Methods for diagnosing colon cancer using MicroRNA signatures
The present invention provides novel methods and compositions for the diagnosis and treatment of solid cancers. The invention also provides methods of identifying inhibitors of tumorigenesis.
1. A method of diagnosing whether a subject has, or is at risk for developing, colon cancer, comprising:
receiving a test sample of colon tissue extracted from a subject having, or suspected of having, colon cancer;
measuring by Northern blot analysis the levels of miR-29b-2 gene product and at least one additional miR gene product in the test sample, wherein the at least one additional miR gene product comprises miR-21, and wherein the Northern blot analysis comprises:
extracting nucleic acids from the test sample;
treating the extracted nucleic acids with DNase to remove DNA;
separating RNA molecules in the extracted nucleic acids by gel electrophoresis;
hybridizing the separated RNA molecules to labeled probes; and
detecting and quantifying the hybridization to determine the level of at least miR-29b-2 and miR-21 gene products;
comparing the levels of the miR-29b-2 gene product and the at least one additional miR gene product in the test sample from the subject to the levels of corresponding gene products in a control sample of normal colon tissue; and
diagnosing the subject as having, or being at risk for developing, colon cancer if the levels of miR-29b-2 gene product the at least one additional miR gene product in the test sample are higher than the levels of corresponding gene products in the control sample.
2. A method of diagnosing whether a subject has, or is at risk for developing, colon cancer, comprising:
receiving a test sample of colon tissue extracted from a subject having, or suspected of having, colon cancer;
reverse transcribing at least two miR gene products from the test sample to provide at least two corresponding miR gene product target oligonucleotides;
hybridizing at least two miR target oligodeoxynucleotides to a microarray comprising miRNA-specific probe oligonucleotides that include at least two corresponding miRNA-specific probe oligonucleotides to provide a hybridization profile for the test sample, wherein the probe oligonucleotides are labeled with one of biotin, a radionuclide, a heavy metal, a fluorescent molecule, a chemiluminescent molecule, or an enzyme;
comparing the test sample hybridization profile to a control hybridization profile for at least the two miR gene products in normal colon tissue; and
diagnosing the subject as having, or being at risk for developing, colon cancer if the level of the at least two gene products in the test sample is greater than the control level of the two gene products, wherein the two miR gene products comprise miR-29b-2 and miR-21.
3. The method of claim 2 , wherein the probes are covalently attached to a polymeric matrix.
4. The method of claim 2 , wherein the miRNA-specific probe oligonucleotides comprise a sequence having at least about 70% complementarity to miR-29b-2.
5. The method of claim 2 , wherein the miRNA-specific probe oligonucleotides comprise a sequence having at least about 70% complementarity to miR-21.
6. A method of diagnosing whether a subject has, or is at risk for developing, colon cancer, comprising:
receiving a test sample of colon tissue extracted from a subject having, or suspected of having, colon cancer;
reverse transcribing at least two miR gene products from the test sample to provide at least two corresponding miR gene product target oligonucleotides;
hybridizing at least two miR target oligodeoxynucleotides to a microarray comprising miRNA-specific probe oligonucleotides that include at least two corresponding miRNA-specific probe oligonucleotides covalently attached to a polymeric matrix, to provide a hybridization profile for the test sample;
comparing the test sample hybridization profile to a control hybridization profile for the at least two miR gene products in normal colon tissue; and
diagnosing the subject as having, or being at risk for developing, colon cancer if the level of the at least two gene products in the test sample is greater than the control level of the two gene products, wherein the two miR gene products comprise miR-29b-2 and miR-21.