title: Development of DNA damage response signaling biomarkers using automated, quantitative image analysis.
abstract: The DNA damage response (DDR) coordinates DNA repair with cell cycle checkpoints to ameliorate or mitigate the pathological effects of DNA damage. Automated quantitative analysis (AQUA) and Tissue Studio are commercial technologies that use digitized immunofluorescence microscopy images to quantify antigen expression in defined tissue compartments. Because DDR is commonly activated in cancer and may reflect genetic instability within the lesion, a method to quantify DDR in cancer offers potential diagnostic and/or prognostic value. In this study, both AQUA and Tissue Studio algorithms were used to quantify the DDR in radiation-damaged skin fibroblasts, melanoma cell lines, moles, and primary and metastatic melanomas. Digital image analysis results for three markers of DDR (γH2AX, P-ATM, P-Chk2) correlated with immunoblot data for irradiated fibroblasts, whereas only γH2AX and P-Chk2 correlated with immunoblot data in melanoma cell lines. Melanoma cell lines displayed substantial variation in γH2AX and P-Chk2 expression, and P-Chk2 expression was significantly correlated with radioresistance. Moles, primary melanomas, and melanoma metastases in brain, lung and liver displayed substantial variation in γH2AX expression, similar to that observed in melanoma cell lines. Automated digital analysis of immunofluorescent images stained for DDR biomarkers may be useful for predicting tumor response to radiation and chemotherapy.
authors.affiliation: Translational Pathology Laboratory (NNF, SMC, BM, MO, CRM), University of North Carolina School of Medicine, NC, USA.
authors.forename: Nana
authors.initials: N
authors.lastname: Nikolaishvilli-Feinberg
authors.forename: Stephanie M
authors.initials: SM
authors.lastname: Cohen
authors.forename: Bentley
authors.initials: B
authors.lastname: Midkiff
authors.forename: Yingchun
authors.initials: Y
authors.lastname: Zhou
authors.forename: Mark
authors.initials: M
authors.lastname: Olorvida
authors.forename: Joseph G
authors.initials: JG
authors.lastname: Ibrahim
authors.forename: Bernard
authors.initials: B
authors.lastname: Omolo
authors.forename: Janiel M
authors.initials: JM
authors.lastname: Shields
authors.forename: Nancy E
authors.initials: NE
authors.lastname: Thomas
authors.forename: Pamela A
authors.initials: PA
authors.lastname: Groben
authors.forename: William K
authors.initials: WK
authors.lastname: Kaufmann
authors.forename: C Ryan
authors.initials: CR
authors.lastname: Miller
mesh_terms: D064007:Ataxia Telangiectasia Mutated Proteins / Q000378:metabolism*; D014408:Biomarkers, Tumor / Q000378:metabolism*; D045744:Cell Line, Tumor; D064447:Checkpoint Kinase 2 / Q000378:metabolism*; D004249:DNA Damage; D005347:Fibroblasts / Q000378:metabolism / Q000528:radiation effects; D006657:Histones / Q000378:metabolism*; D006801:Humans; D007091:Image Processing, Computer-Assisted; D008545:Melanoma / Q000175:diagnosis / Q000378:metabolism; D009362:Neoplasm Metastasis; D009506:Nevus / Q000175:diagnosis / Q000378:metabolism; D012867:Skin / Q000166:cytology; D012878:Skin Neoplasms / Q000175:diagnosis / Q000378:metabolism
chemical_list: D014408:Biomarkers, Tumor; C468783:H2AX protein, human; D006657:Histones; D064447:Checkpoint Kinase 2; D064007:Ataxia Telangiectasia Mutated Proteins; C578634:CHEK2 protein, human
keywords: DNA damage response; Digital-image-analysis; P-Chk2; automated quantitative analysis; immunofluorescence; ionizing radiation; melanoma; γH2AX