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  • 1

    Umeå University Medical Dissertations

    New Series nr. 1235 ISSN: 0346-6612 ISBN: 978-91-7264-708-4

    Investigating the function of

    Anaplastic Lymphoma Kinase

    Emma Vernersson Lindahl

    Department of Medical Bioscience, Pathology

    Umeå University, Sweden

    Umeå 2008

  • 2

    Umeå University Medical Dissertations

    New Series nr. 1235 ISSN: 0346-6612 ISBN: 978-91-7264-708-4

    Cover: wild type mouse embryo, developmental day 13.5. ALK is visualized (red) in part of

    the diencephalon. Immunohistochemistry conducted 041215, picture acquired 041216

    by Emma Vernersson Lindahl

    Copyright© 2008 Emma Vernersson Lindahl

    Printed by Arkitektkopia, Umeå, Sweden

    Umeå 2008

  • 3

    Abstract

    naplastic Lymphoma Kinase (ALK) was discovered in 1994, as a chromosomal

    translocation, t(2;5)(p23;q35), often seen in Anaplastic Large Cell Lymphomas (ALCL).

    Since then ALK has been extensively studied in this disease as well as in different model

    organisms. Due to its expression pattern within the central and peripheral nervous system

    ALK has been implicated in neuronal development. This hypothesis has been further

    strengthened by studies from Drosophila which have shown ALK to have an important role in

    optic lobe development. A recently described ALK mouse knockout model do not indicate an

    essential role for ALK in development, although a potential role within the central nervous

    system was strengthened. This since ALK -/-

    animals has an increased number of progenitor

    cells in the hippocampus and display altered behavior.

    The overall aim of the studies included in this thesis was to elucidate the function of ALK in

    the mouse. As a first step toward this goal we conducted an analysis of ALK mRNA and

    protein expression patterns during development. The strong expression of ALK in neuronal

    structures supports a role for ALK in neuronal development during embryogenesis.

    To further investigate the function of ALK in a physiological context we have developed two

    different ALK knockout strains, the ALK Kinase knockout (KO) and the ALK exon1 KO. The

    only visible phenotype in these strains is a reduction of total body weight which is apparent in

    the ALK -/-

    population when compared to wild type littermates. This size difference seems to

    take place after birth and is not due to an alteration in food consumption. We have also

    extensively studied the ALK Kinase KO with respect to gross development, the

    gastrointestinal canal and the olfactory system. ALK displays a very distinct expression

    pattern within the gastrointestinal canal being confined to enteric neuron precursors during

    embryogenesis and enteric nerves in the adult tissue. From these studies we conclude that

    ALK is not needed for development and viability in mice although it does play a role in

    regulation of body weight via a presently unknown mechanism.

    In addition, we have investigated the relationship between the Drosophila and mouse ALK

    receptor by examining the ability of the Drosophila Alk ligand Jelly-Belly, Jeb, to activate

    mouse ALK. Using different in vivo and in vitro techniques, we have shown that activation of

    mouse ALK cannot be accomplished by Drosophila Jeb. From this study we draw the

    conclusion that during development ligands for the Drosophila and mouse ALK has diverged

    to a level at which they can no longer substitute for each other.

    Keywords: ALK, nervous system, knockout, development, Jelly-Belly

    Author: Emma Vernersson, Department of Medical Bioscience, Pathology, Building 6M, 2 nd

    floor, Umeå University, S-901 85 UMEÅ, Sweden. E-mail:

    [email protected]; phone: +46 (0)90 785 12 96

    A

    mailto:[email protected]

  • 4

    Table of Content

    Papers Included in this Thesis 6 1. Published Articles 6 2. Manuscripts 6

    Abbreviations 7

    Introduction 10 1. Receptor Tyrosine Kinases 10 2. Receptor Tyrosine Kinases in Cancer 10 3. Discovery of the RTK Anaplastic Lymphoma Kinase 11 4. Structural Features of Anaplastic Lymphoma Kinase 11 5. Function of Anaplastic Lymphoma Kinase 13

    5.1 Drosophila Anaplastic lymphoma kinase 13

    5.2 C. elegans Anaplastic Lymphoma Kinase 14

    5.3 Mammalian Anaplastic Lymphoma Kinase 14 6. Anaplastic Lymphoma Kinase in Disease 19

    6.1 Anaplastic Lymphoma Kinase Fusion Proteins in Cancer 19

    6.1.1 Anaplastic Large Cell Lymphoma 19 6.1.2 Mouse Models for ALK Driven ALCL 20

    6.1.3 ALK Fusion Proteins in ALCL 21

    6.1.4 Oncogenic Signaling via ALK Fusion Proteins 22 6.1.5 Inflammatory Myofibroblastic Tumour 27

    6.1.6 Non-Small Cell Lung Cancer 27

    6.1.7 ALK Positive Diffuse B-cell lymphoma 28 6.2 Anaplastic Lymphoma Kinase Overexpression in Cancer 28

    6.3 Anaplastic Lymphoma Kinase Gain-of-Function in Cancer 29

    6.3.1 Neuroblastoma 29 7. Potential Treatments Strategies for ALK Positive Cancers 31

  • 5

    Aim 33 1. Overall Aim 33

    1.1 Specific Aim: Article I 33

    1.2 Specific Aim: Article II 33 1.3 Specific Aim: Manuscript I 33

    1.4 Specific Aim: Manuscript II 33

    Results and Discussion 34 1. Article I: ALK Expression during Development 34 2. Article II: Can Drosophila Jeb activate mouse ALK? 35

    2.1 Why is Drosophila Jeb unable to activate mouse ALK? 36 2.2 What about the LDLa domain? 36

    3. Manuscript I and II: What is the physiological consequence of loss of ALK function? 37

    3.1 The ALK Kinase KO and the ALK exon1 KO 37

    3.2 What function might ALK have during mouse 37 embryogenesis?

    3.3 Is there a role for ALK in the adult olfactory bulb? 38 3.4 The ALK Kinase KO and the ALK exon1 KO are small…

    …but why? 39 3.5 Why are animals smaller? Lessons from the literature. 40

    3.6 Why is ALK not vital for life in the mouse? 40

    Conclusions 42 1. Article I 42 2. Article II 42 3. Manuscript I and II 42

    Acknowledgement 43

    References 45

    Original Papers and Manuscripts 60

  • 6

    Papers Included in this Thesis

    1. Published Articles

    I. Vernersson E, Khoo NK, Henriksson ML, Roos G, Palmer RH, Hallberg B. (2006) Characterization of the expression of the ALK receptor tyrosine kinase in mice.

    Gene Expr. Patterns, 6(5):448-61 *

    II. Yang HL, Eriksson T, Vernersson E, Vigny M, Hallberg B, Palmer RH. (2007) The ligand Jelly Belly (Jeb) activates the Drosophila Alk RTK to drive PC12 cell

    differentiation, but is unable to activate the mouse ALK RTK. J. Exp. Zoolog. B

    Mol. Dev. Evol, 308(3):269-82 **

    2. Manuscripts

    I. Vernersson E, Khoo NK, Roos G, Palmer RH, Hallberg B. (2008) The ALK KO display an idiopathic reduction of total body weight. Manuscript

    II. Vernersson E, Berghard A, Bohm S, Roos G, Palmer RH, Hallberg B. (2008) ALK expression within the olfactory bulb. Manuscript

    *Reprinted from Gene Expression Patterns, 6(5), Vernersson E, Khoo NK, Henriksson ML, Roos G, Palmer RH, Hallberg B, Characterization of the expression of the ALK receptor tyrosine kinase in mice, page 448-461,

    copyright (2006), with permission from Elsevier

    **Reprinted from Journal of experimental zoology. part B, molecular and developmental evolution, 308(3), Yang

    HL, Eriksson T, Vernersson E, Vigny M, Hallberg B, Palmer RH, The ligand Jelly Belly (Jeb) activates the Drosophila Alk RTK to drive PC12 cell differentiation, but is unable to activate the mouse ALK RTK, page 269-

    282, copyright (2007), with permission of Wiley-Liss, Inc. a subsidiary of John Wiley & Sons, Inc.

    http://www.ncbi.nlm.nih.gov/sites/entrez?Db=pubmed&Cmd=Search&Term=%22Vernersson%20E%22%5BAuthor%5D&itool=EntrezSystem2.PEntrez.Pubmed.Pubmed_ResultsPanel.Pubmed_DiscoveryPanel.Pubmed_RVAbstractPlus http://www.ncbi.nlm.nih.gov/sites/entrez?Db=pubmed&Cmd=Search&Term=%22Khoo%20NK%22%5BAuthor%5D&itool=EntrezSystem2.PEntrez.Pubmed.Pubmed_ResultsPanel.Pubmed_DiscoveryPanel.Pubmed_RVAbstractPlus http://www.ncbi.nlm.nih.gov/sites/entrez?Db=pubmed&Cmd=Search&Term=%22Henriksson%20ML%22%5BAuthor%5D&itool=EntrezSystem2.PEntrez.Pubmed.Pubmed_ResultsPanel.Pubmed_DiscoveryPanel.Pubmed_RVAbstractPlus http://www.ncbi.nlm.nih.gov/sites/entrez?Db=pubmed&Cmd=Search&Term=%22Roos%20G%22%5BAuthor%5D&itool=EntrezSystem2.PEntrez.Pubmed.Pubmed_ResultsPanel.Pubmed_DiscoveryPanel.Pubmed_RVAbstractPlus http://www.ncbi.nlm.nih.gov/sites/entrez?Db=pubmed&Cmd=Search&Term=%22Palmer%20RH%22%5BAuthor%5D&itool=EntrezSystem2.PEntrez.Pubmed.Pubmed_ResultsPanel.Pubmed_D