title: Downregulation of c-Myc is involved in TLR3-mediated tumor death of neuroblastoma xenografts.
abstract: Neuroblastoma (NB) is the deadliest pediatric solid tumor due to its pleomorphic molecular characteristics. In the innate immune system, toll-like receptor 3 (TLR3) recognizes viral double-stranded RNAs to initiate immune signaling. Positive TLR3 expression indicates a favorable prognosis in NB patients, and is associated with MYCN-non-amplified. However, TLR3-mediated innate immune responses remain elusive in NB. In this study, we attempted to dissect the molecular mechanism underlying TLR3-agonist polyinosinic-polycytidylic acid [poly(I:C)] treatment in NB in vivo. We established NB xenograft models in non-obese diabetic/severe combined immunodeficiency (NOD/SCID) mice with MYCN-amplified SK-N-DZ (DZ) cells or MYCN-non-amplified SK-N-AS (AS) cells. Poly(I:C) treatment led to significant tumor regression in AS xenografts, but not in DZ xenografts. Through immunohistochemical analysis, significant suppression of tumor proliferation, downregulation of c-Myc expression, and upregulation of TLR3 expression were found in the treatment group. Poly(I:C) inducing activation of TLR3/IRF3-mediated innate immunity associated with downregulation of c-Myc can be found in MYCN-non-amplified SK-N-AS cells, but not in MYCN-amplified BE(2)-M17 cells. Knockdown of TLR3 disturbed poly(I:C)-induced suppression of c-Myc and upregulation of p-IRF3 in AS cells. Furthermore, poly(I:C) treatment upregulated active NF-κB, mitochondrial antioxidant manganese superoxide dismutase and 8-hydroxydeoxyguanosine, which works with reactive oxygen species (ROS) generation and DNA damage. Upregulation of active caspase 3 and cleaved poly [ADP-ribose] polymerase 1 were found in poly(I:C)-treated AS xenografts, which indicates the induction of apoptosis. Thus, our results suggest that c-Myc overexpression may increase sensitivity to poly(I:C)-induced tumor growth arrest and ROS-mediated apoptosis in NB. This study demonstrates that c-Myc protein expression has an important role in TLR3-induced innate immune responses, providing future treatment recommendations.
authors.affiliation: Department of Pediatric Surgery, Kaohsiung Chang Gung Memorial Hospital, Chang Gung University College of Medicine, Kaohsiung, Taiwan.|Department of Medical Research, Kaohsiung Chang Gung Memorial Hospital, Chang Gung University College of Medicine, Kaohsiung, Taiwan.
authors.forename: Li-Ling
authors.initials: LL
authors.lastname: Lin
authors.affiliation: Biobank and Tissue Bank, Kaohsiung Chang Gung Memorial Hospital, Kaohsiung, Taiwan.|Department of Pathology, Kaohsiung Chang Gung Memorial Hospital and Chang Gung University College of Medicine, Kaohsiung, Taiwan.
authors.forename: Chao-Cheng
authors.initials: CC
authors.lastname: Huang
authors.affiliation: Department of Pediatric Surgery, Kaohsiung Chang Gung Memorial Hospital, Chang Gung University College of Medicine, Kaohsiung, Taiwan.
authors.forename: Chia-Ling
authors.initials: CL
authors.lastname: Wu
authors.affiliation: Department of Pediatric Surgery, Kaohsiung Chang Gung Memorial Hospital, Chang Gung University College of Medicine, Kaohsiung, Taiwan.
authors.forename: Min-Tsui
authors.initials: MT
authors.lastname: Wu
authors.affiliation: Department of Surgery, National Taiwan University Hospital and National Taiwan University College of Medicine, Taipei, Taiwan.
authors.forename: Wen-Ming
authors.initials: WM
authors.lastname: Hsu
authors.affiliation: Department of Pediatric Surgery, Kaohsiung Chang Gung Memorial Hospital, Chang Gung University College of Medicine, Kaohsiung, Taiwan.
authors.forename: Jiin-Haur
authors.initials: JH
authors.lastname: Chuang
mesh_terms: D000818:Animals; D017209:Apoptosis / Q000187:drug effects; D053148:Caspase 3 / Q000378:metabolism; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D015536:Down-Regulation / Q000187:drug effects; D016259:Genes, myc; D064593:Heterografts; D006801:Humans; D007113:Immunity, Innate; D007167:Immunotherapy; D008297:Male; D051379:Mice; D016688:Mice, Inbred NOD; D016513:Mice, SCID; D016328:NF-kappa B / Q000378:metabolism; D009447:Neuroblastoma / Q000235:genetics* / Q000378:metabolism* / Q000628:therapy; D011070:Poly I-C / Q000494:pharmacology; D016271:Proto-Oncogene Proteins c-myc / Q000235:genetics* / Q000378:metabolism*; D034622:RNA Interference; D017382:Reactive Oxygen Species / Q000378:metabolism; D051196:Toll-Like Receptor 3 / Q000819:agonists / Q000235:genetics / Q000378:metabolism*; D015854:Up-Regulation / Q000187:drug effects
chemical_list: C489427:MYC protein, human; D016328:NF-kappa B; D016271:Proto-Oncogene Proteins c-myc; D017382:Reactive Oxygen Species; C495344:TLR3 protein, human; D051196:Toll-Like Receptor 3; C505407:CASP3 protein, human; D053148:Caspase 3; D011070:Poly I-C