The role of histone demethylase KDM4B in Myc signaling in neuroblastoma.
BACKGROUND
Epigenetic alterations, such as histone methylation, modulate Myc signaling, a pathway central to oncogenesis. We investigated the role of the histone demethylase KDM4B in N-Myc-mediated neuroblastoma pathogenesis.


METHODS
Spearman correlation was performed to correlate MYCN and KDM4B expression. RNA interference, microarray analysis, gene set enrichment analysis, and real-time polymerase chain reaction were used to define the functions of KDM4B. Immunoprecipitation and immunofluorescence were used to assess protein-protein interactions between N-Myc and KDM4B. Chromatin immunoprecipitation was used to assess the binding of Myc targets. Constitutive and inducible lentiviral-mediated KDM4B knockdown with shRNA was used to assess the effects on tumor growth. Kaplan-Meier survival analysis was used to assess the prognostic value of KDM4B expression. All statistical tests were two-sided.


RESULTS
KDM4B and MYCN expression were found to be statistically significantly correlated in a variety of cancers, including neuroblastoma (R = 0.396, P < .001). Functional studies demonstrated that KDM4B regulates the Myc pathway. N-Myc was found to physically interact with and recruit KDM4B. KDM4B was found to regulate neuroblastoma cell proliferation and differentiation in vitro and xenograft growth in vivo (5 mice/group, two-tailed t test, P ≤ 0.001). Finally, together with MYCN amplification, KDM4B was found to stratify a subgroup of poor-prognosis patients (122 case patients, P < .001).


CONCLUSIONS
Our findings provide insight into the epigenetic regulation of Myc via histone demethylation and proof-of-concept for inhibition of histone demethylases to target Myc signaling in cancers such as neuroblastoma.
Department of Surgery (JY, AMA, DH, PHC, CLM, AMD) and Department of Bioinformatics (CQ), St. Jude Children's Research Hospital, Memphis, TN; Yale Cardiovascular Research Center, Yale School of Medicine, New Haven, CT (YW); Department of Oncology, University College London Cancer Institute, London, UK (ACN); Texas Children's Cancer Center, Department of Pediatrics, Baylor College of Medicine, Houston, TX (JMS); Division of Biomedical Research, Foundation of Research and Technology-Hellas, Institute of Molecular Biology and Biotechnology, Ioannina, Greece (TF); Laboratory of Biological Chemistry, Medical School, University of Ioannina, Ioannina, Greece (TF); Department of Oncogenomics, Academic Medical Center, University of Amsterdam, Amsterdam, the Netherlands (JK, RV); Kinki University Faculty of Medicine, Osaka-Sayama, Osaka, Japan (HO); Molecular Oncology Laboratories, Weatherall Institute of Molecular Medicine, University of Oxford, John Radcliffe Hospital, Oxford, UK (ALH).
Jun
J
Yang
Department of Surgery (JY, AMA, DH, PHC, CLM, AMD) and Department of Bioinformatics (CQ), St. Jude Children's Research Hospital, Memphis, TN; Yale Cardiovascular Research Center, Yale School of Medicine, New Haven, CT (YW); Department of Oncology, University College London Cancer Institute, London, UK (ACN); Texas Children's Cancer Center, Department of Pediatrics, Baylor College of Medicine, Houston, TX (JMS); Division of Biomedical Research, Foundation of Research and Technology-Hellas, Institute of Molecular Biology and Biotechnology, Ioannina, Greece (TF); Laboratory of Biological Chemistry, Medical School, University of Ioannina, Ioannina, Greece (TF); Department of Oncogenomics, Academic Medical Center, University of Amsterdam, Amsterdam, the Netherlands (JK, RV); Kinki University Faculty of Medicine, Osaka-Sayama, Osaka, Japan (HO); Molecular Oncology Laboratories, Weatherall Institute of Molecular Medicine, University of Oxford, John Radcliffe Hospital, Oxford, UK (ALH).
Alaa M
AM
AlTahan
Department of Surgery (JY, AMA, DH, PHC, CLM, AMD) and Department of Bioinformatics (CQ), St. Jude Children's Research Hospital, Memphis, TN; Yale Cardiovascular Research Center, Yale School of Medicine, New Haven, CT (YW); Department of Oncology, University College London Cancer Institute, London, UK (ACN); Texas Children's Cancer Center, Department of Pediatrics, Baylor College of Medicine, Houston, TX (JMS); Division of Biomedical Research, Foundation of Research and Technology-Hellas, Institute of Molecular Biology and Biotechnology, Ioannina, Greece (TF); Laboratory of Biological Chemistry, Medical School, University of Ioannina, Ioannina, Greece (TF); Department of Oncogenomics, Academic Medical Center, University of Amsterdam, Amsterdam, the Netherlands (JK, RV); Kinki University Faculty of Medicine, Osaka-Sayama, Osaka, Japan (HO); Molecular Oncology Laboratories, Weatherall Institute of Molecular Medicine, University of Oxford, John Radcliffe Hospital, Oxford, UK (ALH).
Dongli
D
Hu
Department of Surgery (JY, AMA, DH, PHC, CLM, AMD) and Department of Bioinformatics (CQ), St. Jude Children's Research Hospital, Memphis, TN; Yale Cardiovascular Research Center, Yale School of Medicine, New Haven, CT (YW); Department of Oncology, University College London Cancer Institute, London, UK (ACN); Texas Children's Cancer Center, Department of Pediatrics, Baylor College of Medicine, Houston, TX (JMS); Division of Biomedical Research, Foundation of Research and Technology-Hellas, Institute of Molecular Biology and Biotechnology, Ioannina, Greece (TF); Laboratory of Biological Chemistry, Medical School, University of Ioannina, Ioannina, Greece (TF); Department of Oncogenomics, Academic Medical Center, University of Amsterdam, Amsterdam, the Netherlands (JK, RV); Kinki University Faculty of Medicine, Osaka-Sayama, Osaka, Japan (HO); Molecular Oncology Laboratories, Weatherall Institute of Molecular Medicine, University of Oxford, John Radcliffe Hospital, Oxford, UK (ALH).
Yingdi
Y
Wang
Department of Surgery (JY, AMA, DH, PHC, CLM, AMD) and Department of Bioinformatics (CQ), St. Jude Children's Research Hospital, Memphis, TN; Yale Cardiovascular Research Center, Yale School of Medicine, New Haven, CT (YW); Department of Oncology, University College London Cancer Institute, London, UK (ACN); Texas Children's Cancer Center, Department of Pediatrics, Baylor College of Medicine, Houston, TX (JMS); Division of Biomedical Research, Foundation of Research and Technology-Hellas, Institute of Molecular Biology and Biotechnology, Ioannina, Greece (TF); Laboratory of Biological Chemistry, Medical School, University of Ioannina, Ioannina, Greece (TF); Department of Oncogenomics, Academic Medical Center, University of Amsterdam, Amsterdam, the Netherlands (JK, RV); Kinki University Faculty of Medicine, Osaka-Sayama, Osaka, Japan (HO); Molecular Oncology Laboratories, Weatherall Institute of Molecular Medicine, University of Oxford, John Radcliffe Hospital, Oxford, UK (ALH).
Pei-Hsin
PH
Cheng
Department of Surgery (JY, AMA, DH, PHC, CLM, AMD) and Department of Bioinformatics (CQ), St. Jude Children's Research Hospital, Memphis, TN; Yale Cardiovascular Research Center, Yale School of Medicine, New Haven, CT (YW); Department of Oncology, University College London Cancer Institute, London, UK (ACN); Texas Children's Cancer Center, Department of Pediatrics, Baylor College of Medicine, Houston, TX (JMS); Division of Biomedical Research, Foundation of Research and Technology-Hellas, Institute of Molecular Biology and Biotechnology, Ioannina, Greece (TF); Laboratory of Biological Chemistry, Medical School, University of Ioannina, Ioannina, Greece (TF); Department of Oncogenomics, Academic Medical Center, University of Amsterdam, Amsterdam, the Netherlands (JK, RV); Kinki University Faculty of Medicine, Osaka-Sayama, Osaka, Japan (HO); Molecular Oncology Laboratories, Weatherall Institute of Molecular Medicine, University of Oxford, John Radcliffe Hospital, Oxford, UK (ALH).
Christopher L
CL
Morton
Department of Surgery (JY, AMA, DH, PHC, CLM, AMD) and Department of Bioinformatics (CQ), St. Jude Children's Research Hospital, Memphis, TN; Yale Cardiovascular Research Center, Yale School of Medicine, New Haven, CT (YW); Department of Oncology, University College London Cancer Institute, London, UK (ACN); Texas Children's Cancer Center, Department of Pediatrics, Baylor College of Medicine, Houston, TX (JMS); Division of Biomedical Research, Foundation of Research and Technology-Hellas, Institute of Molecular Biology and Biotechnology, Ioannina, Greece (TF); Laboratory of Biological Chemistry, Medical School, University of Ioannina, Ioannina, Greece (TF); Department of Oncogenomics, Academic Medical Center, University of Amsterdam, Amsterdam, the Netherlands (JK, RV); Kinki University Faculty of Medicine, Osaka-Sayama, Osaka, Japan (HO); Molecular Oncology Laboratories, Weatherall Institute of Molecular Medicine, University of Oxford, John Radcliffe Hospital, Oxford, UK (ALH).
Chunxu
C
Qu
Department of Surgery (JY, AMA, DH, PHC, CLM, AMD) and Department of Bioinformatics (CQ), St. Jude Children's Research Hospital, Memphis, TN; Yale Cardiovascular Research Center, Yale School of Medicine, New Haven, CT (YW); Department of Oncology, University College London Cancer Institute, London, UK (ACN); Texas Children's Cancer Center, Department of Pediatrics, Baylor College of Medicine, Houston, TX (JMS); Division of Biomedical Research, Foundation of Research and Technology-Hellas, Institute of Molecular Biology and Biotechnology, Ioannina, Greece (TF); Laboratory of Biological Chemistry, Medical School, University of Ioannina, Ioannina, Greece (TF); Department of Oncogenomics, Academic Medical Center, University of Amsterdam, Amsterdam, the Netherlands (JK, RV); Kinki University Faculty of Medicine, Osaka-Sayama, Osaka, Japan (HO); Molecular Oncology Laboratories, Weatherall Institute of Molecular Medicine, University of Oxford, John Radcliffe Hospital, Oxford, UK (ALH).
Amit C
AC
Nathwani
Department of Surgery (JY, AMA, DH, PHC, CLM, AMD) and Department of Bioinformatics (CQ), St. Jude Children's Research Hospital, Memphis, TN; Yale Cardiovascular Research Center, Yale School of Medicine, New Haven, CT (YW); Department of Oncology, University College London Cancer Institute, London, UK (ACN); Texas Children's Cancer Center, Department of Pediatrics, Baylor College of Medicine, Houston, TX (JMS); Division of Biomedical Research, Foundation of Research and Technology-Hellas, Institute of Molecular Biology and Biotechnology, Ioannina, Greece (TF); Laboratory of Biological Chemistry, Medical School, University of Ioannina, Ioannina, Greece (TF); Department of Oncogenomics, Academic Medical Center, University of Amsterdam, Amsterdam, the Netherlands (JK, RV); Kinki University Faculty of Medicine, Osaka-Sayama, Osaka, Japan (HO); Molecular Oncology Laboratories, Weatherall Institute of Molecular Medicine, University of Oxford, John Radcliffe Hospital, Oxford, UK (ALH).
Jason M
JM
Shohet
Department of Surgery (JY, AMA, DH, PHC, CLM, AMD) and Department of Bioinformatics (CQ), St. Jude Children's Research Hospital, Memphis, TN; Yale Cardiovascular Research Center, Yale School of Medicine, New Haven, CT (YW); Department of Oncology, University College London Cancer Institute, London, UK (ACN); Texas Children's Cancer Center, Department of Pediatrics, Baylor College of Medicine, Houston, TX (JMS); Division of Biomedical Research, Foundation of Research and Technology-Hellas, Institute of Molecular Biology and Biotechnology, Ioannina, Greece (TF); Laboratory of Biological Chemistry, Medical School, University of Ioannina, Ioannina, Greece (TF); Department of Oncogenomics, Academic Medical Center, University of Amsterdam, Amsterdam, the Netherlands (JK, RV); Kinki University Faculty of Medicine, Osaka-Sayama, Osaka, Japan (HO); Molecular Oncology Laboratories, Weatherall Institute of Molecular Medicine, University of Oxford, John Radcliffe Hospital, Oxford, UK (ALH).
Theodore
T
Fotsis
Department of Surgery (JY, AMA, DH, PHC, CLM, AMD) and Department of Bioinformatics (CQ), St. Jude Children's Research Hospital, Memphis, TN; Yale Cardiovascular Research Center, Yale School of Medicine, New Haven, CT (YW); Department of Oncology, University College London Cancer Institute, London, UK (ACN); Texas Children's Cancer Center, Department of Pediatrics, Baylor College of Medicine, Houston, TX (JMS); Division of Biomedical Research, Foundation of Research and Technology-Hellas, Institute of Molecular Biology and Biotechnology, Ioannina, Greece (TF); Laboratory of Biological Chemistry, Medical School, University of Ioannina, Ioannina, Greece (TF); Department of Oncogenomics, Academic Medical Center, University of Amsterdam, Amsterdam, the Netherlands (JK, RV); Kinki University Faculty of Medicine, Osaka-Sayama, Osaka, Japan (HO); Molecular Oncology Laboratories, Weatherall Institute of Molecular Medicine, University of Oxford, John Radcliffe Hospital, Oxford, UK (ALH).
Jan
J
Koster
Department of Surgery (JY, AMA, DH, PHC, CLM, AMD) and Department of Bioinformatics (CQ), St. Jude Children's Research Hospital, Memphis, TN; Yale Cardiovascular Research Center, Yale School of Medicine, New Haven, CT (YW); Department of Oncology, University College London Cancer Institute, London, UK (ACN); Texas Children's Cancer Center, Department of Pediatrics, Baylor College of Medicine, Houston, TX (JMS); Division of Biomedical Research, Foundation of Research and Technology-Hellas, Institute of Molecular Biology and Biotechnology, Ioannina, Greece (TF); Laboratory of Biological Chemistry, Medical School, University of Ioannina, Ioannina, Greece (TF); Department of Oncogenomics, Academic Medical Center, University of Amsterdam, Amsterdam, the Netherlands (JK, RV); Kinki University Faculty of Medicine, Osaka-Sayama, Osaka, Japan (HO); Molecular Oncology Laboratories, Weatherall Institute of Molecular Medicine, University of Oxford, John Radcliffe Hospital, Oxford, UK (ALH).
Rogier
R
Versteeg
Department of Surgery (JY, AMA, DH, PHC, CLM, AMD) and Department of Bioinformatics (CQ), St. Jude Children's Research Hospital, Memphis, TN; Yale Cardiovascular Research Center, Yale School of Medicine, New Haven, CT (YW); Department of Oncology, University College London Cancer Institute, London, UK (ACN); Texas Children's Cancer Center, Department of Pediatrics, Baylor College of Medicine, Houston, TX (JMS); Division of Biomedical Research, Foundation of Research and Technology-Hellas, Institute of Molecular Biology and Biotechnology, Ioannina, Greece (TF); Laboratory of Biological Chemistry, Medical School, University of Ioannina, Ioannina, Greece (TF); Department of Oncogenomics, Academic Medical Center, University of Amsterdam, Amsterdam, the Netherlands (JK, RV); Kinki University Faculty of Medicine, Osaka-Sayama, Osaka, Japan (HO); Molecular Oncology Laboratories, Weatherall Institute of Molecular Medicine, University of Oxford, John Radcliffe Hospital, Oxford, UK (ALH).
Hitoshi
H
Okada
Department of Surgery (JY, AMA, DH, PHC, CLM, AMD) and Department of Bioinformatics (CQ), St. Jude Children's Research Hospital, Memphis, TN; Yale Cardiovascular Research Center, Yale School of Medicine, New Haven, CT (YW); Department of Oncology, University College London Cancer Institute, London, UK (ACN); Texas Children's Cancer Center, Department of Pediatrics, Baylor College of Medicine, Houston, TX (JMS); Division of Biomedical Research, Foundation of Research and Technology-Hellas, Institute of Molecular Biology and Biotechnology, Ioannina, Greece (TF); Laboratory of Biological Chemistry, Medical School, University of Ioannina, Ioannina, Greece (TF); Department of Oncogenomics, Academic Medical Center, University of Amsterdam, Amsterdam, the Netherlands (JK, RV); Kinki University Faculty of Medicine, Osaka-Sayama, Osaka, Japan (HO); Molecular Oncology Laboratories, Weatherall Institute of Molecular Medicine, University of Oxford, John Radcliffe Hospital, Oxford, UK (ALH).
Adrian L
AL
Harris
Department of Surgery (JY, AMA, DH, PHC, CLM, AMD) and Department of Bioinformatics (CQ), St. Jude Children's Research Hospital, Memphis, TN; Yale Cardiovascular Research Center, Yale School of Medicine, New Haven, CT (YW); Department of Oncology, University College London Cancer Institute, London, UK (ACN); Texas Children's Cancer Center, Department of Pediatrics, Baylor College of Medicine, Houston, TX (JMS); Division of Biomedical Research, Foundation of Research and Technology-Hellas, Institute of Molecular Biology and Biotechnology, Ioannina, Greece (TF); Laboratory of Biological Chemistry, Medical School, University of Ioannina, Ioannina, Greece (TF); Department of Oncogenomics, Academic Medical Center, University of Amsterdam, Amsterdam, the Netherlands (JK, RV); Kinki University Faculty of Medicine, Osaka-Sayama, Osaka, Japan (HO); Molecular Oncology Laboratories, Weatherall Institute of Molecular Medicine, University of Oxford, John Radcliffe Hospital, Oxford, UK (ALH). andrew.davidoff@stjude.org.
Andrew M
AM
Davidoff
D000818:Animals; D002454:Cell Differentiation; D045744:Cell Line, Tumor; D049109:Cell Proliferation; D047369:Chromatin Immunoprecipitation; D019175:DNA Methylation; D015972:Gene Expression Regulation, Neoplastic; D064593:Heterografts; D006801:Humans; D056484:Jumonji Domain-Containing Histone Demethylases / Q000378:metabolism*; D053208:Kaplan-Meier Estimate; D051379:Mice; D009447:Neuroblastoma / Q000235:genetics / Q000378:metabolism*; D011379:Prognosis; D040081:Protein Array Analysis; D016271:Proto-Oncogene Proteins c-myc / Q000378:metabolism*; D060888:Real-Time Polymerase Chain Reaction
C489427:MYC protein, human; D016271:Proto-Oncogene Proteins c-myc; D056484:Jumonji Domain-Containing Histone Demethylases; C523637:KDM4B protein, human