Antibodies against actin-binding protein girdin and methods of making and using the same

Abstract

Antibodies are provided that specifically bind to Acting Binding Protein Girdin (Akt Phosphorylation Enhancer), a new substrate of serine/threonine kinase. Additionally, methods of making hybridomas for producing said antibodies and methods of detecting girdin using said antibodies are provided herein. The antibodies and methods are useful for detecting girdin in tissue samples, such as tissue samples from subjects having or suspected of having a disorder in which any of cell motility, cell movement, and angiogenesis is involved.

What is claimed is: 
     
         1 . A monoclonal or polyclonal antibody specific to a protein or to a partial peptide thereof containing an amino acid sequence identical with the amino acid sequence represented by SEQ ID NO: 2. 
     
     
         2 . The monoclonal or polyclonal antibody of  claim 1 , wherein said antibody is raised against a carboxyl-terminal fragment of SEQ ID NO: 2. 
     
     
         3 . A method of detecting girdin in a tissue sample, said method comprising immunostaining the tissue sample with the monoclonal or polyclonal antibody according to  claim 1 . 
     
     
         4 . The method of  claim 3 , wherein said tissue sample is selected from the group consisting of primary, metastatic or recurrent breast cancer, prostate cancer, pancreatic cancer, stomach cancer, pulmonary cancer, large intestine cancer (colon cancer, rectal cancer, anus cancer), esophageal cancer, duodenal cancer, head and neck cancer (tongue cancer, pharyngeal cancer, larynx cancer), cerebral tumor, neurilemmoma, non-small cell lung cancer, lung small cell cancer, hepatic cancer, kidney cancer, bile duct cancer, uterus cancer (uterine body cancer, uterine cervix cancer), ovary cancer, bladder cancer, skin cancer, vascular neoplasm, malignant lymphoma, malignant melanoma, thyroid cancer, bone tumor, vascular neoplasm, angiofibroma, retinal sarcoma, penile cancer, pediatric solid cancer, Kaposi's sarcoma, Kaposi's sarcoma resulting from AIDS, maxillary sinus tumor, fibrous histiocytoma, leiomyosarcoma, rhabdomyosarcoma, liposarcoma, uterine myoma, osteoblastoma, bone sarcoma, chondrosarcoma, carcinomatous mesothelioma, tumor such as leukemia, Hodgkin's disease 
     
     
         5 . The method of  claim 3 , wherein said tissue sample is a human breast cancer or a human cervical cancer sample. 
     
     
         6 . A method of detecting girdin in a tissue sample, said method comprising immunostaining the tissue sample with the monoclonal or polyclonal antibody according to  claim 2 . 
     
     
         7 . The method of  claim 6 , wherein said tissue sample is selected from the group consisting of primary, metastatic or recurrent breast cancer, prostate cancer, pancreatic cancer, stomach cancer, pulmonary cancer, large intestine cancer (colon cancer, rectal cancer, anus cancer), esophageal cancer, duodenal cancer, head and neck cancer (tongue cancer, pharyngeal cancer, larynx cancer), cerebral tumor, neurilemmoma, non-small cell lung cancer, lung small cell cancer, hepatic cancer, kidney cancer, bile duct cancer, uterus cancer (uterine body cancer, uterine cervix cancer), ovary cancer, bladder cancer, skin cancer, vascular neoplasm, malignant lymphoma, malignant melanoma, thyroid cancer, bone tumor, vascular neoplasm, angiofibroma, retinal sarcoma, penile cancer, pediatric solid cancer, Kaposi's sarcoma, Kaposi's sarcoma resulting from AIDS, maxillary sinus tumor, fibrous histiocytoma, leiomyosarcoma, rhabdomyosarcoma, liposarcoma, uterine myoma, osteoblastoma, bone sarcoma, chondrosarcoma, carcinomatous mesothelioma, tumor such as leukemia, Hodgkin's disease 
     
     
         8 . The method of  claim 6 , wherein said tissue sample is a human breast cancer or a human cervical cancer sample. 
     
     
         9 . A hybridoma producing a monoclonal antibody specific to a protein or to a partial peptide thereof containing an amino acid sequence identical with the amino acid sequence represented by SEQ ID NO: 2. 
     
     
         10 . A method of making the hybridoma according to  claim 9 , said method comprising:
 immunizing a warm-blooded animal with an antigenic substance;   collecting spleen or lymph node from animals showing antibody titer an appropriate number of days after the final immunization; and   fusing antibody producing cells contained in the spleen or lymph node with a myeloma to prepare monoclonal antibody producing hybridoma.   
     
     
         11 . The method of  claim 10 , wherein said antigenic substance comprises a compound material of an antigen with immunogenicity and a carrier protein. 
     
     
         12 . The method of  claim 11 , wherein said antigen with immunogenicity is a carboxyl-terminal fragment of girdin. 
     
     
         13 . A method of making a monoclonal antibody specific to a protein or to a partial peptide thereof containing an amino acid sequence identical with the amino acid sequence represented by SEQ ID NO: 2, said method comprising:
 making a hybridoma according to the method of  claim 10 ; and   isolating and purifying the monoclonal antibody.   
     
     
         14 . The method of  claim 13 , wherein said antigenic substance comprises a compound material of an antigen with immunogenicity and a carrier protein. 
     
     
         15 . The method of  claim 14 , wherein said antigen with immunogenicity is a carboxyl-terminal fragment of girdin. 
     
     
         16 . An isolated monoclonal antibody obtained by a method according to  claim 12 . 
     
     
         17 . The isolated monoclonal antibody of  claim 16 , wherein said antigenic substance comprises a compound material of an antigen with immunogenicity and a carrier protein. 
     
     
         18 . The isolated monoclonal antibody of  claim 17 , wherein said antigen with immunogenicity is a carboxyl-terminal fragment of girdin. 
     
     
         19 . A method of detecting girdin in a tissue sample, said method comprising immunostaining the tissue sample with the monoclonal antibody according to  claim 16 . 
     
     
         20 . The method of  claim 19 , wherein said tissue sample is selected from the group consisting of primary, metastatic or recurrent breast cancer, prostate cancer, pancreatic cancer, stomach cancer, pulmonary cancer, large intestine cancer (colon cancer, rectal cancer, anus cancer), esophageal cancer, duodenal cancer, head and neck cancer (tongue cancer, pharyngeal cancer, larynx cancer), cerebral tumor, neurilemmoma, non-small cell lung cancer, lung small cell cancer, hepatic cancer, kidney cancer, bile duct cancer, uterus cancer (uterine body cancer, uterine cervix cancer), ovary cancer, bladder cancer, skin cancer, vascular neoplasm, malignant lymphoma, malignant melanoma, thyroid cancer, bone tumor, vascular neoplasm, angiofibroma, retinal sarcoma, penile cancer, pediatric solid cancer, Kaposi's sarcoma, Kaposi's sarcoma resulting from AIDS, maxillary sinus tumor, fibrous histiocytoma, leiomyosarcoma, rhabdomyosarcoma, liposarcoma, uterine myoma, osteoblastoma, bone sarcoma, chondrosarcoma, carcinomatous mesothelioma, tumor such as leukemia, Hodgkin's disease 
     
     
         21 . The method of  claim 19 , wherein said tissue sample is a human breast cancer or a human cervical cancer sample. 
     
     
         22 . A method of detecting girdin in a tissue sample, said method comprising immunostaining the tissue sample with the monoclonal antibody according to  claim 17 . 
     
     
         23 . The method of  claim 22 , wherein said tissue sample is selected from the group consisting of primary, metastatic or recurrent breast cancer, prostate cancer, pancreatic cancer, stomach cancer, pulmonary cancer, large intestine cancer (colon cancer, rectal cancer, anus cancer), esophageal cancer, duodenal cancer, head and neck cancer (tongue cancer, pharyngeal cancer, larynx cancer), cerebral tumor, neurilemmoma, non-small cell lung cancer, lung small cell cancer, hepatic cancer, kidney cancer, bile duct cancer, uterus cancer (uterine body cancer, uterine cervix cancer), ovary cancer, bladder cancer, skin cancer, vascular neoplasm, malignant lymphoma, malignant melanoma, thyroid cancer, bone tumor, vascular neoplasm, angiofibroma, retinal sarcoma, penile cancer, pediatric solid cancer, Kaposi's sarcoma, Kaposi's sarcoma resulting from AIDS, maxillary sinus tumor, fibrous histiocytoma, leiomyosarcoma, rhabdomyosarcoma, liposarcoma, uterine myoma, osteoblastoma, bone sarcoma, chondrosarcoma, carcinomatous mesothelioma, tumor such as leukemia, Hodgkin's disease 
     
     
         24 . The method of  claim 22 , wherein said tissue sample is a human breast cancer or a human cervical cancer sample. 
     
     
         25 . A method of detecting girdin in a tissue sample, said method comprising immunostaining the tissue sample with the monoclonal antibody according to  claim 18 . 
     
     
         26 . The method of  claim 25 , wherein said tissue sample is selected from the group consisting of primary, metastatic or recurrent breast cancer, prostate cancer, pancreatic cancer, stomach cancer, pulmonary cancer, large intestine cancer (colon cancer, rectal cancer, anus cancer), esophageal cancer, duodenal cancer, head and neck cancer (tongue cancer, pharyngeal cancer, larynx cancer), cerebral tumor, neurilemmoma, non-small cell lung cancer, lung small cell cancer, hepatic cancer, kidney cancer, bile duct cancer, uterus cancer (uterine body cancer, uterine cervix cancer), ovary cancer, bladder cancer, skin cancer, vascular neoplasm, malignant lymphoma, malignant melanoma, thyroid cancer, bone tumor, vascular neoplasm, angiofibroma, retinal sarcoma, penile cancer, pediatric solid cancer, Kaposi's sarcoma, Kaposi's sarcoma resulting from AIDS, maxillary sinus tumor, fibrous histiocytoma, leiomyosarcoma, rhabdomyosarcoma, liposarcoma, uterine myoma, osteoblastoma, bone sarcoma, chondrosarcoma, carcinomatous mesothelioma, tumor such as leukemia, Hodgkin's disease 
     
     
         27 . The method of  claim 25 , wherein said tissue sample is a human breast cancer or a human cervical cancer sample.