title: A method for screening and validation of resistant mutations against kinase inhibitors.
abstract: The discovery of BCR/ABL as a driver oncogene in chronic myeloid leukemia (CML) resulted in the development of Imatinib, which, in fact, demonstrated the potential of targeting the kinase in cancers by effectively treating the CML patients. This observation revolutionized drug development to target the oncogenic kinases implicated in various other malignancies, such as, EGFR, B-RAF, KIT and PDGFRs. However, one major drawback of anti-kinase therapies is the emergence of drug resistance mutations rendering the target to have reduced or lost affinity for the drug. Understanding the mechanisms employed by resistant variants not only helps in developing the next generation inhibitors but also gives impetus to clinical management using personalized medicine. We reported a retroviral vector based screening strategy to identify the spectrum of resistance conferring mutations in BCR/ABL, which has helped in developing the next generation BCR/ABL inhibitors. Using Ruxolitinib and JAK2 as a drug target pair, here we describe in vitro screening methods that utilizes the mouse BAF3 cells expressing the random mutation library of JAK2 kinase.
authors.affiliation: Divisions of Experimental Hematology and Cancer Pathology, Cancer Blood Disease Institute, Cincinnati Children's Hospital Medical Center.
authors.forename: Meenu
authors.initials: M
authors.lastname: Kesarwani
authors.affiliation: Divisions of Experimental Hematology and Cancer Pathology, Cancer Blood Disease Institute, Cincinnati Children's Hospital Medical Center.
authors.forename: Erika
authors.initials: E
authors.lastname: Huber
authors.affiliation: Divisions of Experimental Hematology and Cancer Pathology, Cancer Blood Disease Institute, Cincinnati Children's Hospital Medical Center.
authors.forename: Zachary
authors.initials: Z
authors.lastname: Kincaid
authors.affiliation: Divisions of Experimental Hematology and Cancer Pathology, Cancer Blood Disease Institute, Cincinnati Children's Hospital Medical Center; mohammad.azam@cchmc.org.
authors.forename: Mohammad
authors.initials: M
authors.lastname: Azam
mesh_terms: D000818:Animals; D004252:DNA Mutational Analysis / Q000379:methods*; D019008:Drug Resistance, Neoplasm / Q000235:genetics*; D016044:Fusion Proteins, bcr-abl / Q000235:genetics; D005820:Genetic Testing / Q000379:methods*; D006801:Humans; D000068877:Imatinib Mesylate / Q000494:pharmacology; D015464:Leukemia, Myelogenous, Chronic, BCR-ABL Positive / Q000235:genetics; D051379:Mice; D009154:Mutation; D009570:Nitriles; D047428:Protein Kinase Inhibitors / Q000494:pharmacology*; D011720:Pyrazoles / Q000494:pharmacology; D011743:Pyrimidines
chemical_list: D009570:Nitriles; D047428:Protein Kinase Inhibitors; D011720:Pyrazoles; D011743:Pyrimidines; C540383:ruxolitinib; D000068877:Imatinib Mesylate; D016044:Fusion Proteins, bcr-abl