title: Cell culture and Drosophila model systems define three classes of anaplastic lymphoma kinase mutations in neuroblastoma.
abstract: Neuroblastoma is a childhood extracranial solid tumour that is associated with a number of genetic changes. Included in these genetic alterations are mutations in the kinase domain of the anaplastic lymphoma kinase (ALK) receptor tyrosine kinase (RTK), which have been found in both somatic and familial neuroblastoma. In order to treat patients accordingly requires characterisation of these mutations in terms of their response to ALK tyrosine kinase inhibitors (TKIs). Here, we report the identification and characterisation of two novel neuroblastoma ALK mutations (A1099T and R1464STOP), which we have investigated together with several previously reported but uncharacterised ALK mutations (T1087I, D1091N, T1151M, M1166R, F1174I and A1234T). In order to understand the potential role of these ALK mutations in neuroblastoma progression, we have employed cell culture-based systems together with the model organism Drosophila as a readout for ligand-independent activity. Mutation of ALK at position 1174 (F1174I) generates a gain-of-function receptor capable of activating intracellular targets such as ERK (extracellular signal regulated kinase) and STAT3 (signal transducer and activator of transcription 3) in a ligand-independent manner. Analysis of these previously uncharacterised ALK mutants and comparison with ALK(F1174) mutants suggests that ALK mutations observed in neuroblastoma fall into three classes. These classes are: (i) gain-of-function ligand-independent mutations such as ALK(F1174l), (ii) kinase-dead ALK mutants, e.g. ALK(I1250T) (Schönherr et al., 2011a) and (iii) ALK mutations that are ligand-dependent in nature. Irrespective of the nature of the observed ALK mutants, in every case the activity of the mutant ALK receptors could be abrogated by the ALK inhibitor crizotinib (Xalkori/PF-02341066), albeit with differing levels of sensitivity.
authors.affiliation: Department of Molecular Biology, Building 6L, Umeå University, Umeå 901 87, Sweden.
authors.forename: Damini
authors.initials: D
authors.lastname: Chand
authors.forename: Yasuo
authors.initials: Y
authors.lastname: Yamazaki
authors.forename: Kristina
authors.initials: K
authors.lastname: Ruuth
authors.forename: Christina
authors.initials: C
authors.lastname: Schönherr
authors.forename: Tommy
authors.initials: T
authors.lastname: Martinsson
authors.forename: Per
authors.initials: P
authors.lastname: Kogner
authors.forename: Edward F
authors.initials: EF
authors.lastname: Attiyeh
authors.forename: John
authors.initials: J
authors.lastname: Maris
authors.forename: Olena
authors.initials: O
authors.lastname: Morozova
authors.forename: Marco A
authors.initials: MA
authors.lastname: Marra
authors.forename: Miki
authors.initials: M
authors.lastname: Ohira
authors.forename: Akira
authors.initials: A
authors.lastname: Nakagawara
authors.forename: Per-Erik
authors.initials: PE
authors.lastname: Sandström
authors.forename: Ruth H
authors.initials: RH
authors.lastname: Palmer
authors.forename: Bengt
authors.initials: B
authors.lastname: Hallberg
mesh_terms: D000077548:Anaplastic Lymphoma Kinase; D000818:Animals; D018929:Cell Culture Techniques; D049109:Cell Proliferation / Q000187:drug effects; D002471:Cell Transformation, Neoplastic / Q000187:drug effects / Q000473:pathology; D000077547:Crizotinib; D004195:Disease Models, Animal; D004331:Drosophila melanogaster / Q000201:enzymology*; D006801:Humans; D020128:Inhibitory Concentration 50; D050505:Mutant Proteins / Q000378:metabolism; D009154:Mutation / Q000235:genetics*; D016501:Neurites / Q000187:drug effects / Q000378:metabolism / Q000473:pathology; D009447:Neuroblastoma / Q000188:drug therapy / Q000201:enzymology* / Q000235:genetics* / Q000473:pathology; D016716:PC12 Cells; D010641:Phenotype; D010766:Phosphorylation / Q000187:drug effects; D017434:Protein Structure, Tertiary; D011720:Pyrazoles / Q000494:pharmacology / Q000627:therapeutic use; D011725:Pyridines / Q000494:pharmacology / Q000627:therapeutic use; D051381:Rats; D020794:Receptor Protein-Tyrosine Kinases / Q000737:chemistry / Q000235:genetics*
chemical_list: D050505:Mutant Proteins; D011720:Pyrazoles; D011725:Pyridines; D000077547:Crizotinib; C000626173:ALK protein, human; C000626175:Alk protein, rat; D000077548:Anaplastic Lymphoma Kinase; D020794:Receptor Protein-Tyrosine Kinases