title: Sensitization of Pancreatic Cancers to Gemcitabine Chemoradiation by WEE1 Kinase Inhibition Depends on Homologous Recombination Repair.
abstract: To improve the efficacy of chemoradiation therapy for locally advanced pancreatic cancer and begin to establish patient selection criteria, we investigated the combination of the WEE1 inhibitor AZD1775 with gemcitabine-radiation in homologous recombination (HR) repair proficient and deficient pancreatic cancers. Sensitization to gemcitabine-radiation by AZD1775 was assessed in pancreatic cancer cells by clonogenic survival and in patient-derived xenografts by tumor growth. The contributions of HR repair inhibition and G2 checkpoint abrogation to sensitization were assessed by γH2AX, BRCA2 manipulation, and RAD51 focus formation and pHistone H3 flow cytometry, respectively. We found that AZD1775 sensitized to gemcitabine-radiation in BRCA2 wild-type but not BRCA2 mutant pancreatic cancer cells. In all cells, AZD1775 caused inhibition of CDK1 phosphorylation and G2 checkpoint abrogation. However, sensitization by AZD1775 was associated with persistent γH2AX and inhibition of RAD51 focus formation. In HR-proficient (BRCA2 wild-type) or -deficient (BRAC2 null) isogenic cells, AZD1775 sensitized to gemcitabine-radiation in BRCA2 wild-type, but not in BRCA2 null cells, despite significant G2 checkpoint abrogation. In patient-derived pancreatic tumor xenografts, AZD1775 significantly inhibited tumor growth and impaired RAD51 focus formation in response to gemcitabine-radiation. In conclusion, WEE1 inhibition by AZD1775 is an effective strategy for sensitizing pancreatic cancers to gemcitabine chemoradiation. Although this sensitization is accompanied by inhibition of CDK1 phosphorylation and G2 checkpoint abrogation, this mechanism is not sufficient for sensitization. Our findings demonstrate that sensitization to chemoradiation by WEE1 inhibition results from inhibition of HR repair and suggest that patient tumors without underlying HR defects would benefit most from this therapy.
authors.affiliation: Department of Radiation Oncology, University of Michigan Medical School, Ann Arbor, MI 48109.
authors.forename: Tasneem
authors.initials: T
authors.lastname: Kausar
authors.affiliation: Department of Radiation Oncology, University of Michigan Medical School, Ann Arbor, MI 48109.
authors.forename: Jason S
authors.initials: JS
authors.lastname: Schreiber
authors.affiliation: Department of Radiation Oncology, University of Michigan Medical School, Ann Arbor, MI 48109.
authors.forename: David
authors.initials: D
authors.lastname: Karnak
authors.affiliation: Department of Pharmacology, University of Michigan Medical School, Ann Arbor, MI 48109.
authors.forename: Leslie A
authors.initials: LA
authors.lastname: Parsels
authors.affiliation: Department of Pharmacology, University of Michigan Medical School, Ann Arbor, MI 48109.
authors.forename: Joshua D
authors.initials: JD
authors.lastname: Parsels
authors.affiliation: Department of Radiation Oncology, University of Michigan Medical School, Ann Arbor, MI 48109.
authors.forename: Mary A
authors.initials: MA
authors.lastname: Davis
authors.affiliation: Biostatistics Unit, University of Michigan Comprehensive Cancer Center, Ann Arbor, MI 48109.
authors.forename: Lili
authors.initials: L
authors.lastname: Zhao
authors.affiliation: Department of Pharmacology, University of Michigan Medical School, Ann Arbor, MI 48109.
authors.forename: Jonathan
authors.initials: J
authors.lastname: Maybaum
authors.affiliation: Department of Radiation Oncology, University of Michigan Medical School, Ann Arbor, MI 48109.
authors.forename: Theodore S
authors.initials: TS
authors.lastname: Lawrence
authors.affiliation: Department of Radiation Oncology, University of Michigan Medical School, Ann Arbor, MI 48109. Electronic address: mmccrack@med.umich.edu.
authors.forename: Meredith A
authors.initials: MA
authors.lastname: Morgan
mesh_terms: D000818:Animals; D017209:Apoptosis / Q000187:drug effects; D015153:Blotting, Western; D018797:Cell Cycle Proteins / Q000037:antagonists & inhibitors* / Q000378:metabolism; D049109:Cell Proliferation / Q000187:drug effects; D059248:Chemoradiotherapy; D004249:DNA Damage / Q000187:drug effects; D003841:Deoxycytidine / Q000031:analogs & derivatives* / Q000494:pharmacology; D005260:Female; D005434:Flow Cytometry; D005455:Fluorescent Antibody Technique; D006801:Humans; D007124:Immunoenzyme Techniques; D051379:Mice; D008819:Mice, Nude; D009687:Nuclear Proteins / Q000037:antagonists & inhibitors* / Q000378:metabolism; D010190:Pancreatic Neoplasms / Q000235:genetics / Q000473:pathology / Q000628:therapy*; D010766:Phosphorylation / Q000187:drug effects; D047428:Protein Kinase Inhibitors / Q000494:pharmacology*; D011505:Protein-Tyrosine Kinases / Q000037:antagonists & inhibitors* / Q000378:metabolism; D011838:Radiation-Sensitizing Agents / Q000494:pharmacology*; D059767:Recombinational DNA Repair / Q000187:drug effects*; D014407:Tumor Cells, Cultured; D023041:Xenograft Model Antitumor Assays; D000093542:Gemcitabine
chemical_list: D018797:Cell Cycle Proteins; D009687:Nuclear Proteins; D047428:Protein Kinase Inhibitors; D011838:Radiation-Sensitizing Agents; D003841:Deoxycytidine; D011505:Protein-Tyrosine Kinases; C499349:WEE1 protein, human; D000093542:Gemcitabine