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Cell and Drosophila model systems define three classes of anaplastic lymphoma kinase mutations in neuroblastoma
Umeå University, Faculty of Medicine, Department of Molecular Biology (Faculty of Medicine). (Hallberg)
Umeå University, Faculty of Science and Technology, Department of Molecular Biology (Faculty of Science and Technology).
Umeå University, Faculty of Medicine, Department of Molecular Biology (Faculty of Medicine). (Hallberg)
Umeå University, Faculty of Medicine, Department of Molecular Biology (Faculty of Medicine). (Hallberg)
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2013 (English)In: Disease models & mechanisms, ISSN 1754-8411, Vol. 6, no 2, 373-382 p.Article in journal (Refereed) Published
Abstract [en]

Neuroblastoma is a childhood extracranial solid tumor which 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 required 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 1464STOP) 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 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 are: (i) gain-of-function ligand independent mutations such as ALK(F1174), (ii) kinase-dead ALK mutants, e.g. ALK(I1250T)(Schonherr et al 2011a) or (iii) ALK mutations which 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 (PF-02341066, Xalkori), albeit with differing levels of sensitivity.

Place, publisher, year, edition, pages
Cambridge, UK: The Company of Biologists Ltd , 2013. Vol. 6, no 2, 373-382 p.
National Category
Cancer and Oncology
URN: urn:nbn:se:umu:diva-61265DOI: 10.1242/dmm.010348PubMedID: 23104988OAI: diva2:565393
Available from: 2012-11-07 Created: 2012-11-07 Last updated: 2015-09-01Bibliographically approved
In thesis
1. Mechanistic Implications and Characterization of Anaplastic Lymphoma Kinase (ALK) mutations in Neuroblastoma
Open this publication in new window or tab >>Mechanistic Implications and Characterization of Anaplastic Lymphoma Kinase (ALK) mutations in Neuroblastoma
2015 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Anaplastic lymphoma kinase (ALK) is a receptor tyrosine kinase that was first reported as a fusion partner of nucleophosmin in Anaplastic large cell lymphoma in 1994. ALK is involved in myriad of cancers including neuroblastoma which is the most common extracranial solid tumor affecting young children. It arises in the neural crest cells of sympathetic nervous system origin and is responsible for 12% of all childhood cancer deaths. Several point mutations in ALK have been described in both familial and sporadic neuroblastoma.

With the aim to understand the role of ALK in neuroblastoma further, we investigated the point mutations in ALK reported in patients. Using cell culture based methods and Drosophila as a model organism; we first characterized these mutations under three broad categories: 1) Ligand independent mutations that were constitutively active, 2) Kinase dead mutation and 3) Ligand dependent mutations that behaved as inducible wild type. Further, to understand the activation mechanism of ALK, we constructed mutations that could potentially alter ALK’s conformation based on the available crystal structure. From the data generated, we were able to provide a new perspective to the activation of full length ALK receptor. This was more in line with activation mechanism of insulin receptor and different from that suggested for ALK fusion protein. From a clinical point of view, all the mutations in the study were blocked to different degrees using the ALK inhibitor, crizotinib. Lastly, we identified potential downstream targets of ALK using phosphoproteomics. From the various targets identified, we focused on STAT3 and confirmed its role as a mediator in ALK initiated MYCN transcription. We showed that STAT3 inhibition led to reduction of MYCN levels and thereby identifying it as a potential therapeutic target in neuroblastoma. Overall, our study highlights clinical relevance of ALK mutations in neuroblastoma and from a basic biology viewpoint; it reveals important mechanistic insight into receptor activation.

Place, publisher, year, edition, pages
Umeå: Umeå Universitet, 2015. 79 p.
Umeå University medical dissertations, ISSN 0346-6612 ; 1709
neuroblastoma, ALK, crizotinib, receptor tyrosine kinase
National Category
Cell and Molecular Biology
Research subject
Molecular Biology
urn:nbn:se:umu:diva-106663 (URN)978-91-7601-254-3 (ISBN)
Public defence
2015-10-02, KBC-huset, Stora hörsalen, KKB3B1, Umeå universitet, Umeå, 09:00 (English)
Available from: 2015-09-04 Created: 2015-07-28 Last updated: 2015-09-04Bibliographically approved

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Chand, DaminiYamazaki, YasuoRuuth, KristinaSchönherr, ChristinaSandström, Per-ErikPalmer, Ruth HHallberg, Bengt
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Department of Molecular Biology (Faculty of Medicine)Department of Molecular Biology (Faculty of Science and Technology)Paediatrics
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