Development of DNA probes and immunological reagents specific for cell surface-expressed molecules and transformation-associated genes

Abstract

This invention provides a method for preparing a hybridoma cell line which produces an antibody capable of specifically binding to a cell surface-expressed protein which expresses on the surface of one cell type but not the other. This invention also provides a method for preparing a hybridoma cell line which produces an antibody capable of specifically binding to a cell surface-expressed protein. This invention provides a method to prepare a hybridoma cell line which specifically recognizes and binds to a tumor associated antigen associated with a neoplastic, human cell. This invention also provides a method of preparing DNA encoding a cell surface antigen associated with a neoplastic, human cell. This invention further provides an isolated mammalian nucleic acid molecule having the sequence of Prostate Carcinoma Tumor Antigen Gene-1. This invention also provides an isolated mammalian nucleic acid molecule having the sequence of Prostate Tumor Inducing Gene-1. This invention provides an isolated mammalian nucleic acid molecule having the sequence of Prostate Tumor Inducing Gene-2. Finally, this invention provides an isolated mammalian nucleic acid molecule having the sequence of Prostate Tumor Inducing Gene-1.

1 . A method for preparing a hybridoma cell line which produces an antibody which specifically recognizes and binds to a cell surface antigen associated with a neoplastic, human cell which comprises: 
 (a) cotransfecting an established non-human, non-tumorigenic cell line with DNA isolated from a neoplastic, human cell and DNA encoding a selectable or identifiable trait;    (b) selecting transfected cells which express the selectable or identifiable trait;    (c) recovering the transfected cells so selected;    (d) injecting the transfected cells so recovered into a suitable first murine host;    (e) maintaining the resulting first murine host for a period of time effective to induce the injected transfected cells to form a tumor in the first murine host;    (f) isolating the resulting tumor from the first murine host;    (g) obtaining tumor cells from the tumor so isolated;    (h) coating the tumor cells so obtained with an antiserum generated against the established non-human, non-tumorigenic cell line;    (i) injecting the antiserum-coated cells into suitable second hosts;    (j) screening the resulting second hosts to identify hosts which produce serum reactive with the neoplastic, human cell;    (k) removing spleens from the second hosts so identified;    (l) preparing from the spleens so removed hybridomas; and    (m) recovering therefrom a hybridoma cell line which produces an antibody which specifically recognizes and binds to the cell surface antigen.    
     
     
         2 . A method of  claim 1 , wherein the established non-human, non-tumorigenic cell line is the CREF-Trans 6 cell line (ATCC Accession No. CRL 10584).  
     
     
         3 . A method of  claim 1 , wherein the neoplastic, human cell is a benign cell.  
     
     
         4 . A method of  claim 1 , wherein the neoplastic, human cell is metastatic cell.  
     
     
         5 . A method of  claim 1 , wherein the neoplastic, human cell is a human prostatic carcinoma cell derived from cell line LNCaP.  
     
     
         6 . A method of  claim 1 , wherein the neoplastic, human cell is a human breast carcinoma cell derived from cell line T47D.  
     
     
         7 . A method of  claim 1 , wherein the neoplastic, human cell is a human colorectal carcinoma cell derived from cell line SW480.  
     
     
         8 . A method of  claim 1 , wherein the neoplastic, human cell is a human glioblastoma multiform (stage 1V astrocytoma) cell derived from cell line GBM-18.  
     
     
         9 . A method of  claim 1 , wherein the neoplastic, human cell is a human glioblastoma multiform (stage 1V astrocytoma) cell derived from a primary tumor.  
     
     
         10 . A method of  claim 1 , wherein the DNA encoding the selectable or identifiable trait is plasmid DNA encoding resistance to an antibiotic.  
     
     
         11 . A method of  claim 10 , wherein the plasmid DNA comprises pSV2-Neo.  
     
     
         12 . A method of  claim 11 , wherein the antibiotic is G418.  
     
     
         13 . A method of  claim 1 , wherein the suitable second host is a murine host.  
     
     
         14 . A method of  claim 1 , wherein the suitable second host is a non-human primate host.  
     
     
         15 . A method of  claim 1 , wherein the cell surface antigen is a tumor associated antigen.  
     
     
         16 . A method of  claim 1 , wherein the cell surface antigen is a growth factor receptor.  
     
     
         17 . A method of  claim 1 , wherein the cell surface antigen is a viral-encoded, surface-expressed antigen.  
     
     
         18 . A method of  claim 1 , wherein the cell surface antigen is encoded by an oncogene product.  
     
     
         19 . A method of  claim 1 , wherein the cell surface antigen is a surface epitope.  
     
     
         20 . A method of  claim 1 , wherein the cell surface antigen is a membrane protein which mediates classical multi-drug resistance.  
     
     
         21 . A method of  claim 1 , wherein the cell surface antigen is a membrane protein which mediates atypical multi-drug resistance.  
     
     
         22 . A method of  claim 1 , wherein the cell surface antigen is an antigen which mediates a tumorigenic phenotype.  
     
     
         23 . A method of  claim 1 , wherein the cell surface antigen is an antigen which mediates a metastatic phenotype.  
     
     
         24 . A method of  claim 1 , wherein the cell surface antigen is an antigen which suppresses a tumorigenic phenotype.  
     
     
         25 . A method of  claim 1 , wherein the cell surface antigen is an antigen which suppresses a metastatic phenotype.  
     
     
         26 . A method of  claim 1 , wherein the cell surface antigen is an antigen which is recognized by a specific immunological effector cell.  
     
     
         27 . A method of  claim 1 , wherein the specific immunological effector cell is a T-cell.  
     
     
         28 . A method of  claim 1 , wherein the cell surface antigen is an antigen which is recognized by a non-specific immunological effector cell.  
     
     
         29 . A method of  claim 28 , wherein the non-specific immunological effector cell is a macrophage cell.  
     
     
         30 . A method of  claim 28 , wherein the non-specific immunological effector cell is a natural killer cell.  
     
     
         31 - 101 . (canceled)  
     
     
         102 . A method of preparing DNA encoding a cell surface antigen associated with a neoplastic, human cell, the method comprising: 
 (a) cotransfecting a cell line with DNA isolated from a neoplastic human cell and DNA encoding a selectable trait;    (b) selecting transfected cells expressing the selectable trait;    (c) recovering the selected cells;    (d) injecting the recovered cells into a suitable non-human host;    (e) maintaining the host for a period of time effective to allow the injected transfected cells to form a tumor in the host;    (f) isolating the tumor from the host;    (g) isolating tumor cells from the tumor;    (h) recovering DNA encoding the cell surface antigen associated with the neoplastic human cell from the isolated tumor cells.