title: WTp53 induction does not override MTp53 chemoresistance and radioresistance due to gain-of-function in lung cancer cells.
abstract: New molecular cancer treatment strategies aim to reconstitute wild-type p53 (WTp53) function in mutant p53 (MTp53)-expressing tumors as a means of resensitizing cells to chemotherapy or radiotherapy. The success of this approach may depend on whether MTp53 proteins are acting in a dominant-negative or independent gain-of-function mode. Herein, we describe an isogenic, temperature-sensitive p53 model (p53(A138V)) in p53-null human H1299 lung cancer cells in which WTp53 can be selectively coexpressed with a temperature-sensitive MTp53 allele (A138V) during initial DNA damage and subsequent DNA repair. Cells expressing MTp53 alone or coexpressing induced WTp53 and MTp53 were tested for p53 transcription, G(1) and G(2) cell cycle checkpoints, apoptosis, and long-term clonogenic survival following DNA damage. Transient transfection of WTp53 into H1299 cells, or shift-down of H1299-p53(A138V) stable transfectants to 32 degrees C to induce WTp53, led to increased p21(WAF1) expression and G(1) and G(2) arrests following DNA damage but did not increase BAX expression or apoptosis. In contrast, both transient and stable expression of the p53(A138V) mutant in p53-null H1299 cells (e.g. testing gain-of-function) at 37 degrees C blocked p21(WAF1) induction following DNA damage. Cell death was secondary to mitotic catastrophe and/or tumor cell senescence. Overexpression of WTp53 did not resensitize resistant MTp53-expressing cells to ionizing radiation, cisplatinum, or mitomycin C. Our results suggest that human MTp53 proteins can lead to resistant phenotypes independent of WTp53-mediated transcription and checkpoint control. This should be considered when using p53 as a prognostic factor and therapeutic target.
authors.affiliation: Division of Applied Molecular Oncology, Ontario Cancer Institute, Princess Margaret Hospital (University Health Network), University of Toronto, Toronto, Canada
authors.forename: Andrew R
authors.initials: AR
authors.lastname: Cuddihy
authors.forename: Farid
authors.initials: F
authors.lastname: Jalali
authors.forename: Carla
authors.initials: C
authors.lastname: Coackley
authors.forename: Robert G
authors.initials: RG
authors.lastname: Bristow
mesh_terms: D000903:Antibiotics, Antineoplastic / Q000494:pharmacology; D017209:Apoptosis / Q000187:drug effects / Q000528:radiation effects; D049109:Cell Proliferation / Q000187:drug effects / Q000528:radiation effects; D016922:Cellular Senescence; D002945:Cisplatin / Q000494:pharmacology; D050759:Cyclin-Dependent Kinase Inhibitor p21 / Q000378:metabolism; D004249:DNA Damage / Q000187:drug effects / Q000528:radiation effects; D019008:Drug Resistance, Neoplasm / Q000235:genetics*; D005434:Flow Cytometry; D005455:Fluorescent Antibody Technique; D016193:G1 Phase / Q000187:drug effects / Q000528:radiation effects; D016195:G2 Phase / Q000187:drug effects / Q000528:radiation effects; D005720:Gamma Rays; D006801:Humans; D015151:Immunoblotting; D047468:Immunoprecipitation; D008175:Lung Neoplasms / Q000235:genetics* / Q000628:therapy*; D016685:Mitomycin / Q000494:pharmacology; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D011836:Radiation Tolerance / Q000235:genetics*; D011839:Radiation, Ionizing; D011838:Radiation-Sensitizing Agents / Q000494:pharmacology; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014158:Transcription, Genetic / Q000187:drug effects / Q000528:radiation effects; D014162:Transfection; D014407:Tumor Cells, Cultured / Q000187:drug effects / Q000528:radiation effects; D014410:Tumor Stem Cell Assay; D016159:Tumor Suppressor Protein p53 / Q000235:genetics* / Q000378:metabolism; D051028:bcl-2-Associated X Protein / Q000378:metabolism; D001616:beta-Galactosidase / Q000378:metabolism
chemical_list: D000903:Antibiotics, Antineoplastic; C494826:BAX protein, human; D050759:Cyclin-Dependent Kinase Inhibitor p21; D012333:RNA, Messenger; D011838:Radiation-Sensitizing Agents; D016159:Tumor Suppressor Protein p53; D051028:bcl-2-Associated X Protein; D016685:Mitomycin; D001616:beta-Galactosidase; D002945:Cisplatin