title: Evaluation of O6-methylguanine-DNA methyltransferase by immunohistochemistry: best clinical and research practices.
abstract: O6-Methyguanine-DNA methyltransferase (MGMT) repairs DNA damage and acts as a tumor suppressor in normal cells by preventing DNA mutations. Several antibodies against MGMT are used for immunohistochemical assessment of this marker and no universal standard is adopted. We evaluated the immunohistochemical expression of MGMT with 5 commercially available primary antibodies in 59 invasive breast carcinomas. A tissue microarray was constructed using 59 invasive breast carcinoma samples. Five primary antibodies against MGMT were used for immunohistochemistry, including clones MT3.1, SPM287, and MT23.2. Heat-induced antigen retrieval and polymer-based immunohistochemistry were performed. Stains were analyzed by microscopy and automated digital slide technology. qRT-PCR was performed for all tumors. Clone SPM287 had the highest sensitivity (p<0.001), and clone MT3.1 had the lowest sensitivity (p<0.001). Clone MT23.2 generated higher levels of cytoplasmic staining, which was not observed with the other antibodies (p<0.001). Fifty-six samples (94.9%) showed hypoexpression of MGMT compared with normal breast, as measured by qRT-PCR (p<0.001). Only clone SPM287 correlated significantly with the qRT-PCR results (p=0.027). Antibody clone SPM287 is the most sensitive and specific antibody for the immunohistochemical evaluation of MGMT, rendering it a reliable and effective reagent for research and clinical practice in breast cancer.
authors.affiliation: Dept of Anatomic Pathology, Cancer Hospital A C Camargo, São Paulo, Brazil.
authors.forename: Julio Cirullo
authors.initials: JC
authors.lastname: Neto
authors.forename: Katia
authors.initials: K
authors.lastname: Carvalho
authors.forename: Eloisa
authors.initials: E
authors.lastname: Olivieri
authors.forename: Dirce
authors.initials: D
authors.lastname: Carraro
authors.forename: Isabela
authors.initials: I
authors.lastname: Cunha
authors.forename: José
authors.initials: J
authors.lastname: Vassallo
authors.forename: Luciane
authors.initials: L
authors.lastname: Kagohara
authors.forename: Fernando
authors.initials: F
authors.lastname: Soares
authors.forename: Rafael
authors.initials: R
authors.lastname: Rocha
mesh_terms: D000911:Antibodies, Monoclonal; D000918:Antibody Specificity; D014408:Biomarkers, Tumor / Q000032:analysis*; D001943:Breast Neoplasms / Q000201:enzymology* / Q000473:pathology; D018270:Carcinoma, Ductal, Breast / Q000201:enzymology* / Q000473:pathology; D015254:DNA Modification Methylases / Q000032:analysis*; D045643:DNA Repair Enzymes / Q000032:analysis*; D005260:Female; D006801:Humans; D007150:Immunohistochemistry / Q000592:standards*; D017410:Practice Guidelines as Topic; D060888:Real-Time Polymerase Chain Reaction; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012680:Sensitivity and Specificity; D046888:Tissue Array Analysis; D025521:Tumor Suppressor Proteins / Q000032:analysis*
chemical_list: D000911:Antibodies, Monoclonal; D014408:Biomarkers, Tumor; D025521:Tumor Suppressor Proteins; D015254:DNA Modification Methylases; C515491:MGMT protein, human; D045643:DNA Repair Enzymes