Druggable targets in pediatric neurocutaneous melanocytosis: Molecular and drug sensitivity studies in xenograft and ex vivo tumor cell culture to identify agents for therapy.
BACKGROUND
Neurocutaneous melanocytosis (NCM) is a rare congenital disorder that presents with pigmented cell lesions of the brain or leptomeninges in children with large or multiple congenital melanocytic nevi. Although the exact pathological processes involved are currently unclear, NCM appears to arise from an abnormal development of melanoblasts or melanocyte precursors. Currently, it has an extremely poor prognosis due to rapid disease progression and lack of effective treatment modalities.


METHODS
In this study, we report on an experimental approach to examining NCM cells by establishing subcutaneous tumors in nude mice, which can be further expanded for conducting molecular and drug sensitivity experiments.


RESULTS
Analysis of the NRAS gene-coding sequences of an established NCM cell line (YP-MEL) and NCM patient cells revealed heterogeneity in NRAS Q61K that activated mutation and possibly consequential differential sensitivity to MEK inhibition. Gene expression studies were performed to compare the molecular profiles of NCM cells with normal skin fibroblasts. In vitro cytotoxicity screens of libraries of targeted small-molecule inhibitors revealed prospective agents for further evaluation.


CONCLUSIONS
Our studies provide an experimental platform for the generation of NCM cells for preclinical studies and the production of molecular and in vitro data with which to identify druggable targets for the treatment.
Division of Pediatric Oncology, Alberta Children's Hospital and POETIC Laboratory for Preclinical and Drug Discovery Studies, University of Calgary, Calgary, Alberta, Canada (Y.R., A.J., X.L., R.A., A.N.); Department of Biological Sciences, University of Lethbridge, Lethbridge, Canada (A.K., O.K.); Department of Pathology, Yokohama City University School of Medicine and Division of Diagnostic Pathology, Tokyo Women's Medical University Hospital, Tokyo, Japan (Y.N.); Department of Pathology and Laboratory Medicine, Alberta Children's Hospital and Calgary Laboratory Services, Calgary, Canada (J.R.W., A.P.); Department of Neurology, Alberta Children's Hospital, Calgary, Canada (A.K.).
Yibing
Y
Ruan
Division of Pediatric Oncology, Alberta Children's Hospital and POETIC Laboratory for Preclinical and Drug Discovery Studies, University of Calgary, Calgary, Alberta, Canada (Y.R., A.J., X.L., R.A., A.N.); Department of Biological Sciences, University of Lethbridge, Lethbridge, Canada (A.K., O.K.); Department of Pathology, Yokohama City University School of Medicine and Division of Diagnostic Pathology, Tokyo Women's Medical University Hospital, Tokyo, Japan (Y.N.); Department of Pathology and Laboratory Medicine, Alberta Children's Hospital and Calgary Laboratory Services, Calgary, Canada (J.R.W., A.P.); Department of Neurology, Alberta Children's Hospital, Calgary, Canada (A.K.).
Anna
A
Kovalchuk
Division of Pediatric Oncology, Alberta Children's Hospital and POETIC Laboratory for Preclinical and Drug Discovery Studies, University of Calgary, Calgary, Alberta, Canada (Y.R., A.J., X.L., R.A., A.N.); Department of Biological Sciences, University of Lethbridge, Lethbridge, Canada (A.K., O.K.); Department of Pathology, Yokohama City University School of Medicine and Division of Diagnostic Pathology, Tokyo Women's Medical University Hospital, Tokyo, Japan (Y.N.); Department of Pathology and Laboratory Medicine, Alberta Children's Hospital and Calgary Laboratory Services, Calgary, Canada (J.R.W., A.P.); Department of Neurology, Alberta Children's Hospital, Calgary, Canada (A.K.).
Aarthi
A
Jayanthan
Division of Pediatric Oncology, Alberta Children's Hospital and POETIC Laboratory for Preclinical and Drug Discovery Studies, University of Calgary, Calgary, Alberta, Canada (Y.R., A.J., X.L., R.A., A.N.); Department of Biological Sciences, University of Lethbridge, Lethbridge, Canada (A.K., O.K.); Department of Pathology, Yokohama City University School of Medicine and Division of Diagnostic Pathology, Tokyo Women's Medical University Hospital, Tokyo, Japan (Y.N.); Department of Pathology and Laboratory Medicine, Alberta Children's Hospital and Calgary Laboratory Services, Calgary, Canada (J.R.W., A.P.); Department of Neurology, Alberta Children's Hospital, Calgary, Canada (A.K.).
Xueqing
X
Lun
Division of Pediatric Oncology, Alberta Children's Hospital and POETIC Laboratory for Preclinical and Drug Discovery Studies, University of Calgary, Calgary, Alberta, Canada (Y.R., A.J., X.L., R.A., A.N.); Department of Biological Sciences, University of Lethbridge, Lethbridge, Canada (A.K., O.K.); Department of Pathology, Yokohama City University School of Medicine and Division of Diagnostic Pathology, Tokyo Women's Medical University Hospital, Tokyo, Japan (Y.N.); Department of Pathology and Laboratory Medicine, Alberta Children's Hospital and Calgary Laboratory Services, Calgary, Canada (J.R.W., A.P.); Department of Neurology, Alberta Children's Hospital, Calgary, Canada (A.K.).
Yoji
Y
Nagashima
Division of Pediatric Oncology, Alberta Children's Hospital and POETIC Laboratory for Preclinical and Drug Discovery Studies, University of Calgary, Calgary, Alberta, Canada (Y.R., A.J., X.L., R.A., A.N.); Department of Biological Sciences, University of Lethbridge, Lethbridge, Canada (A.K., O.K.); Department of Pathology, Yokohama City University School of Medicine and Division of Diagnostic Pathology, Tokyo Women's Medical University Hospital, Tokyo, Japan (Y.N.); Department of Pathology and Laboratory Medicine, Alberta Children's Hospital and Calgary Laboratory Services, Calgary, Canada (J.R.W., A.P.); Department of Neurology, Alberta Children's Hospital, Calgary, Canada (A.K.).
Olga
O
Kovalchuk
Division of Pediatric Oncology, Alberta Children's Hospital and POETIC Laboratory for Preclinical and Drug Discovery Studies, University of Calgary, Calgary, Alberta, Canada (Y.R., A.J., X.L., R.A., A.N.); Department of Biological Sciences, University of Lethbridge, Lethbridge, Canada (A.K., O.K.); Department of Pathology, Yokohama City University School of Medicine and Division of Diagnostic Pathology, Tokyo Women's Medical University Hospital, Tokyo, Japan (Y.N.); Department of Pathology and Laboratory Medicine, Alberta Children's Hospital and Calgary Laboratory Services, Calgary, Canada (J.R.W., A.P.); Department of Neurology, Alberta Children's Hospital, Calgary, Canada (A.K.).
James R
JR
Wright
Division of Pediatric Oncology, Alberta Children's Hospital and POETIC Laboratory for Preclinical and Drug Discovery Studies, University of Calgary, Calgary, Alberta, Canada (Y.R., A.J., X.L., R.A., A.N.); Department of Biological Sciences, University of Lethbridge, Lethbridge, Canada (A.K., O.K.); Department of Pathology, Yokohama City University School of Medicine and Division of Diagnostic Pathology, Tokyo Women's Medical University Hospital, Tokyo, Japan (Y.N.); Department of Pathology and Laboratory Medicine, Alberta Children's Hospital and Calgary Laboratory Services, Calgary, Canada (J.R.W., A.P.); Department of Neurology, Alberta Children's Hospital, Calgary, Canada (A.K.).
Alfredo
A
Pinto
Division of Pediatric Oncology, Alberta Children's Hospital and POETIC Laboratory for Preclinical and Drug Discovery Studies, University of Calgary, Calgary, Alberta, Canada (Y.R., A.J., X.L., R.A., A.N.); Department of Biological Sciences, University of Lethbridge, Lethbridge, Canada (A.K., O.K.); Department of Pathology, Yokohama City University School of Medicine and Division of Diagnostic Pathology, Tokyo Women's Medical University Hospital, Tokyo, Japan (Y.N.); Department of Pathology and Laboratory Medicine, Alberta Children's Hospital and Calgary Laboratory Services, Calgary, Canada (J.R.W., A.P.); Department of Neurology, Alberta Children's Hospital, Calgary, Canada (A.K.).
Adam
A
Kirton
Division of Pediatric Oncology, Alberta Children's Hospital and POETIC Laboratory for Preclinical and Drug Discovery Studies, University of Calgary, Calgary, Alberta, Canada (Y.R., A.J., X.L., R.A., A.N.); Department of Biological Sciences, University of Lethbridge, Lethbridge, Canada (A.K., O.K.); Department of Pathology, Yokohama City University School of Medicine and Division of Diagnostic Pathology, Tokyo Women's Medical University Hospital, Tokyo, Japan (Y.N.); Department of Pathology and Laboratory Medicine, Alberta Children's Hospital and Calgary Laboratory Services, Calgary, Canada (J.R.W., A.P.); Department of Neurology, Alberta Children's Hospital, Calgary, Canada (A.K.).
Ronald
R
Anderson
Division of Pediatric Oncology, Alberta Children's Hospital and POETIC Laboratory for Preclinical and Drug Discovery Studies, University of Calgary, Calgary, Alberta, Canada (Y.R., A.J., X.L., R.A., A.N.); Department of Biological Sciences, University of Lethbridge, Lethbridge, Canada (A.K., O.K.); Department of Pathology, Yokohama City University School of Medicine and Division of Diagnostic Pathology, Tokyo Women's Medical University Hospital, Tokyo, Japan (Y.N.); Department of Pathology and Laboratory Medicine, Alberta Children's Hospital and Calgary Laboratory Services, Calgary, Canada (J.R.W., A.P.); Department of Neurology, Alberta Children's Hospital, Calgary, Canada (A.K.).
Aru
A
Narendran
D000818:Animals; D000970:Antineoplastic Agents / Q000008:administration & dosage* / Q000627:therapeutic use; D002675:Child, Preschool; D020558:GTP Phosphohydrolases / Q000235:genetics; D064593:Heterografts / Q000187:drug effects / Q000473:pathology; D006801:Humans; D008297:Male; D008548:Melanosis / Q000188:drug therapy* / Q000235:genetics* / Q000473:pathology; D008565:Membrane Proteins / Q000235:genetics; D051379:Mice; D008819:Mice, Nude; D009154:Mutation; D020752:Neurocutaneous Syndromes / Q000188:drug therapy* / Q000235:genetics* / Q000473:pathology; D015398:Signal Transduction / Q000187:drug effects; D054852:Small Molecule Libraries / Q000008:administration & dosage / Q000627:therapeutic use; D059467:Transcriptome; D014407:Tumor Cells, Cultured; D023041:Xenograft Model Antitumor Assays / Q000379:methods*
D000970:Antineoplastic Agents; D008565:Membrane Proteins; D054852:Small Molecule Libraries; D020558:GTP Phosphohydrolases; C579846:NRAS protein, human
molecular profiling; neurocutaneous melanocytosis; targeted therapy