title: Antibody validation and scoring guidelines for ABCG2 immunohistochemical staining in formalin-fixed paraffin-embedded colon cancer tissue.
abstract: Overexpression of the ATP-dependent drug efflux pump ABCG2 is a major molecular mechanism of multidrug resistance in cancer and might be a predictive biomarker for drug response. Contradictory results have been reported for immunohistochemical studies of ABCG2 protein expression in colorectal cancer (CRC), probably because of the use of different antibodies and scoring approaches. In this study, we systematically studied six commercially available anti-ABCG2 antibodies, using cell lines with up-regulation of ABCG2, and selected one antibody for validation in CRC tissue. Furthermore, we established scoring guidelines for ABCG2 expression based on the clinically used guidelines for HER2 immunohistochemistry assessment in gastric cancer. The guidelines provide a semi-quantitative measure of the basolateral membrane staining of ABCG2 and disregard the apical membrane staining and the cytoplasmic signal. Intra-tumor heterogeneity in ABCG2 immunoreactivity was observed; however, statistical analyses of tissue microarrays (TMAs) and the corresponding whole sections from primary tumors of 57 metastatic CRC patients revealed a strong positive correlation between maximum TMA scores and whole sections, especially when more than one core was used. In conclusion, here, we provide validated results to guide future studies on the associations between ABCG2 immunoreactivity in tumor cells and the benefits of chemotherapeutic treatment in patients with CRC.
authors.affiliation: Faculty of Health and Medical Sciences, Department of Veterinary Disease Biology, Section for Molecular Disease Biology, University of Copenhagen, Strandboulevarden 49, DK- 2100 Copenhagen Ø, Denmark.
authors.forename: Camilla Natasha
authors.initials: CN
authors.lastname: Cederbye
authors.affiliation: Department of Oncology, Herlev and Gentofte Hospital, University of Copenhagen, Herlev Ringvej 75, DK-2730 Herlev, Denmark.
authors.forename: Jesper Andreas
authors.initials: JA
authors.lastname: Palshof
authors.affiliation: Department of Pathology, Hvidovre Hospital, Kettegaard Alle 30, DK- 2650 Hvidovre, Denmark.
authors.forename: Tine Plato
authors.initials: TP
authors.lastname: Hansen
authors.affiliation: Danish Cancer Society, Strandboulevarden 49, DK- 2100 Copenhagen Ø, Denmark.
authors.forename: Anne Katrine
authors.initials: AK
authors.lastname: Duun-Henriksen
authors.affiliation: Department of Pathology, Herlev and Gentofte Hospital, University of Copenhagen, Herlev Ringvej 75, DK-2730 Herlev, Denmark.
authors.forename: Dorte
authors.initials: D
authors.lastname: Linnemann
authors.affiliation: Faculty of Health and Medical Sciences, Department of Veterinary Disease Biology, Section for Molecular Disease Biology, University of Copenhagen, Strandboulevarden 49, DK- 2100 Copenhagen Ø, Denmark.
authors.forename: Jan
authors.initials: J
authors.lastname: Stenvang
authors.affiliation: Department of Oncology, Herlev and Gentofte Hospital, University of Copenhagen, Herlev Ringvej 75, DK-2730 Herlev, Denmark.
authors.forename: Dorte Lisbet
authors.initials: DL
authors.lastname: Nielsen
authors.affiliation: Faculty of Health and Medical Sciences, Department of Veterinary Disease Biology, Section for Molecular Disease Biology, University of Copenhagen, Strandboulevarden 49, DK- 2100 Copenhagen Ø, Denmark.
authors.forename: Nils
authors.initials: N
authors.lastname: Brünner
authors.affiliation: Faculty of Health and Medical Sciences, Department of Veterinary Disease Biology, Section for Molecular Disease Biology, University of Copenhagen, Strandboulevarden 49, DK- 2100 Copenhagen Ø, Denmark.
authors.forename: Birgitte Martine
authors.initials: BM
authors.lastname: Viuff
mesh_terms: D000070997:ATP Binding Cassette Transporter, Subfamily G, Member 2 / Q000276:immunology / Q000378:metabolism*; D058846:Antibodies, Monoclonal, Murine-Derived / Q000737:chemistry* / Q000276:immunology; D000918:Antibody Specificity; D014408:Biomarkers, Tumor / Q000276:immunology / Q000378:metabolism*; D045744:Cell Line, Tumor; D003110:Colonic Neoplasms / Q000175:diagnosis / Q000378:metabolism*; D003429:Cross Reactions; D015536:Down-Regulation; D005404:Fixatives / Q000737:chemistry; D005557:Formaldehyde / Q000737:chemistry; D006801:Humans; D007150:Immunohistochemistry; D009363:Neoplasm Proteins / Q000276:immunology / Q000378:metabolism*; D016612:Paraffin Embedding; D034622:RNA Interference; D034741:RNA, Small Interfering / Q000235:genetics; D016707:Tissue Fixation
chemical_list: C116614:ABCG2 protein, human; D000070997:ATP Binding Cassette Transporter, Subfamily G, Member 2; D058846:Antibodies, Monoclonal, Murine-Derived; D014408:Biomarkers, Tumor; D005404:Fixatives; D009363:Neoplasm Proteins; D034741:RNA, Small Interfering; D005557:Formaldehyde