Compositions and methods for the early diagnosis of ovarian cancer

Abstract

The disclosed nucleic acid primer sets, used in combination with quantitative amplification (PCR) of tissue cDNA, can indicate the presence of specific proteases in a tissue sample. The detected proteases are themselves specifically overexpressed in certain cancers, and their presence may serve for early detection of associated ovarian and other malignancies, and for the design of interactive therapies for cancer treatment.

What is claimed is:  
     
         1 . A method of diagnosing cancer in an individual, comprising the steps of: 
 (a) obtaining a biological sample from an individual; and    (b) detecting PUMP-1 in said sample, wherein the presence of PUMP-1 in said sample is indicative of the presence of cancer in said individual, wherein the absence of PUMP-1 in said sample is indicative of the absence of cancer in said individual.    
     
     
         2 . The method of  claim 1 , wherein said biological sample is selected from the group consisting of blood, urine, saliva, tears, interstitial fluid, ascites fluid, tumor tissue biopsy and circulating tumor cells.  
     
     
         3 . The method of  claim 1 , wherein said detection of said PUMP-1 is by means selected from the group consisting of Northern blot, Western blot, PCR, dot blot, ELIZA sandwich assay, radioimmunoassay, DNA array chips and flow cytometry.  
     
     
         4 . The method of  claim 1 , wherein said cancer is selected from the group consisting of ovarian, breast, lung, colon, prostate and others in which PUMP-1 is overexpressed.  
     
     
         5 . A method for detecting malignant hyperplasia in a biological sample, comprising the steps of: 
 (a) isolating mRNA from said sample; and    (b) detecting PUMP-1 mRNA in said sample, wherein the presence of said PUMP-1 mRNA in said sample is indicative of the presence of malignant hyperplasia, wherein the absence of said PUMP-1 mRNA in said sample is indicative of the absence of malignant hyperplasia.    
     
     
         6 . The method of  claim 5 , further comprising the step of: 
 comparing said PUMP-1 mRNA to reference information, wherein said comparison provides a diagnosis of said malignant hyperplasia.    
     
     
         7 . The method of  claim 5  further comprising the step of: 
 comparing said PUMP-1 mRNA to reference information, wherein said comparison determines a treatment of said malignant hyperplasia.  
 
     
     
         8 . The method of  claim 5 , wherein said detection of said PUMP-1 mRNA is by PCR amplification.  
     
     
         9 . The method of  claim 8 , wherein said PCR amplification uses primers selected from the group consisting of SEQ ID No. 8 and SEQ ID 9.  
     
     
         10 . The method of  claim 5 , wherein said biological sample is selected from the group consisting of blood, urine, saliva, tears, interstitial fluid, ascites fluid, tumor tissue biopsy and circulating tumor cells.  
     
     
         11 . A method for detecting malignant hyperplasia in a biological sample, comprising the steps of: 
 (a) isolating protein from said sample; and    (b) detecting PUMP-1 protein in said sample, wherein the presence of said PUMP-1 protein in said sample is indicative of the presence of malignant hyperplasia, wherein the absence of said PUMP-1 protein in said sample is indicative of the absence of malignant hyperplasia.    
     
     
         12 . The method of  claim 11 , further comprising the step of: 
 comparing said PUMP-1 protein to reference information, wherein said comparison provides a diagnosis of said malignant hyperplasia.    
     
     
         13 . The method of  claim 11 , further comprising the step of: 
 comparing said PUMP-1 protein to reference information, wherein said comparison determines a treatment of said malignant hyperplasia.    
     
     
         14 . The method of  claim 11 , wherein said detection is by immunoaffinity to an antibody, wherein said antibody is specific for PUMP-1.  
     
     
         15 . The method of  claim 11 , wherein said biological sample is selected from the group consisting of blood, urine, saliva, tears, interstitial fluid, ascites fluid, tumor tissue biopsy and circulating tumor cells.  
     
     
         16 . A method of inhibiting expression of endogenous PUMP-1 in a cell, comprising the step of: 
 introducing a vector into a cell, wherein said vector comprises a PUMP-1 gene in opposite orientation operably linked to elements necessary for expression, wherein expression of said vector in said cell produces PUMP-1 antisense mRNA, wherein said PUMP-1 antisense mRNA hybridizes to endogenous PUMP-1 mRNA, thereby inhibiting expression of endogenous PUMP-1 in said cell.    
     
     
         17 . A method of inhibiting PUMP-1 protein in a cell, comprising the step of: 
 introducing an antibody into a cell, wherein said antibody is specific for a PUMP-1 protein or a fragment thereof, wherein binding of said antibody to said PUMP-1 protein inhibits said PUMP-1 protein.    
     
     
         18 . A method of targeted therapy to an individual, comprising the step of: 
 administering a compound to an individual, wherein said compound has a targeting moiety and a therapeutic moiety, wherein said targeting moiety is specific for PUMP-1.    
     
     
         19 . The method of  claim 18 , wherein said targeting moiety is selected from the group consisting of an antibody specific for PUMP-1 and a ligand or ligand binding domain that binds PUMP-1.  
     
     
         20 . The method of  claim 18 , wherein said therapeutic moiety is selected from the group consisting of a radioisotope, a toxin, a chemotherapeutic agent, an immune stimulant and a cytotoxic agent.  
     
     
         21 . The method of  claim 18 , wherein said individual suffers from a disease selected from the group consisting of ovarian cancer, lung cancer, prostate cancer, colon cancer and other cancers in which PUMP-1 is overexpressed.  
     
     
         22 . A method of vaccinating an individual against PUMP-1, comprising the step of: 
 inoculating an individual with a PUMP-1 protein or fragment thereof, wherein said PUMP-1 protein or fragment thereof lack PUMP-1 propane activity, wherein said inoculation with said PUMP-1 protein or fragment thereof elicits an immune response in said individual, thereby vaccinating said individual against PUMP-1.    
     
     
         23 . The method of  claim 22 , wherein said individual has cancer, is suspected of having cancer or is at risk of getting cancer.  
     
     
         24 . The method of  claim 22 , wherein said PUMP-1 fragment is selected from the group consisting of a 9-residue fragment up to a 20-residue fragment.  
     
     
         25 . The method of  claim 24 , wherein said 9-residue fragment is selected from the group consisting of SEQ ID Nos. 30, 31, 32, 33, 50, 70, 110, 111, 150, 151 and 152.  
     
     
         26 . A method of producing immune-activated cells directed toward PUMP-1, comprising the steps of: 
 exposing dendritic cells to a PUMP-1 protein or fragment thereof, wherein said PUMP-1 protein or fragment thereof lacks PUMP-1 protease activity, wherein said exposure to said PUMP-1 protein or fragment thereof activates said dendritic cells, thereby producing immune-activated cells directed toward PUMP-1.    
     
     
         27 . The method of  claim 26 , wherein said immune-activated cells are selected from the group consisting of B-cells, T-cells and dendrites.  
     
     
         28 . The method of  claim 26 , wherein said PUMP-1 fragment is selected from the group consisting of a 9-residue fragment up to a 20-residue fragment.  
     
     
         29 . The method of  claim 28 , wherein said 9-residue fragment is selected from the group consisting of SEQ ID Nos. 30, 31, 32, 33, 50, 70, 110, 111, 150, 151 and 152.  
     
     
         30 . The method of  claim 26 , wherein said dendritic cells are isolated from an individual prior to said exposure, wherein said activated dendritic cells are reintroduced into said individual subsequent to said exposure.  
     
     
         31 . The method of  claim 30 , wherein said individual has cancer, is suspected of having cancer or is at risk of getting cancer.  
     
     
         32 . An immunogenic composition, comprising an immunogenic fragment of a PUMP-1 protein and an appropriate adjuvant.  
     
     
         33 . The immunogenic composition of  claim 32 , wherein said fragment is selected from the group consisting of a 9-residue fragment up to a 20-residue fragment.  
     
     
         34 . The immunogenic composition of  claim 33 , wherein said 9-residue fragment is selected from the group consisting of SEQ ID Nos. 30, 31, 32, 33, 50, 70, 110, 111, 150, 151 and 152.  
     
     
         35 . An oligonucleotide having a sequence complementary to SEQ ID No. 29.  
     
     
         36 . A composition comprising the oligonucleotide of  claim 35  and a physiologically acceptable carrier.  
     
     
         37 . A method of treating a neoplastic state in an individual in need of such treatment, comprising the step of: administering to said individual an effective dose of the oligonucleotide of  claim 35 .  
     
     
         38 . The method of  claim 37 , wherein said neoplastic state is selected from the group consisting of ovarian cancer, breast cancer, lung cancer, colon cancer, prostate cancer and other cancers in which PUMP-1 is overexpressed.  
     
     
         39 . A method of screening for compounds that inhibit PUMP-1 activity, comprising the steps of: 
 contacting a sample with a compound, wherein said sample comprises PUMP-1 protein; and    assaying for PUMP-1 protease activity, wherein a decrease in said PUMP-1 protease activity in the presence of said compound relative to PUMP-1 protease activity in the absence of said compound indicatives the compound inhibits PUMP-1 activity.    
     
     
         40 . A method for detecting ovarian malignant hyperplasia in a biological sample, comprising the steps of: 
 (a) isolating the proteases or protease mRNA present in said biological sample; and    (b) detecting specific proteases or protease mRNA present in said biological sample, wherein said proteases are selected from the group consisting of hepsin, protease M, complement factor B, SCCE, other serine proteases indicated in lanes 2 and 4 of FIG. 1, cathepsin L and PUMP-1.    
     
     
         41 . The method of  claim 40 , further comprising the step of: comparing the specific proteases or protease mRNA detected to reference information, wherein said comparison provides a diagnoses of said malignant hyperplasia.  
     
     
         42 . The method of  claim 40 , further comprising the step of: comparing the specific proteases or protease mRNA detected to reference information, wherein said comparison determines a treatment of said malignant hyperplasia.  
     
     
         43 . The method of  claim 40 , wherein said protease mRNA is detected by amplification of total mRNA.  
     
     
         44 . The method of  claim 40 , wherein said protease is detected with an antibody.  
     
     
         45 . The method of  claim 40 , wherein said biological sample is selected from the group consisting of blood, urine, saliva, tears, interstitial fluid, ascites fluid, tumor tissue biopsy and circulating tumor cells.