title: Platform comparison for evaluation of ALK protein immunohistochemical expression, genomic copy number and hotspot mutation status in neuroblastomas.
abstract: ALK is an established causative oncogenic driver in neuroblastoma, and is likely to emerge as a routine biomarker in neuroblastoma diagnostics. At present, the optimal strategy for clinical diagnostic evaluation of ALK protein, genomic and hotspot mutation status is not well-studied. We evaluated ALK immunohistochemical (IHC) protein expression using three different antibodies (ALK1, 5A4 and D5F3 clones), ALK genomic status using single-color chromogenic in situ hybridization (CISH), and ALK hotspot mutation status using conventional Sanger sequencing and a next-generation sequencing platform (Ion Torrent Personal Genome Machine (IT-PGM)), in archival formalin-fixed, paraffin-embedded neuroblastoma samples. We found a significant difference in IHC results using the three different antibodies, with the highest percentage of positive cases seen on D5F3 immunohistochemistry. Correlation with ALK genomic and hotspot mutational status revealed that the majority of D5F3 ALK-positive cases did not possess either ALK genomic amplification or hotspot mutations. Comparison of sequencing platforms showed a perfect correlation between conventional Sanger and IT-PGM sequencing. Our findings suggest that D5F3 immunohistochemistry, single-color CISH and IT-PGM sequencing are suitable assays for evaluation of ALK status in future neuroblastoma clinical trials.
authors.affiliation: Department of Pathology and Laboratory Medicine, KK Women's and Children's Hospital, Singapore, Singapore.
authors.forename: Benedict
authors.initials: B
authors.lastname: Yan
authors.affiliation: Department of Pathology and Laboratory Medicine, KK Women's and Children's Hospital, Singapore, Singapore.
authors.forename: Chik Hong
authors.initials: CH
authors.lastname: Kuick
authors.affiliation: Department of Pathology and Laboratory Medicine, KK Women's and Children's Hospital, Singapore, Singapore.
authors.forename: Malcolm
authors.initials: M
authors.lastname: Lim
authors.affiliation: Saw Swee Hock School of Public Health, National University of Singapore and National University Health System, Singapore, Singapore.
authors.forename: Kavita
authors.initials: K
authors.lastname: Venkataraman
authors.affiliation: Genome Institute of Singapore, Singapore, Singapore.
authors.forename: Chandana
authors.initials: C
authors.lastname: Tennakoon
authors.affiliation: Department of Pathology and Laboratory Medicine, KK Women's and Children's Hospital, Singapore, Singapore.
authors.forename: Eva
authors.initials: E
authors.lastname: Loh
authors.affiliation: Department of Pathology and Laboratory Medicine, KK Women's and Children's Hospital, Singapore, Singapore.
authors.forename: Derrick
authors.initials: D
authors.lastname: Lian
authors.affiliation: Department of Pathology and Laboratory Medicine, KK Women's and Children's Hospital, Singapore, Singapore.
authors.forename: May Ying
authors.initials: MY
authors.lastname: Leong
authors.affiliation: Mouse Models for Human Cancer Unit, Institute of Molecular and Cell Biology, Singapore, Singapore.
authors.forename: Manikandan
authors.initials: M
authors.lastname: Lakshmanan
authors.affiliation: Mouse Models for Human Cancer Unit, Institute of Molecular and Cell Biology, Singapore, Singapore.
authors.forename: Vinay
authors.initials: V
authors.lastname: Tergaonkar
authors.affiliation: Genome Institute of Singapore, Singapore, Singapore; School of Computing, National University of Singapore, Singapore, Singapore.
authors.forename: Wing-Kin
authors.initials: WK
authors.lastname: Sung
authors.affiliation: Haematology/Oncology Service, Department of Paediatric Subspecialties, KK Women's and Children's Hospital, Singapore, Singapore.
authors.forename: Shui Yen
authors.initials: SY
authors.lastname: Soh
authors.affiliation: Department of Pathology and Laboratory Medicine, KK Women's and Children's Hospital, Singapore, Singapore.
authors.forename: Kenneth T E
authors.initials: KT
authors.lastname: Chang
mesh_terms: D000077548:Anaplastic Lymphoma Kinase; D000818:Animals; D002648:Child; D002675:Child, Preschool; D005260:Female; D006801:Humans; D007150:Immunohistochemistry; D017403:In Situ Hybridization; D007223:Infant; D007231:Infant, Newborn; D051379:Mice; D008807:Mice, Inbred BALB C; D009154:Mutation; D009447:Neuroblastoma / Q000235:genetics* / Q000378:metabolism*; D020794:Receptor Protein-Tyrosine Kinases / Q000235:genetics / Q000378:metabolism*
chemical_list: C000626173:ALK protein, human; C000626174:Alk protein, mouse; D000077548:Anaplastic Lymphoma Kinase; D020794:Receptor Protein-Tyrosine Kinases