Method for the detection of cancer

Abstract

The present invention relates to a method for the diagnosis and/or the follow up of the evolution of cancer, which includes the analysis and quantification of over expressed and amplified genes in the plasma/serum of cancer patients or persons suspected to harbor cancer. This is achieved by analyzing together the amount of DNA and RNA of certain genes in the plasma/serum of cancer patients that are the reflection of a gene amplification and/or a gene over expression in comparison to healthy controls.

The invention claimed is: 
     
       1. A method for the diagnosis or the follow up of the evolution of cancers which comprises measuring both gene over-expression (RNA) and gene amplification (DNA) of a gene present in the bodily fluids of a patient that is both amplified and over-expressed in cancer cells and comparing to healthy controls. 
     
     
       2. The method according to  claim 1 , wherein RNA and DNA are extracted from a bodily fluid, purified and amplified, and the over-expressed RNA and amplified DNA are analyzed and compared to a house keeping gene. 
     
     
       3. The method according to  claim 2 , wherein the genes analyzed are selected from hTERT, hTR, TEP1, MYCN, MYCC, ErbB2, Her2, Her2/Neu, Her 1, Cyclin A and D1, ABL, SKP2, ETV6 (TELgene), MGC2177, PLAG1, PSMC6P and LYN. 
     
     
       4. The method according to  claim 2 , wherein the nucleic acids are amplified by reverse transcriptase chain reaction (RT-PCR). 
     
     
       5. The method according to  claim 3 , wherein the nucleic acids are amplified by reverse transcriptase chain reaction (RT-PCR). 
     
     
       6. The method according to  claim 1 , wherein the genes analyzed are selected from the group consisting of hTERT, hTR, TEP1, MYCN, MYCC, ErbB2, Her2, Her2/Neu, Her 1, Cyclin A and D1, ABL, SKP2, ETV6 (TELgene), MGC2177, PLAG1, PSMC6P and LYN. 
     
     
       7. The method according to  claim 6 , wherein the nucleic acids are amplified by reverse transcriptase chain reaction (RT-PCR). 
     
     
       8. The method according to  claim 1 , wherein the nucleic acids are amplified by reverse transcriptase chain reaction (RT-PCR). 
     
     
       9. The method according to  claim 1 , wherein the genes analyzed are compared to a reference nucleic acid extract (DNA and RNA) corresponding to the expression (RNA) and quantity (DNA) of a house keeping gene. 
     
     
       10. The method according to  claim 1 , wherein the genes analyzed are compared to a reference RNA corresponding to the expression of a house keeping gene. 
     
     
       11. The method according to  claim 1 , wherein the genes analyzed are compared to a reference DNA corresponding to a housekeeping gene. 
     
     
       12. The method according to  claim 1 , wherein the gene quantification may be estimated in reference to a standard curve obtained with nucleic acids of a cell line. 
     
     
       13. The method according to  claim 1 , wherein the nucleic acids RNA and DNA are analyzed by gel coloration, by radioactive immunological technique (RIA), by enzyme linked immunosorbant test (ELISA) or by a microchip test (gene array), and quantified by any method for nucleic acid quantification. 
     
     
       14. The method according to  claim 1 , wherein the nucleic acids RNA and DNA are quantified by real time RT PCR. 
     
     
       15. The method according to  claim 1 , wherein the RNA and DNA are extracted from the patient and measured simultaneously. 
     
     
       16. A method for measuring both gene over-expression (RNA) and gene amplification (DNA) in a patient suspected of having cancer, wherein the gene is selected from the group consisting of hTERT, hTR, TEP1, MYCN, MYCC, ErbB2, Her2, Her2/Neu, Her 1, Cyclin A, Cyclin D1, ABL, SKP2, ETV6 (TELgene), MGC2177, PLAG1, PSMC6P and LYN, comprising:
 obtaining a sample of a bodily fluid from a patient, and extracting both RNA and DNA simultaneously from said sample,   measuring both gene over-expression (RNA) and gene amplification (DNA) at the same time in said sample, and   comparing said measurement to a healthy control.   
     
     
       17. The method according to  claim 16 , wherein RNA and DNA are extracted from a bodily fluid, purified and amplified, and the over-expressed RNA and amplified DNA are analyzed and compared to a house keeping gene. 
     
     
       18. The method according to  claim 17 , wherein the over-expressed and amplified gene is hTERT. 
     
     
       19. The method according to  claim 18 , wherein the nucleic acids are amplified by reverse transcriptase chain reaction (RT-PCR). 
     
     
       20. The method according to  claim 16 , wherein the genes analyzed are compared to a reference nucleic acid extract (DNA and RNA) corresponding to the expression (RNA) and quantity (DNA) of a house keeping gene. 
     
     
       21. The method according to  claim 16 , wherein the genes analyzed are compared to a reference RNA corresponding to the expression of a house keeping gene. 
     
     
       22. A method for detecting gene amplification of one or more genes that occurs in cancer cells of a subject having cancer, the gene amplification of the one or more genes being indicative of cancer, the method comprising:
 (i) obtaining a blood sample from the subject;   (ii) separating cell-free deoxyribonucleic acid (DNA) from a plasma or serum fraction of the blood sample;   (iii) amplifying DNA of the one or more genes of the subject present in the cell-free DNA separated from the plasma or serum fraction;   (iv) amplifying DNA of a reference gene present in the cell-free DNA separated from the plasma or the serum fraction;   (v) measuring amounts of the amplified DNA of the one or more genes;   (vi) measuring an amount of the amplified DNA of the reference gene, wherein the reference gene is different than the one or more genes;   (vii) determining if the amounts of the amplified DNA of the one or more genes are increased relative to the amount of amplified DNA of the reference gene; and   (viii) detecting the gene amplification in cancer cells of the one or more genes if the amounts of the amplified DNA of the one or more genes are determined to be increased relative to the amount of amplified DNA of the reference gene.   
     
     
       23. The method according to claim 22, wherein the one or more genes are a plurality of genes. 
     
     
       24. The method according to claim 22, wherein the one or more genes include oncogenes. 
     
     
       25. The method according to claim 22, further comprising detecting a tumor-related characteristic in the one or more genes present in the plasma or serum fraction. 
     
     
       26. The method according to claim 22, wherein the separating comprises extracting cell-free DNA from the plasma or serum fraction. 
     
     
       27. The method according to claim 26, further comprising amplifying DNA of the one or more genes of the extracted cell-free DNA. 
     
     
       28. The method according to claim 27, wherein the amplifying comprises using polymerase chain reaction. 
     
     
       29. The method according to claim 22, wherein the one or more genes are selected from the group consisting of hTERT, hTR, TEPI, MYCN, MYCC, ErbB2, Her2, Her2/Neu, Her1, Cyclin A and D1, ABL, SKP2, ETV6 (TELgene), MGC2177, PLAG!, PSMC6P and LYN. 
     
     
       30. The method according to claim 22, wherein the cancer is neuroblastoma, and wherein the one or more genes include MYCN. 
     
     
       31. The method according to claim 22, wherein the cancer is selected from the group consisting of esophageal cancer, breast cancer, and ovarian cancer, and wherein the one or more genes include ErbB2. 
     
     
       32. The method according to claim 22, wherein the cancer is breast cancer or lung cancer, and wherein the one or more genes are selected from the group consisting of: Her2, Her2/Neu, and Her1. 
     
     
       33. The method according to claim 22, wherein the cancer is colorectal cancer or laryngeal cancer, and wherein the one or more genes include Cyclin A or Cyclin D1. 
     
     
       34. The method according to claim 22, wherein the cancer is selected from the group consisting of: leukemia and lymphoma, and wherein the one or more genes include ABL. 
     
     
       35. The method according to claim 22, wherein the cancer is non-small cell lung cancer, and wherein the one or more genes include SKP2. 
     
     
       36. The method according to claim 22, wherein the cancer is myelodysplastic syndrome, and wherein the one or more genes include ETV6. 
     
     
       37. The method according to claim 22, wherein the cancer is selected from the group consisting of breast cancer, ovarian cancer, head and neck cancer, pancreatic cancer, liver cancer, stomach cancer, and colorectal cancer, and wherein the one or more genes include hTR, hTERT, or TEPI. 
     
     
       38. The method according to claim 22, wherein the cancer is nasopharyngeal cancer, and wherein the one or more genes is not Epstein Barr Virus DNA. 
     
     
       39. The method according to claim 22, further comprising following up with the subject based on the presence of the gene amplification.