title: Rapid Identification of Chemoresistance Mechanisms Using Yeast DNA Mismatch Repair Mutants.
abstract: Resistance to cancer therapy is a major obstacle in the long-term treatment of cancer. A greater understanding of drug resistance mechanisms will ultimately lead to the development of effective therapeutic strategies to prevent resistance from occurring. Here, we exploit the mutator phenotype of mismatch repair defective yeast cells combined with whole genome sequencing to identify drug resistance mutations in key pathways involved in the development of chemoresistance. The utility of this approach was demonstrated via the identification of the known CAN1 and TOP1 resistance targets for two compounds, canavanine and camptothecin, respectively. We have also experimentally validated the plasma membrane transporter HNM1 as the primary drug resistance target of mechlorethamine. Furthermore, the sequencing of mitoxantrone-resistant strains identified inactivating mutations within IPT1, a gene encoding inositolphosphotransferase, an enzyme involved in sphingolipid biosynthesis. In the case of bactobolin, a promising anticancer drug, the endocytosis pathway was identified as the drug resistance target responsible for conferring resistance. Finally, we show that that rapamycin, an mTOR inhibitor previously shown to alter the fitness of the ipt1 mutant, can effectively prevent the formation of mitoxantrone resistance. The rapid and robust nature of these techniques, using Saccharomyces cerevisiae as a model organism, should accelerate the identification of drug resistance targets and guide the development of novel therapeutic combination strategies to prevent the development of chemoresistance in various cancers.
authors.affiliation: Department of Molecular Biology, Princeton University, Princeton, New Jersey 08544.
authors.forename: Irene
authors.initials: I
authors.lastname: Ojini
authors.affiliation: Department of Molecular Biology, Princeton University, Princeton, New Jersey 08544 agammie@princeton.edu.
authors.forename: Alison
authors.initials: A
authors.lastname: Gammie
mesh_terms: D001578:Benzopyrans / Q000494:pharmacology; D053843:DNA Mismatch Repair; D055808:Drug Discovery; D019008:Drug Resistance, Neoplasm; D004354:Drug Screening Assays, Antitumor; D004705:Endocytosis / Q000187:drug effects / Q000235:genetics; D016681:Genome, Fungal; D023281:Genomics / Q000379:methods; D059014:High-Throughput Nucleotide Sequencing; D008466:Mechlorethamine / Q000494:pharmacology; D009154:Mutation; D012441:Saccharomyces cerevisiae / Q000187:drug effects* / Q000235:genetics* / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics / Q000378:metabolism; D054852:Small Molecule Libraries; D013107:Sphingolipids / Q000096:biosynthesis
chemical_list: D001578:Benzopyrans; D029701:Saccharomyces cerevisiae Proteins; D054852:Small Molecule Libraries; D013107:Sphingolipids; D008466:Mechlorethamine; C025268:bactobolin
keywords: DNA mismatch repair; cancer; drug resistance; mutator; whole genome sequencing