METHODS FOR DETERMINING RESPONSIVENESS TO A DRUG BASED UPON DETERMINATION OF RAS MUTATION AND/OR RAS AMPLIFICATION
The present disclosure provides methods for predicting the sensitivity (e.g., responsiveness) of a cell and/or biological sample obtained from a subject (e.g., a human) to a drug (e.g., a DHFR inhibitor). Such methods may comprise determining the presence or absence of one or more Ras mutations and/or determining the presence or absence of an amplification of the Ras gene in the cell and/or biological sample. The methods may be used to predict the responsiveness of a subject to treatment with a drug.
1 . A method for predicting sensitivity of a test cell to a drug, the method comprising:
a. obtaining a test cell;
b. assaying the test cell for one or more Ras mutations;
c. assaying the test cell for amplification of a Ras gene;
d. determining if one or more Ras mutations are present or absent in the test cell and determining if an amplification of the Ras gene is present or absent in the test cell; and
e. employing the determination of the presence or absence of a Ras mutation in the test cell and the presence or absence of an amplification of Ras in the test cell to predict sensitivity of the test cell to the drug.
2 . The method of claim 1 , wherein Ras is k-Ras (SEQ ID NO: 1), n-Ras (SEQ ID NO: 2) or h-Ras (SEQ ID NO: 3).
3 . The method of claim 2 , wherein the k-Ras mutations are at one or more of positions 12, 13 or 61.
4 . The method of claim 3 , wherein the k-Ras mutations are selected from the group consisting of: G12A, G12N, G12R, G12C, G12S, G12V, G13N and Q61H.
5 . The method of claim 2 , wherein the h-Ras or n-Ras mutations are at one or more of positions 12, 13 or 61.
6 . The method of claim 1 , wherein the drug is a chemotherapeutic agent.
7 . The method of claim 1 , wherein the drug is an antifolate.
8 . The method of claim 7 , wherein the antifolate is a dihydrofolate reductase (DHFR) inhibitor.
9 . The method of claim 8 , wherein the DHFR inhibitor is Methotrexate or Pemetrexed.
10 . The method of claim 1 , wherein the drug is a tyrosine kinase inhibitor that targets HER1 (EGFR), HER2/neu, HER3, or any combination thereof.
11 . The method of claim 10 , wherein the tyrosine kinase inhibitor is an antibody.
12 . The method of claim 11 , wherein the antibody is monoclonal antibody.
13 . The method of claim 12 , wherein the monoclonal antibody is cetuximab (Erbitux), panitumumab, zalutumumab, nimotuzumab or matuzmab.
14 . The method of claim 10 , wherein the tyrosine kinase inhibitor is a small molecule inhibitor.
15 . The method of claim 14 , wherein the small molecule inhibitor is gefitinib, erlotinib or lapatinib.
16 . The method of claim 1 , wherein the test cell is obtained from a subject that has a disease or disorder.
17 . The method of claim 16 , wherein the disease or disorder is cancer.
18 . The method of claim 17 , wherein the cancer is selected from the group consisting of gastrointestinal cancer, prostate cancer, ovarian cancer, breast cancer, head and neck cancer, lung cancer, non-small cell lung cancer, cancer of the nervous system, kidney cancer, retina cancer, skin cancer, liver cancer, pancreatic cancer, genital urinary cancer and bladder cancer.
19 . The method of claim 16 , wherein the subject is a cancer patient.
20 . The method of claim 1 , wherein the test cell is assayed for one or more Ras mutations and an amplification of Ras by analyzing nucleic acid obtained from the test cell.
21 . The method of claim 1 , wherein the test cell is assayed for one or more Ras mutations by analyzing proteins obtained from the test cell.
22 . The method of claim 1 , wherein test cell is obtained from a tumor biopsy.
23 . The method of claim 1 , wherein the test cell is obtained from an aspirate, blood or serum.
24 . The method of claim 1 , wherein the test cell is predicted to be sensitive to the drug where one or more Ras mutations are determined to be present in the test cell and an amplification of Ras is determined to be present in the test cell.
25 . The method of claim 1 , wherein the test cell is predicted to be sensitive to the drug where one or more Ras mutations are determined to be present in the test cell and amplification of Ras is determined to be absent in the test cell.
26 . The method of claim 1 , wherein the test cell is predicted to be sensitive to the drug where Ras mutations are determined to be absent in the test cell and an amplification of Ras is determined to be present in the test cell.
27 . The method of claim 1 , wherein the test cell is predicted to be sensitive to the Drug where Ras mutations are determined to be absent in the test cell and amplification of Ras is determined to be absent in the test cell.
28 . The method of claim 1 , wherein the test cell is predicted to be insensitive to the drug where one ore more Ras mutations are determined to be present in the test cell and an amplification of Ras is determined to be present in the test cell.
29 . The method of claim 1 , wherein the test cell is predicted to be insensitive to the drug where one or more Ras mutations are determined to be present in the test cell and amplification of Ras is determined to be present in the test cell.
30 . The method of claim 1 , wherein the test cell is predicted to be insensitive to the drug where Ras mutations are determined to be absent in the test cell and an amplification of Ras is determined to be present in the test cell.
31 . The method of claim 1 , wherein the test cell is predicted to be insensitive to the drug where Ras mutations are determined to be absent in the test cell and amplification of Ras is determined to be absent in the test cell.
32 . The method of claim 1 , wherein the step of assaying the test cell for one or more Ras mutations and amplification of Ras is performed by in situ hybridization (TSH), northern blot, qRT-PCT or microarray analysis.