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DNA Repair and Damage Signaling in Hypoxic
Cells
Robert G. Bristow, MD PhD FRCPC
Applied Molecular Oncology Division, Ontario Cancer Institute and Princess
Margaret Hospital and Departments of Radiaiton Oncology and Medical
Biophysics, University of Toronto
Email: rob.bristow@rmp.uhn.on.ca
Phone: 416-946-2129
FAX: 416-946-3586
Our laboratory is interested in determining those factors which control
genetic instability and the emergence of aggressive tumour cell variants
within hypoxic tumour subpopulations. We are especially interested in how
well hypoxic normal and tumour cells repair DNA double-strand breaks
(DNA-dsbs). DNA-dsbs are created in the nucleus by exogenous agents such as
radiotherapy or chemotherapy or can be generated endogenously during DNA
replication in the S-phase of the cell cycle. Human cells have two main
DNA-dsb repair pathways: (1) homologous recombination (HR) driven by the
activity of the RAD51-BRCA2 repair complex and (2), non-homologous
recombination or end-joining (NHEJ) driven by the DNA-PK and XRCC4-Ligase IV
complexes. Although many details of these pathways have been discovered
using yeast and mammalian genetics under oxic conditions, the data is sparse
regarding the effecitveness of these pathways under hypoxic conditions.
Our laboratory uses in vitro and in vivo
(xenograft) models of prostate, lung, and cervix cancer under oxic and hypoxic conditions to study genetic
instability within tumour cells. We also utilize experimental methods
pertaining to the COMET assay (alkaline and neutral lysis conditions),
continuous-field gel electrophoresis (CFGE), plasmid-based HR and chromosomal damage assays, apoptosis and clonogenic cell kill assays, and
immunofluorescence biology. We are particularly interested in the study of
the formation of DNA repair complexes in situ within the nucleus under
hypoxic conditions using immunofluorescent confocal and multi-photon
microscopy.
It is hoped that by studying DNA repair under oxic and hypoxic conditions,
we will be able to understand the process of prostate and cervix carcinogenesis and understand the consequences of the cancer "mutator"
phenotype. We hope to use this information to discover new
biomarkers for tumour prognosis and develop novel therapeutic agents directed against
hypoxic cells during cellular carcinogenesis and tumour
progression.
2004 Lab Group
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