binding by the sense strands. Sense strand removal and hybridization is optional in the third embodiment, where the bound three-primer PCR products can be detected by a detectible signal. Quantification of the target template may preferably be achieved in all embodiments by an enzyme-linked immunosorbent assay (ELISA), using ELISA data analysis software and standard curves.

摘要翻译： 公开了聚合，非竞争性杂交和测定技术的组合。 在该方法的一个方面，将规则或锚定引物对的一个成员修饰为包含能够与反应物形成紧密键合（对碱性变性环境中的解偶联的抗性）的偶联剂。 进行竞争性PCR，PCR产物通过偶联剂偶联到固相载体表面的反应物上。 在这个和所有实施方案中，反应物和固相载体之间的键在碱性变性环境中也抵抗解偶联。 在该方法的另一方面，引物与结合的反应物紧密耦合，并且在固相上进行竞争剂和靶核酸的聚合。 第三个实施方案使用至少三个引物，其中一个引物位于PCR模板的内部，并结合到其上进行整个PCR的固相载体上。 在前两个实施方案中，有义链通过PCR产物的结合反义链从溶液中完全除去。 然后在不存在通过有义链结合的竞争的情况下进行与序列特异性探针的杂交。 在第三实施方案中，有义链去除和杂交是任选的，其中结合的三引物PCR产物可以通过可检测信号检测。 可以使用ELISA数据分析软件和标准曲线，优选通过酶联免疫吸附测定（ELISA）在所有实施方案中实现靶模板的定​​量。

公开(公告)号：US20080226599A1

公开(公告)日：2008-09-18

申请号：US11799353

申请日：2007-05-01

申请人： Nobuyuki Miyasaka ,  Hitoshi Kohsaka

发明人： Nobuyuki Miyasaka ,  Hitoshi Kohsaka

IPC分类号： A61K35/76 ,  A61K38/00 ,  G01N33/00 ,  C12Q1/68 ,  A61P19/00 ,  A01K67/027 ,  C12Q1/02 ,  A61K31/70

CPC分类号： A61K38/005 ,  A61K38/1709 ,  A61K48/00 ,  C12Q1/6897 ,  G01N33/68 ,  G01N33/6863 ,  G01N2333/4739 ,  G01N2500/00 ,  G01N2500/04

摘要： The onset ratios and pathological conditions of collagen-induced arthritis and adjuvant arthritis in model mice and rats, respectively, were successfully ameliorated by topically expressing the cyclin-dependent kinase inhibitors p16INK4a and p21Cip1 p in articular tissues using adenoviral vectors. In the synovial cells of CDKI-transduced mice, expression of inflammatory cytokines was inhibited. Described are the use of the p21Cip1 protein for inhibiting abnormal proliferation of synovial tissues, inflammation in synovial tissues and/or expression of inflammatory cytokines in synovial tissues; the p21Cip1 gene; compounds promoting the activity or expression of the p21Cip1 protein; and pharmaceutical compositions containing these molecules. Also provided are method of screening for compounds participating in the abnormal proliferation of synovial tissues, inflammation in synovial tissues and/or the expression of inflammatory cytokines in synovial tissues targeting the p21Cip1 protein. Rheumatoid arthritis and other disorders associated with inflammation of the synovial tissue can be prevented or treated by promoting expression or function of p21Cip1 protein.

摘要翻译： 通过局部表达细胞周期蛋白依赖性激酶抑制剂p16 INK4a和p21Cip1p，分别成功地改善了模型小鼠和大鼠中胶原诱导的关节炎和辅助性关节炎的发病率和病理状况 在使用腺病毒载体的关节组织中。 在CDKI转导小鼠的滑膜细胞中，炎性细胞因子的表达被抑制。 描述了用于抑制滑膜组织的异常增殖，滑膜组织中的炎症和/或滑膜组织中炎性细胞因子的表达的用途。 p21 Cip1基因; 促进p21 Cip1蛋白的活性或表达的化合物; 和含有这些分子的药物组合物。 还提供了筛选参与滑膜组织异常增殖，滑膜组织炎症和/或靶向p21蛋白质Cip1蛋白的滑膜组织中炎性细胞因子表达的化合物的方法。 可通过促进p21蛋白质的表达或功能来预防或治疗类风湿性关节炎和与滑膜组织炎症相关的其它疾病。

公开(公告)号：US08697050B2

公开(公告)日：2014-04-15

申请号：US12791577

申请日：2010-06-01

申请人： Nobuyuki Miyasaka ,  Hitoshi Kohsaka

发明人： Nobuyuki Miyasaka ,  Hitoshi Kohsaka

IPC分类号： A01N63/00 ,  A01N65/00

CPC分类号： A61K38/005 ,  A61K38/1709 ,  A61K48/00 ,  C12Q1/6897 ,  G01N33/68 ,  G01N33/6863 ,  G01N2333/4739 ,  G01N2500/00 ,  G01N2500/04

摘要： The onset ratios and pathological conditions of collagen-induced arthritis and adjuvant arthritis in model mice and rats, respectively, were successfully ameliorated by topically expressing the cyclin-dependent kinase inhibitors p161NK4a and p21Cip1 in articular tissues using adenoviral vectors. In the synovial cells of CDKI-transduced mice, expression of inflammatory cytokines was inhibited. Described are the use of the p21Cip1 protein for inhibiting abnormal proliferation of synovial tissues, inflammation in synovial tissues and/or expression of inflammatory cytokines in synovial tissues; the p21Cip1 gene; compounds promoting the activity or expression of the p21Cip1 protein; and pharmaceutical compositions containing these molecules. Also provided are method of screening for compounds participating in the abnormal proliferation of synovial tissues, inflammation in synovial tissues and/or the expression of inflammatory cytokines in synovial tissues targeting the p21Clip1 protein. Rheumatoid arthritis and other disorders associated with inflammation of the synovial tissue can be prevented or treated by promoting expression or function of p21Cip1 protein.

摘要翻译： 通过使用腺病毒载体在关节组织中局部表达细胞周期蛋白依赖性激酶抑制剂p161NK4a和p21Cip1，分别模拟小鼠和大鼠中胶原诱导的关节炎和辅助性关节炎的发病率和病理状况得到了改善。 在CDKI转导小鼠的滑膜细胞中，炎性细胞因子的表达被抑制。 描述了p21Cip1蛋白用于抑制滑膜组织的异常增生，滑液组织中的炎症和/或滑膜组织中炎症细胞因子的表达; p21Cip1基因; 促进p21Cip1蛋白活性或表达的化合物; 和含有这些分子的药物组合物。 还提供了筛选参与滑膜组织的异常增殖，滑膜组织炎症和/或靶向p21Clip1蛋白的滑膜组织中炎性细胞因子表达的化合物的方法。 可通过促进p21Cip1蛋白的表达或功能来预防或治疗类风湿性关节炎和与滑膜组织炎症相关的其它疾病。

公开(公告)号：US20100310508A1

公开(公告)日：2010-12-09

申请号：US12791577

申请日：2010-06-01

申请人： Nobuyuki Miyasaka ,  Hitoshi Kohsaka

发明人： Nobuyuki Miyasaka ,  Hitoshi Kohsaka

IPC分类号： A61K38/20 ,  A61K31/7088 ,  A61K38/02 ,  A61P29/00 ,  A61P19/02 ,  A61K38/18 ,  A61K31/22 ,  A61K31/16 ,  A61K31/23 ,  A61K31/593 ,  A61K31/203 ,  A61K31/4412 ,  A61K31/35 ,  A61K36/185

CPC分类号： A61K38/005 ,  A61K38/1709 ,  A61K48/00 ,  C12Q1/6897 ,  G01N33/68 ,  G01N33/6863 ,  G01N2333/4739 ,  G01N2500/00 ,  G01N2500/04

摘要： The onset ratios and pathological conditions of collagen-induced arthritis and adjuvant arthritis in model mice and rats, respectively, were successfully ameliorated by topically expressing the cyclin-dependent kinase inhibitors p161nk4a and p21Cip1 in articular tissues using adenoviral vectors. In the synovial cells of CDKI-transduced mice, expression of inflammatory cytokines was inhibited. Described are the use of the p21Cip1 protein for inhibiting abnormal proliferation of synovial tissues, inflammation in synovial tissues and/or expression of inflammatory cytokines in synovial tissues; the p21Cip1 gene; compounds promoting the activity or expression of the p21Cip1 protein; and pharmaceutical compositions containing these molecules. Also provided are method of screening for compounds participating in the abnormal proliferation of synovial tissues, inflammation in synovial tissues and/or the expression of inflammatory cytokines in synovial tissues targeting the p21Clip1 protein. Rheumatoid arthritis and other disorders associated with inflammation of the synovial tissue can be prevented or treated by promoting expression or function of p21Cip1 protein.

摘要翻译： 通过使用腺病毒载体在关节组织中局部表达细胞周期蛋白依赖性激酶抑制剂p161nk4a和p21Cip1，分别模拟小鼠和大鼠中胶原诱导的关节炎和辅助性关节炎的发病率和病理状况得到改善。 在CDKI转导小鼠的滑膜细胞中，炎性细胞因子的表达被抑制。 描述了p21Cip1蛋白用于抑制滑膜组织的异常增生，滑液组织中的炎症和/或滑膜组织中炎症细胞因子的表达; p21Cip1基因; 促进p21Cip1蛋白活性或表达的化合物; 和含有这些分子的药物组合物。 还提供了筛选参与滑膜组织的异常增殖，滑膜组织炎症和/或靶向p21Clip1蛋白的滑膜组织中炎性细胞因子表达的化合物的方法。 可通过促进p21Cip1蛋白的表达或功能来预防或治疗类风湿性关节炎和与滑膜组织炎症相关的其它疾病。