US2003113743A1PendingUtilityA1

Method for detection of micrometastatic prostate cancer

52
Assignee: BAYLOR COLLEGE OF MEDICINE HYBPriority: Nov 14, 1996Filed: Apr 22, 2002Published: Jun 19, 2003
Est. expiryNov 14, 2016(expired)· nominal 20-yr term from priority
Inventors:Kevin SlawinDonald J. TindallCharles Young
C12N 9/6445C07K 16/40C12Q 1/6886C12Q 2600/112
52
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Claims

Abstract

Prostate cancer is detected by determining the presence of hK2 RNA in a physiological sample.

Claims

exact text as granted — not AI-modified
What is claimed is:  
     
         1 . A diagnostic method for detecting hK2 DNA comprising: 
 (a) contacting an amount of DNA obtained by reverse transcription of RNA from a human physiological sample which comprises cells suspected of containing hK2 RNA with an amount of at least two oligonucleotides under conditions effective to amplify the DNA by a polymerase chain reaction so as to yield an amount of amplified hK2 DNA, wherein the conditions are effective to amplify the amount of DNA obtained by reverse transcription of RNA from at least one cell containing hK2 in a sample which comprises at least about 10 7  to about 10 9  cells; and    (b) detecting the presence of the amplified hK2 DNA.    
     
     
         2 . A method for detecting metastatic prostate cancer in a human, comprising: 
 (a) contacting an amount of DNA obtained by reverse transcription of RNA from a human physiological sample which comprises cells suspected of containing hK2 RNA with an amount of at least two oligonucleotides under conditions effective to amplify the DNA by a polymerase chain reaction so as to yield an amount of amplified hK2 DNA, wherein the conditions are effective to amplify the amount of DNA obtained by reverse transcription of RNA from at least one cell containing hK2 in a sample which comprises at least about 10 7  to about 10 9  cells;    (b) detecting the presence of the amplified hK2 DNA, wherein the presence of hK2 DNA is indicative of the presence of metastatic prostate cancer in said human.    
     
     
         3 . The method of  claim 1  or  2  wherein the physiological sample is a fluid.  
     
     
         4 . The method of  claim 3  wherein the fluid is whole blood.  
     
     
         5 . The method of  claim 1  or  2  wherein the physiological sample is a tissue sample.  
     
     
         6 . The method of  claim 5  wherein the tissue is selected from the group consisting of prostate, prostate capsule, seminal vesicle, bone marrow and lymph node.  
     
     
         7 . The method of  claim 5  wherein the tissue is non-prostate tissue.  
     
     
         8 . The method of  claim 1  or  2  wherein the amplified hK2 DNA is subjected to agarose gel electrophoresis prior to detection.  
     
     
         9 . The method of  claim 1  or  2  further comprising quantifying the amount of amplified hK2 DNA.  
     
     
         10 . The method of  claim 1  or  2  wherein the amplified hK2 DNA is obtained from hK2 RNA that is alternatively spliced.  
     
     
         11 . The method of  claim 1  or  2  further comprising: 
 (c) contacting a second amount of the DNA obtained by reverse transcription of RNA from the human physiological sample with an amount of at least two oligonucleotides under conditions effective to amplify prostate specific antigen (PSA) DNA, but not hK2 DNA, by polymerase chain reaction so as to yield amplified PSA DNA; and  
 (d) detecting the presence of the amplified PSA DNA.  
 
     
     
         12 . The method of  claim 1  or  2  wherein one oligonucleotide comprises SEQ ID NO:27.  
     
     
         13 . The method of  claim 1  or  2  wherein one oligonucleotide comprises SEQ ID NO:28.  
     
     
         14 . A diagnostic method for detecting hK2 RNA comprising: 
 (a) extracting RNA from a physiological sample obtained from a human, wherein the sample comprises cells;    (b) reverse transcribing the extracted RNA to yield DNA;    (c) contacting the DNA with an amount of at least two oligonucleotides under conditions effective to amplify the DNA by polymerase chain reaction so as to yield an amount amplified hK2 DNA, wherein the conditions are effective to amplify the amount of DNA obtained by reverse transcription of RNA from at least one cell containing hK2 in a sample which comprises at least about 10 7  to about 10 9  cells; and    (d) detecting the presence of the amplified hK2 DNA, wherein the presence of the amplified hK2 DNA is indicative of metastatic prostate cancer in the human.    
     
     
         15 . The method of  claim 14  wherein the sample is a physiological fluid sample.  
     
     
         16 . The method of  claim 14  wherein the sample is a tissue sample.  
     
     
         17 . The method of  claim 14  wherein the human has had a radical prostatectomy, and wherein the presence of hK2 DNA is indicative of the presence of persistent prostate cancer in the human.  
     
     
         18 . The method of  claim 14  wherein the amplified hK2 DNA is obtained from hK2 RNA that is alternatively spliced.  
     
     
         19 . The method of  claim 14  wherein one oligonucleotide comprises SEQ ID NO:27.  
     
     
         20 . The method of  claim 14  wherein one oligonucleotide comprises SEQ ID NO:28.  
     
     
         21 . A method for monitoring the progression of prostate cancer, comprising: 
 (a) contacting an amount of DNA obtained by reverse transcription of RNA from a physiological sample obtained from a human afflicted with prostate cancer with an amount of at least two oligonucleotides under conditions effective to amplify the DNA by a polymerase chain reaction so as to yield an amount of amplified hK2 DNA, wherein the physiological sample comprises cells, and wherein the conditions are effective to amplify an amount of DNA obtained by reverse transcription of RNA from at least one cell containing hK2 in a sample which comprises at least about 10 7  to about 10 9  cells;    (b) detecting or determining the amount of the amplified hK2 DNA;    (c) repeating steps (a) and (b) at a point later in time; and    (d) comparing the result of step (b) with the result of step (c), wherein an increase in the amount of hK2 DNA is indicative of the progression of prostate cancer in said human.    
     
     
         22 . A method for pathologically staging prostate cancer, comprising: 
 (a) contacting an amount of DNA obtained by reverse transcription of RNA from a physiological sample obtained from a human afflicted with prostate cancer with an amount of at least two oligonucleotides under conditions effective to amplify the DNA by a polymerase chain reaction so as to yield an amount of amplified hK2 DNA, wherein the physiological sample comprises cells, and wherein the conditions are effective to amplify an amount of DNA obtained by reverse transcription of RNA from at least one cell containing hK2 in a sample which comprises at least about 10 7  to about 10 9  cells; and    (b) detecting or determining the presence or amount of the amplified hK2 DNA, wherein the presence or amount of amplified hK2 DNA is indicative of the pathological stage of the prostate cancer.    
     
     
         23 . The method of  claim 21  or  22  wherein the human is a candidate for radical prostatectomy.  
     
     
         24 . The method of  claim 21  or  22  wherein the sample is a physiological fluid sample.  
     
     
         25 . The method of  claim 21  or  22  wherein the sample is a non-prostate tissue sample.  
     
     
         26 . The method of  claim 21  or  22  wherein the fluid is whole blood.  
     
     
         27 . The method of  claim 21  or  22  wherein one oligonucleotide comprises SEQ ID NO:27.  
     
     
         28 . The method of  claim 21  or  22  wherein one oligonucleotide comprises SEQ ID NO:28.  
     
     
         29 . An oligonucleotide which consists of at least about 7-50 nucleotides, and which has at least about 80% identity to, or has complementarity to, a nucleotide sequence comprising SEQ ID NO:27.  
     
     
         30 . An oligonucleotide which consists of at least about 7-50 nucleotides, and which has at least about 80% identity to, or has complementarity to, a nucleotide sequence comprising SEQ ID NO:28.  
     
     
         31 . A diagnostic method for detecting hK2 DNA comprising: 
 (a) contacting an amount of DNA obtained by reverse transcription of RNA from a human physiological sample which comprises cells suspected of containing hK2 RNA, with an amount of at least two oligonucleotides under conditions effective to amplify the DNA by a polymerase chain reaction so as to yield an amount of amplified hK2 DNA, wherein at least one oligonucleotide is an hK2-specific oligonucleotide; and    (b) detecting the presence of the amplified hK2 DNA.    
     
     
         32 . A method for detecting metastatic prostate cancer in a human, comprising: 
 (a) contacting an amount of DNA obtained by reverse transcription of RNA from a human physiological sample which comprises cells suspected of containing hK2 RNA, with an amount of at least two oligonucleotides under conditions effective to amplify the DNA by a polymerase chain reaction so as to yield an amount of amplified hK2 DNA, wherein at least one oligonucleotide is an hK2-specific oligonucleotide; and    (b) detecting the presence of the amplified hK2 DNA, wherein the presence of hK2 DNA is indicative of the presence of metastatic prostate cancer in said human.    
     
     
         33 . The method of  claim 31  or  32  wherein the physiological sample is a tissue sample.  
     
     
         34 . The method of  claim 33  wherein the tissue is selected from the group consisting of prostate, prostate capsule, seminal vesicle, bone marrow and lymph node.  
     
     
         35 . The method of  claim 33  wherein the tissue is non-prostate tissue.  
     
     
         36 . The method of  claim 31  or  32  wherein the physiological sample is a fluid.  
     
     
         37 . The method of  claim 36  wherein the fluid is selected from the group consisting of whole blood, blood serum and seminal fluid.  
     
     
         38 . The method of  claim 37  wherein the fluid is whole blood.  
     
     
         39 . The method of  claim 31  or  32  wherein the amplified hK2 DNA is subjected to agarose gel electrophoresis prior to detection.  
     
     
         40 . The method of  claim 31  or  32  further comprising quantifying the amount of amplified hK2 DNA.  
     
     
         41 . A diagnostic method for detecting hK2 RNA comprising: 
 (a) extracting RNA from a physiological sample obtained from a human;    (b) reverse transcribing the extracted RNA to yield DNA;    (c) contacting the DNA with an amount of at least two oligonucleotides under conditions effective to amplify the DNA by polymerase chain reaction so as to yield an amount amplified hK2 DNA, wherein at least one oligonucleotide is an hK2-specific oligonucleotide; and    (d) detecting the presence of the amplified hK2 DNA, wherein the presence of the amplified hK2 DNA is indicative of metastatic prostate cancer in the human.    
     
     
         42 . The method of  claim 41  wherein the sample is a tissue sample.  
     
     
         43 . The method of  claim 41  wherein the sample is a physiological fluid sample.  
     
     
         44 . The method of  claim 41  wherein the human has had a radical prostatectomy, and wherein the presence of hK2 DNA is indicative of the presence of persistent prostate cancer in the human.  
     
     
         45 . A method for monitoring the progression of prostate cancer, comprising: 
 (a) contacting an amount of DNA obtained by reverse transcription of RNA from a physiological sample obtained from a human afflicted with prostate cancer with an amount of at least two oligonucleotides under conditions effective to amplify the DNA by a polymerase chain reaction so as to yield an amount of amplified hK2 DNA, wherein at least one oligonucleotide is an hK2-specific oligonucleotide;    (b) detecting or determining the amount of the amplified hK2 DNA;    (c) repeating steps (a) and (b) at a point later in time; and    (d) comparing the result of step (b) with the result of step (c), wherein an increase in the amount of hK2 DNA is indicative of the progression of prostate cancer in said human.    
     
     
         46 . A method for pathologically staging prostate cancer, comprising: 
 (a) contacting an amount of DNA obtained by reverse transcription of RNA from a physiological sample obtained from a human afflicted with prostate cancer with an amount of at least two oligonucleotides under conditions effective to amplify the DNA by a polymerase chain reaction so as to yield an amount of amplified hK2 DNA, wherein at least one oligonucleotide is an hK2-specific oligonucleotide; and    (b) detecting or determining the presence or amount of the amplified hK2 DNA, wherein the presence or amount of amplified hK2 DNA is indicative of the pathological stage of the prostate cancer.    
     
     
         47 . The method of  claim 45  or  46  wherein the human is a candidate for radical prostatectomy.  
     
     
         48 . The method of  claim 45  where the sample of step (a) is obtained prior to the human undergoing hormone therapy.  
     
     
         49 . The method of  claim 48  wherein the hormone therapy is androgen therapy.  
     
     
         50 . The method of  claim 49  wherein the androgen therapy is androgen provocative therapy.  
     
     
         51 . The method of  claim 45  or  46  wherein the sample is a non-prostate tissue sample.  
     
     
         52 . The method of  claim 45  or  46  wherein the sample is a physiological fluid sample.  
     
     
         53 . The method of  claim 52  wherein the fluid is whole blood.

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