Hiroyuki Shimada, MD, PhD, FRCPA (Hon), is Professor of Pathology and of Pediatrics at the Stanford University Medical Center. He was born in Tokyo, Japan, and completed MD (1973) and PhD (1982) at the Yokohama City University School of Medicine, Yokohama, Japan, and also completed his pathology training at the Children's Hospital (now the Nationwide Children’s Hospital) and the Ohio State University, Columbus, Ohio, USA (1988). Before moving to the Stanford University in 2019, he was Professor of Pathology (Clinical Scholar) at the University of Southern California Keck School of Medicine and working at the Children’s Hospital Los Angeles.
Dr. Shimada was Chair of the International Neuroblastoma Pathology Committee (1999-2017) and the founder of the International Neuroblastoma Pathology Classification (INPC). As Director of the COG (Children’s Oncology Group) Neuroblastoma Pathology Reference Laboratory (since 2001), he has been actively reviewing pathology samples of ~700 neuroblastoma cases per year from United States, Canada, Australia, and New Zealand. Pathology review results according to the INPC have been providing critical information for patient stratification and protocol assignment in the COG international neuroblastoma clinical trials.

Clinical Focus

  • Pathology
  • neuroblastoma

Academic Appointments

Honors & Awards

  • Enid Gilbert-Barness Prize, Society for Pediatric Pathology (2018)
  • Honorary Fellowship, Royal College of Pathologists of Australasia (2012)
  • Eleanor Humpherys Visiting Professorship, University of Chicago (2005)
  • Lotte Straus Prize, Society for Pediatric Pathology (1989)

Professional Education

  • Residency: Nationwide Children's Hospital Pediatric Pathology (1988) OH
  • Board Certification: Japanese Society of Pathology, Pathology (1981)
  • Medical Education: Yokohama City University School of Medicine (1973) Japan

All Publications

  • Composite Neuroblastoma Metastatic to a Lymph Node: The Novel Histopathologic Diagnosis of a Unique Multiclonal Neoplasm ANNALS OF CLINICAL AND LABORATORY SCIENCE Kurtz, J. L., Shimada, H., Hazard, F. K. 2021; 51 (4): 573-579
  • Composite Neuroblastoma Metastatic to a Lymph Node: The Novel Histopathologic Diagnosis of a Unique Multiclonal Neoplasm. Annals of clinical and laboratory science Kurtz, J. L., Shimada, H., Hazard, F. K. 2021; 51 (4): 573-579


    OBJECTIVE: Composite neuroblastoma is a tumor composed of multiple tumoral clones within the neuroblastoma family. To date, establishing this unique histopathologic diagnosis has required the evaluation of the primary tumor mass. We report a case of composite neuroblastoma diagnosed by evaluation of a metastatic lymph node.METHODS: One abdominal lymph node involved by tumor was evaluated in a 6-year-old boy. The primary abdominal mass was not examined. Following histopathologic examination, clonality studies using comparative genomic hybridization (CGH) and fluorescence in situ hybridization (FISH) were also performed.RESULTS: Two distinct tumor components were identified by histopathologic evaluation and classified as differentiating neuroblastoma (component A) and poorly differentiated neuroblastoma (component B). Based on the patient's age, each clone was further classified as Unfavorable Histology. The presence of these two different tumoral clones was confirmed by CGH and FISH.CONCLUSION: This case affirms the histopathologic approach to evaluating composite tumors, as established by the International Neuroblastoma Pathology Classification (INPC) model for ganglioneuroblastoma, nodular tumors. Also, when both components are metastatic, this case demonstrates that composite tumors can be diagnosed by the evaluation of metastatic lesions alone. Finally, it supports the addition of composite neuroblastoma to a future version of the INPC.

    View details for PubMedID 34452899

  • Combining inhibitors of Brd4 and cyclin-dependent kinase can decrease tumor growth in neuroblastoma with MYCN amplification. Journal of pediatric surgery Wood, L., Huang, M., Zeki, J., Gong, M., Taylor, J., Shimada, H., Chiu, B. 2021: 60215


    INTRODUCTION: High-risk neuroblastoma is a deadly disease; poor prognosticators are MYCN-amplification and TERT-overexpression. We hypothesized that Gene Set Enrichment Analysis (GSEA) could identify pathways associated with MYCN-amplification and that inhibition of these pathways could decrease tumor growth.METHODS: We analyzed the Neuroblastoma-Kocak dataset (GSE45547, n=649) and identified pathways associated with MYCN-amplification. Inhibitors were selected from upregulated gene sets for in vitro cytotoxicity testing using ST16-patient-derived primary neuroblastoma cells and in vivo testing using orthotopic ST16-patient-derived xenografts (PDX) in mice. Tumor volume was measured with ultrasound and tumor sections examined after H&E staining.RESULTS: GSEA identified significantly overexpressed gene sets in MYCN-amplified tumors including MYC targets, cell cycle mitotic genes, TERT associated genes, loss of RB1 gene sets, and E2Fs targets. Several genes were potential Bromodomain-containing protein 4 (Brd4) targets, making Brd4 inhibitors - JQ1, AZD5153 - and cyclin-dependent kinase (Brd4's binding partner) inhibitors - dinaciclib - potential therapeutic agents. JQ1 and dinaciclib were synergistic in inducing cytotoxicity in vitro. Dinaciclib-AZD5153 in vivo decreased tumor size compared to control, and increased tumor lymphocyte infiltration and necrosis on histology.CONCLUSIONS: GSEA is a powerful approach to identify upregulated genes and potential therapeutic targets. Dinaciclib-AZD5153 combination therapy can be effective against MYCN-amplified and TERT-overexpressing neuroblastoma tumors.

    View details for DOI 10.1016/j.jpedsurg.2021.03.037

    View details for PubMedID 33838899

  • A unique composite tumour of the adrenal gland in a paediatric patient, with adrenal cortical and neural crest cell-like components. Pathology Taege, L., Shimada, H. 2021

    View details for DOI 10.1016/j.pathol.2020.10.012

    View details for PubMedID 33454141

  • Revised Neuroblastoma Risk Classification System: A Report From the Children's Oncology Group. Journal of clinical oncology : official journal of the American Society of Clinical Oncology Irwin, M. S., Naranjo, A., Zhang, F. F., Cohn, S. L., London, W. B., Gastier-Foster, J. M., Ramirez, N. C., Pfau, R., Reshmi, S., Wagner, E., Nuchtern, J., Asgharzadeh, S., Shimada, H., Maris, J. M., Bagatell, R., Park, J. R., Hogarty, M. D. 2021: JCO2100278


    Treatment planning for children with neuroblastoma requires accurate assessment of prognosis. The most recent Children's Oncology Group (COG) risk classification system used tumor stage as defined by the International Neuroblastoma Staging System. Here, we validate a revised classifier using the International Neuroblastoma Risk Group Staging System (INRGSS) and incorporate segmental chromosome aberrations (SCA) as an additional genomic biomarker.Newly diagnosed patients enrolled on the COG neuroblastoma biology study ANBL00B1 between 2007 and 2017 with known age, International Neuroblastoma Staging System, and INRGSS stage were identified (N = 4,832). Tumor MYCN status, ploidy, SCA status (1p and 11q), and International Neuroblastoma Pathology Classification histology were determined centrally. Survival analyses were performed for combinations of prognostic factors used in COG risk classification according to the prior version 1, and to validate a revised algorithm (version 2).Most patients with locoregional tumors had excellent outcomes except for those with image-defined risk factors (INRGSS L2) with MYCN amplification (5-year event-free survival and overall survival: 76.3% ± 5.8% and 79.9% ± 5.5%, respectively) or patients age ≥ 18 months with L2 MYCN nonamplified tumors with unfavorable International Neuroblastoma Pathology Classification histology (72.7% ± 5.4% and 82.4% ± 4.6%), which includes the majority of L2 patients with SCA. For patients with stage M (metastatic) and MS (metastatic, special) disease, genomic biomarkers affected risk group assignment for those < 12 months (MYCN) or 12-18 months (MYCN, histology, ploidy, and SCA) of age. In a retrospective analysis of patient outcome, the 5-year event-free survival and overall survival using COG version 1 were low-risk: 89.4% ± 1.1% and 97.9% ± 0.5%; intermediate-risk: 86.1% ± 1.3% and 94.9% ± 0.8%; high-risk: 50.8% ± 1.4% and 61.9% ± 1.3%; and using COG version 2 were low-risk: 90.7% ± 1.1% and 97.9% ± 0.5%; intermediate-risk: 85.1% ± 1.4% and 95.8% ± 0.8%; high-risk: 51.2% ± 1.4% and 62.5% ± 1.3%, respectively.A revised 2021 COG neuroblastoma risk classifier (version 2) that uses the INRGSS and incorporates SCAs has been adopted to prospectively define COG clinical trial eligibility and treatment assignment.

    View details for DOI 10.1200/JCO.21.00278

    View details for PubMedID 34319759

  • Clinical Relevance of CD4 Cytotoxic T Cells in High-Risk Neuroblastoma. Frontiers in immunology Tang, X. X., Shimada, H., Ikegaki, N. 2021; 12: 650427


    Neuroblastoma is the most common extracranial childhood solid tumor. The majority of high-risk neuroblastoma is resistant/refractory to the current high intensity therapy, and the survival of these patients remains poor for the last three decades. To effectively treat these extremely unfavorable neuroblastomas, innovative immunotherapy approaches would be the most promising. In this article, we discuss the identity of tumor-infiltrating effector cells and immunosuppressive cells in high-risk neuroblastoma. Neuroblastoma is unique in that it expresses little or no classical HLA Class I and II. In contrast, high-risk neuroblastomas express the stress-responsive non-classical Class I, HLA-E molecule. HLA-E is the ligand of activating receptors NKG2C/E that are expressed on memory NK cells, CD8+T cells and CD4 CTLs. By examining a comprehensive RNA-seq gene expression dataset, we detected relatively high levels of CD4 expression in high-risk neuroblastoma tissues. The majority of CD4+ cells were CD3+, and thus they were likely tumor-associated CD4+T cells. In addition, high-level of both CD4 and NKG2C/E expression was associated with prolonged survival of the high-risk neuroblastoma patients, but CD8 levels were not, further suggesting that the CD4+ NKG2C/E+ T cells or CD4 CTL conferred cytotoxicity against the neuroblastoma cells. However, this T cell mediated- "protective effect" declined over time, in part due to the progressive formation of immunosuppressive tumor microenvironment. These observations suggest that to improve survival of high-risk neuroblastoma patients, it is essential to gain insights into how to enhance CD4 CTL cytotoxicity and control the immunosuppressive tumor microenvironment during the course of the disease.

    View details for DOI 10.3389/fimmu.2021.650427

    View details for PubMedID 33968044

  • Randomized Phase II Trial of MIBG Versus MIBG, Vincristine, and Irinotecan Versus MIBG and Vorinostat for Patients With Relapsed or Refractory Neuroblastoma: A Report From NANT Consortium. Journal of clinical oncology : official journal of the American Society of Clinical Oncology DuBois, S. G., Granger, M. M., Groshen, S., Tsao-Wei, D., Ji, L., Shamirian, A., Czarnecki, S., Goodarzian, F., Berkovich, R., Shimada, H., Villablanca, J. G., Vo, K. T., Pinto, N., Mosse, Y. P., Maris, J. M., Shusterman, S., Cohn, S. L., Goldsmith, K. C., Weiss, B., Yanik, G. A., Twist, C. J., Irwin, M. S., Haas-Kogan, D. A., Park, J. R., Marachelian, A., Matthay, K. K. 2021: JCO2100703


    131I-metaiodobenzylguanidine (MIBG) is an active radiotherapeutic for neuroblastoma. The primary aim of this trial was to identify which of three MIBG regimens was likely associated with the highest true response rate.Patients 1-30 years were eligible if they had relapsed or refractory neuroblastoma, at least one MIBG-avid site, and adequate autologous stem cells. Patients received MIBG 18 mCi/kg on day 1 and autologous stem cell on day 15. Patients randomly assigned to arm A received only MIBG; patients randomly assigned to arm B received intravenous vincristine on day 0 and irinotecan daily on days 0-4; patients randomly assigned to arm C received vorinostat (180 mg/m2/dose) orally once daily on days 1 to 12. The primary end point was response after one course by New Approaches to Neuroblastoma Therapy criteria. The trial was designed with 105 patients to ensure an 80% chance that the arm with highest response rate was selected.One hundred fourteen patients were enrolled, with three ineligible and six unevaluable, leaving 105 eligible and evaluable patients (36 in arm A, 35 in arm B, and 34 in arm C; 55 boys; and median age 6.5 years). After one course, the response rates (partial response or better) on arms A, B, and C were 14% (95% CI, 5 to 30), 14% (5 to 31), and 32% (18 to 51). An additional five, five, and four patients met New Approaches to Neuroblastoma Therapy Minor Response criteria on arms A, B, and C, respectively. On arms A, B, and C, rates of any grade 3+ nonhematologic toxicity after first course were 19%, 49%, and 35%.Vorinostat and MIBG is likely the arm with the highest true response rate, with manageable toxicity. Vincristine and irinotecan do not appear to improve the response rate to MIBG and are associated with increased toxicity.

    View details for DOI 10.1200/JCO.21.00703

    View details for PubMedID 34270348

  • Stage 4S Neuroblastoma: Molecular, Histologic, and Immunohistochemical Characteristics and Presence of 2 Distinct Patterns of MYCN Protein Overexpression-A Report From the Children's Oncology Group. The American journal of surgical pathology Kawano, A., Hazard, F. K., Chiu, B., Naranjo, A., LaBarre, B., London, W. B., Hogarty, M. D., Cohn, S. L., Maris, J. M., Park, J. R., Gastier-Foster, J. M., Ikegaki, N., Shimada, H. 2020


    Stage 4S neuroblastoma (4SNB) is associated with spontaneous tumor regression and an excellent prognosis. However, a small group of the patients have a poor prognosis. One hundred eighty-five 4SNB cases filed at the Children's Oncology Group Neuroblastoma Pathology Reference Laboratory were studied. MYCN oncogene status [non-amplified (NA) vs. Amplified (A)] determined by fluorescence in situ hybridization, MYC-family (MYCN/MYC) protein expression [no-overexpression(-)/(+/-) vs. overexpression(+)] by immunohistochemistry and histopathology by International Neuroblastoma Pathology Classification [Favorable Histology (FH) vs. Unfavorable Histology (UH)] with particular attention to nucleolar hypertrophy [NH(-) vs. (+)] were assessed with patient survival. One hundred forty-seven (79.5%) tumors were MYCN-NA, FH, MYC-family protein(-)/(+/-), and NH(-) with a good prognosis [88.5%+3.1% 5-y event-free survival (EFS); 94.1%+2.3% 5-y overall survival (OS)]. Among MYCN-NA tumors, 11 demonstrated MYCN protein(+) with a moderate and uniform (M/U) staining pattern: they were FH(10/11), NH(-), 1 showed MYC protein(+) simultaneously, and all patients are alive. Also found were 5 MYC protein(+) and MYCN(-)/(+/-) tumors; they were FH without NH (4/5), and all patients are alive. Among MYCN-A tumors, 18 had MYCN protein(+) with a strong and heterogeneous (S/H) staining pattern, 9 had UH (44.4%+23.4% EFS/OS) and 9 had FH (68.6%+19.2% EFS/OS), and 15 showed NH(+). Two tumors had MYCN protein(-)/(+/-) despite MYCN-A; both were FH and NH(-), and 1 patient died. S/H staining pattern of MYCN protein overexpression by immunohistochemistry was associated with MYCN amplification, NH(+) and a poor prognosis. In contrast, the M/U staining pattern was associated with MYCN nonamplification and NH(-), and had no adverse prognostic effects for the 4SNB patients.

    View details for DOI 10.1097/PAS.0000000000001647

    View details for PubMedID 33739795

  • Gene Expression Signature in Human Neuroblastoma with TERT Overexpression Can Be Identified by Gene Set Enrichment Analysis and Epigenetically Targeted in an Orthotopic Mouse Xenograft Model Huang, M., Wood, L., Zeki, J. C., Diyaolu, M., Gong, M., Taylor, J. S., Shimada, H., Chiu, B. ELSEVIER SCIENCE INC. 2020: S199
  • Outcomes and toxicities in patients (pts) non-randomly assigned to immunotherapy Children's Oncology Group (COG) ANBL0032. Desai, A., Gilman, A., Ozkaynak, M., Naranjo, A., London, W. B., Tenney, S., Smith, M., Seibel, N., Shimada, H., Matthay, K. K., Cohn, S., Maris, J. M., Bagatell, R., Sondel, P. M., Park, J. R., Yu, A. L. AMER SOC CLINICAL ONCOLOGY. 2020
  • Randomized phase II trial of MIBG versus MIBG/vincristine/irinotecan versus MIBG/vorinostat for relapsed/refractory neuroblastoma: A report from the New Approaches to Neuroblastoma Therapy Consortium. DuBois, S. G., Granger, M., Groshen, S. G., Tsao-Wei, D., Shamirian, A., Czarnecki, S., Goodarzian, F., Berkovich, R., Shimada, H., Mosse, Y. P., Shusterman, S., Cohn, S., Goldsmith, K. C., Weiss, B. D., Yanik, G. A., Twist, C., Irwin, M., Park, J. R., Marachelian, A., Matthay, K. K. AMER SOC CLINICAL ONCOLOGY. 2020
  • The Role of the Clinical Laboratory in the Diagnosis of Neuroblastoma JOURNAL OF APPLIED LABORATORY MEDICINE Hazard, F. K., Shimada, H. 2020; 5 (2): 254–56
  • Enhancing sustained-release local therapy: Single versus dual chemotherapy for the treatment of neuroblastoma. Surgery Taylor, J. S., Yavuz, B., Zeki, J., Wood, L., Ikegaki, N., Coburn, J., Harrington, K., Shimada, H., Kaplan, D. L., Chiu, B. 2020


    BACKGROUND: Neuroblastoma is the most common pediatric extracranial solid malignancy with limited effective treatment. We have shown that sustained-release, single drugs delivered locally through a silk-based biomaterial are effective in decreasing orthotopic neuroblastoma xenograft growth. We further optimized this approach and hypothesized that increasing doses of local chemotherapy or delivering 2 chemotherapeutic agents simultaneously inhibit additional tumor growth.METHODS: MYCN-amplified and non-MYCN-amplified neuroblastoma cells were treated with combinations of cisplatin, vincristine, doxorubicin, and etoposide to determine cytotoxicity and synergy. Drug-loaded silk material was created, and the amounts of drug released from the material over time were recorded. Murine orthotopic neuroblastoma xenografts were generated; tumors were implanted with single- or dual-agent chemotherapy-loaded silk. Ultrasound was used to monitor tumor growth, and tumor histology was evaluated.RESULTS: Invitro, vincristine/cisplatin combination was synergistic and significantly decreased cell viability relative to other combinations. Both drugs loaded into silk could be released effectively for over 2 weeks. Locally implanted vincristine/cisplatin silk induced increased tumor growth suppression compared with either agent alone in MYCN-amplified tumors (P < .05). The dose-dependent effect seen in MYCN-amplified tumors treated with combination therapy diminished at higher doses in non-MYCN-amplified tumors, with little benefit with doses >50 mug to 500 mug for vincristine-cisplatin, respectively. Tumor histology demonstrated tumor cell necrosis adjacent to drug-loaded silk material and presence of large cell neuroblastoma.CONCLUSION: Local delivery of sustained release chemotherapy can suppress tumor growth especially at high doses or with 2 synergistic drugs. Locally delivered dual therapy is a promising approach for future clinical testing.

    View details for DOI 10.1016/j.surg.2020.01.012

    View details for PubMedID 32122657

  • Local delivery of dinutuximab from lyophilized silk fibroin foams for treatment of an orthotopic neuroblastoma model. Cancer medicine Ornell, K. J., Taylor, J. S., Zeki, J., Ikegaki, N., Shimada, H., Coburn, J. M., Chiu, B. 2020


    Immunotherapy targeting GD2 is a primary treatment for patients with high-risk neuroblastoma. Dinutuximab is a monoclonal antibody with great clinical promise but is limited by side effects such as severe pain. Local delivery has emerged as a potential mechanism to deliver higher doses of therapeutics into the tumor bed, while limiting systemic toxicity. We aim to deliver dinutuximab locally in a lyophilized silk fibroin foam for the treatment of an orthotopic neuroblastoma mouse model. Dinutuximab-loaded silk fibroin foams were fabricated through lyophilization. In vitro release profile and bioactivity of the release through complement-dependent cytotoxicity were characterized. MYCN-amplified neuroblastoma cells (KELLY) were injected into the left gland of mice to generate an orthotopic neuroblastoma model. Once the tumor volume reached 100mm3 , dinutuximab-, human IgG-, or buffer-loaded foams were implanted into the tumor and growth was monitored using high-resolution ultrasound. Post-resection histology was performed on tumors. Dinutuximab-loaded silk fibroin foams exhibited a burst release, with slow release thereafter in vitro with maintenance of bioactivity. The dinutuximab-loaded foam significantly inhibited xenograft tumor growth compared to IgG- and buffer-loaded foams. Histological analysis revealed the presence of dinutuximab within the tumor and neutrophils and macrophages infiltrating into dinutuximab-loaded silk foam. Tumors treated with local dinutuximab had decreased MYCN expression on histology compared to control or IgG-treated tumors. Silk fibroin foams offer a mechanism for local release of dinutuximab within the neuroblastoma tumor. This local delivery achieved a significant decrease in tumor growth rate in a mouse orthotopic tumor model.

    View details for DOI 10.1002/cam4.2936

    View details for PubMedID 32096344

  • Association of heterogeneous MYCN amplification with clinical features, biological characteristics and outcomes in neuroblastoma: A report from the Children's Oncology Group. European journal of cancer (Oxford, England : 1990) Campbell, K. n., Naranjo, A. n., Hibbitts, E. n., Gastier-Foster, J. M., Bagatell, R. n., Irwin, M. S., Shimada, H. n., Hogarty, M. n., Park, J. R., DuBois, S. G. 2020; 133: 112–19


    MYCN amplification (MNA) is associated with poor outcomes in neuroblastoma. Less is known about heterogeneous MNA within a tumour. We compared clinical characteristics, biologic features and clinical outcomes of patients with heterogeneous MNA to patients with either homogeneous MNA or MYCN wild-type tumours.In this retrospective cohort study, we categorized patients as having tumours with MYCN wild-type, homogeneous MNA (>20% amplified tumour cells) or heterogeneous MNA (≤20% amplified tumour cells). We used chi-squared or Fisher's exact tests to compare features between groups. We used log-rank tests and Cox models to compare event-free survival (EFS) and overall survival (OS) between groups.MYCN status and heterogeneity status (if amplified) could be ascertained in diagnostic tumour samples from 5975 patients, including 57 (1%) with heterogeneous MNA, 981 (16.4%) with homogeneous MNA, and 4937 (82.6%) with MYCN wild-type tumours. Multiple clinical and biological features differed between patients with heterogeneous vs. homogeneous MNA, including enrichment for thoracic primary sites and paucity of 1p loss of heterozygosity with heterogeneous MNA (p < 0.0001). Importantly, EFS and OS were not significantly different between patients with heterogeneous vs. homogeneous MNA. Further, EFS and OS for patients with heterogeneous MNA were significantly inferior to patients with wild-type MYCN.Although neuroblastomas with heterogeneous MNA demonstrate significantly different biological and clinical patterns compared with homogeneous MNA, prognosis is similar between the two groups. These results support current practice that treats patients with heterogeneous MNA similarly to patients with homogeneous MNA.

    View details for DOI 10.1016/j.ejca.2020.04.007

    View details for PubMedID 32492633

  • The Role of the Clinical Laboratory in the Diagnosis of Neuroblastoma. The journal of applied laboratory medicine Hazard, F. K., Shimada, H. n. 2020; 5 (2): 254–56

    View details for DOI 10.1093/jalm/jfz005

    View details for PubMedID 32445372

  • MYC transcription activation mediated by OCT4 as a mechanism of resistance to 13-cisRA-mediated differentiation in neuroblastoma. Cell death & disease Wei, S. J., Nguyen, T. H., Yang, I. H., Mook, D. G., Makena, M. R., Verlekar, D. n., Hindle, A. n., Martinez, G. M., Yang, S. n., Shimada, H. n., Reynolds, C. P., Kang, M. H. 2020; 11 (5): 368


    Despite the improvement in clinical outcome with 13-cis-retinoic acid (13-cisRA) + anti-GD2 antibody + cytokine immunotherapy given in first response ~40% of high-risk neuroblastoma patients die of recurrent disease. MYCN genomic amplification is a biomarker of aggressive tumors in the childhood cancer neuroblastoma. MYCN expression is downregulated by 13-cisRA, a differentiating agent that is a component of neuroblastoma therapy. Although MYC amplification is rare in neuroblastoma at diagnosis, we report transcriptional activation of MYC medicated by the transcription factor OCT4, functionally replacing MYCN in 13-cisRA-resistant progressive disease neuroblastoma in large panels of patient-derived cell lines and xenograft models. We identified novel OCT4-binding sites in the MYC promoter/enhancer region that regulated MYC expression via phosphorylation by MAPKAPK2 (MK2). OCT4 phosphorylation at the S111 residue by MK2 was upstream of MYC transcriptional activation. Expression of OCT4, MK2, and c-MYC was higher in progressive disease relative to pre-therapy neuroblastomas and was associated with inferior patient survival. OCT4 or MK2 knockdown decreased c-MYC expression and restored the sensitivity to 13-cisRA. In conclusion, we demonstrated that high c-MYC expression independent of genomic amplification is associated with disease progression in neuroblastoma. MK2-mediated OCT4 transcriptional activation is a novel mechanism for activating the MYC oncogene in progressive disease neuroblastoma that provides a therapeutic target.

    View details for DOI 10.1038/s41419-020-2563-4

    View details for PubMedID 32409685

    View details for PubMedCentralID PMC7224192

  • Age Inherently Links to Histology to Define Histoprognostic Classification of Peripheral Neuroblastic Tumors. Journal of clinical oncology : official journal of the American Society of Clinical Oncology Beiske, K. n., Ikegaki, N. n., Shimada, H. n. 2020: JCO2001839

    View details for DOI 10.1200/JCO.20.01839

    View details for PubMedID 32931395

  • Replicating and identifying large cell neuroblastoma using high-dose intra-tumoral chemotherapy and automated digital analysis. Journal of pediatric surgery Taylor, J. S., Sha, L., Ikegaki, N., Zeki, J., Deaton, R., Harris, J., Coburn, J., Yavuz, B., Sethi, A., Shimada, H., Kaplan, D. L., Gann, P., Chiu, B. 2019


    PURPOSE: Large cell neuroblastomas (LCN) are frequently seen in recurrent, high-risk neuroblastoma but are rare in primary tumors. LCN, characterized by large nuclei with prominent nucleoli, predict a poor prognosis. We hypothesize that LCN can be created with high-dose intra-tumoral chemotherapy and identified by a digital analysis system.METHODS: Orthotopic mouse xenografts were created using human neuroblastoma and treated with high-dose chemotherapy delivered locally via sustained-release silk platforms, inducing tumor remission. After recurrence, LCN populations were identified on H&E sections manually. Clusters of typical LCN and non-LCN cells were divided equally into training and test sets for digital analysis. Marker-controlled watershed segmentation was used to identify nuclei and characterize their features. Logistic regression was developed to distinguish LCN from non-LCN.RESULTS: Image analysis identified 15,000 nuclei and characterized 70 nuclear features. A 19-feature model provided AUC >0.90 and 100% accuracy when >30% nuclei/cluster were predicted as LCN. Overall accuracy was 87%.CONCLUSIONS: We recreated LCN using high-dose chemotherapy and developed an automated method for defining LCN histologically. Features in the model provide insight into LCN nuclear phenotypic changes that may be related to increased activity. This model could be adapted to identify LCN in human tumors and correlated with clinical outcomes.

    View details for DOI 10.1016/j.jpedsurg.2019.08.022

    View details for PubMedID 31519361

  • Down-regulation of MYCN protein by CX-5461 leads to neuroblastoma tumor growth suppression. Journal of pediatric surgery Taylor, J. S., Zeki, J., Ornell, K., Coburn, J., Shimada, H., Ikegaki, N., Chiu, B. 2019


    PURPOSE: MYCN oncogene amplification is an independent predictor of poor prognosis in neuroblastoma. CX-5461 is a small molecular inhibitor that prevents initiation of ribosomal RNA (rRNA) synthesis by RNA Pol I, down-regulating MYCN/MYC proteins. We hypothesize that neuroblastoma tumor growth can be suppressed by CX-5461.METHODS: MYCN-amplified (KELLY, IMR5) and nonamplified (SY5Y, SKNAS) neuroblastoma cells were treated with CX-5461. MYCN/MYC expression after 24-48 h was determined by Western blot. Orthotopic neuroblastoma tumors created in mice using KELLY cells were treated with CX-5461-loaded silk films implanted locally. Tumor growth was monitored using ultrasound. Histologic evaluation of tumors was performed.RESULTS: IC50 for KELLY, IMR5, SY5Y, and SKNAS cells to CX-5461 was 0.75 muM, 0.02 muM, 0.8 muM, and 1.7 muM, respectively. CX-5461 down-regulated MYCN and MYC proteins at 0.25-1.0 muM on Western blot analysis. CX-5461-loaded silk film released 23.7±3 mug of the drug in 24 h and 48.2±3.9 mug at 120 h. KELLY tumors treated with CX-5461-loaded film reached 800 mm3 after 7.8±1.4 days, while those treated with control film reached the same size on 5.1±0.6 days (p=0.03). CX-5461-treated tumors showed collapse of nucleolar hypertrophy and MYCN protein downregulation.CONCLUSION: We demonstrated that local delivery of CX-5461 via sustained release platform can suppress orthotopic neuroblastoma tumor growth, especially those with MYCN/MYC overexpression.

    View details for PubMedID 30879743

  • Anti-CD105 Antibody Eliminates Tumor Microenvironment Cells and Enhances Anti-GD2 Antibody Immunotherapy of Neuroblastoma with Activated Natural Killer Cells. Clinical cancer research : an official journal of the American Association for Cancer Research Wu, H. W., Sheard, M. A., Malvar, J. n., Fernandez, G. E., DeClerck, Y. A., Blavier, L. n., Shimada, H. n., Theuer, C. P., Sposto, R. n., Seeger, R. C. 2019; 25 (15): 4761–74


    We determined whether elimination of CD105+ cells in the tumor microenvironment (TME) with anti-CD105 antibodies enhanced anti-disialoganglioside (GD2) antibody dinutuximab therapy of neuroblastoma when combined with activated natural killer (aNK) cells.The effect of MSCs and monocytes on antibody-dependent cellular cytotoxicity (ADCC) mediated by dinutuximab with aNK cells against neuroblastoma cells was determined in vitro. ADCC with anti-CD105 mAb TRC105 and aNK cells against MSCs, monocytes, and endothelial cells, which express CD105, was evaluated. Anti-neuroblastoma activity in immunodeficient NSG mice of dinutuximab with aNK cells without or with anti-CD105 mAbs was determined using neuroblastoma cell lines and a patient-derived xenograft.ADCC mediated by dinutuximab with aNK cells against neuroblastoma cells in vitro was suppressed by addition of MSCs and monocytes, and dinutuximab with aNK cells was less effective against neuroblastomas formed with coinjected MSCs and monocytes in NSG mice than against those formed by tumor cells alone. Anti-CD105 antibody TRC105 with aNK cells mediated ADCC against MSCs, monocytes, and endothelial cells. Neuroblastomas formed in NSG mice by two neuroblastoma cell lines or a patient-derived xenograft coinjected with MSCs and monocytes were most effectively treated with dinutuximab and aNK cells when anti-human (TRC105) and anti-mouse (M1043) CD105 antibodies were added, which depleted human MSCs and murine endothelial cells and macrophages from the TME.Immunotherapy of neuroblastoma with anti-GD2 antibody dinutuximab and aNK cells is suppressed by CD105+ cells in the TME, but suppression is overcome by adding anti-CD105 antibodies to eliminate CD105+ cells.

    View details for DOI 10.1158/1078-0432.CCR-18-3358

    View details for PubMedID 31068371

  • MYC-family protein overexpression and prominent nucleolar formation represent prognostic indicators and potential therapeutic targets for aggressive high-MKI neuroblastomas: a report from the children's oncology group. Oncotarget Niemas-Teshiba, R. n., Matsuno, R. n., Wang, L. L., Tang, X. X., Chiu, B. n., Zeki, J. n., Coburn, J. n., Ornell, K. n., Naranjo, A. n., Van Ryn, C. n., London, W. B., Hogarty, M. D., Gastier-Foster, J. M., Look, A. T., Park, J. R., Maris, J. M., Cohn, S. L., Seeger, R. C., Asgharzadeh, S. n., Ikegaki, N. n., Shimada, H. n. 2018; 9 (5): 6416–32


    Neuroblastomas with a high mitosis-karyorrhexis index (High-MKI) are often associated with MYCN amplification, MYCN protein overexpression and adverse clinical outcome. However, the prognostic effect of MYC-family protein expression on these neuroblastomas is less understood, especially when MYCN is not amplified. To address this, MYCN and MYC protein expression in High-MKI cases (120 MYCN amplified and 121 non-MYCN amplified) was examined by immunohistochemistry. The majority (101) of MYCN-amplified High-MKI tumors were MYCN(+), leaving one MYC(+), 2 both(+), and 16 both(-)/(+/-), whereas non-MYCN-amplified cases appeared heterogeneous, including 7 MYCN(+), 36 MYC(+), 3 both(+), and 75 both(-)/(+/-) tumors. These MYC-family proteins(+), or MYC-family driven tumors, were most likely to have prominent nucleolar (PN) formation (indicative of augmented rRNA synthesis). High-MKI neuroblastoma patients showed a poor survival irrespective of MYCN amplification. However, patients with MYC-family driven High-MKI neuroblastomas had significantly lower survival than those with non-MYC-family driven tumors. MYCN(+), MYC-family protein(+), PN(+), and clinical stage independently predicted poor survival. Specific inhibition of hyperactive rRNA synthesis and protein translation was shown to be an effective way to suppress MYC/MYCN protein expression and neuroblastoma growth. Together, MYC-family protein overexpression and PN formation should be included in new neuroblastoma risk stratification and considered for potential therapeutic targets.

    View details for PubMedID 29464082

    View details for PubMedCentralID PMC5814222

  • Dose Escalation Study of No-Carrier-Added I-131-Metaiodobenzylguanidine for Relapsed or Refractory Neuroblastoma: New Approaches to Neuroblastoma Therapy Consortium Trial JOURNAL OF NUCLEAR MEDICINE Matthay, K. K., Weiss, B., Villablanca, J. G., Maris, J. M., Yanik, G. A., DuBois, S. G., Stubbs, J., Groshen, S., Tsao-Wei, D., Hawkins, R., Jackson, H., Goodarzian, F., Daldrup-Link, H., Panigrahy, A., Towbin, A., Shimada, H., Barrett, J., LaFrance, N., Babich, J. 2012; 53 (7): 1155-1163


    (131)I-metaiodobenzylguanidine (MIBG) is specifically taken up in neuroblastoma, with a response rate of 20%-37% in relapsed disease. Nonradioactive carrier MIBG molecules inhibit uptake of (131)I-MIBG, theoretically resulting in less tumor radiation and increased risk of cardiovascular toxicity. Our aim was to establish the maximum tolerated dose of no-carrier-added (NCA) (131)I-MIBG, with secondary aims of assessing tumor and organ dosimetry and overall response.Eligible patients were 1-30 y old with resistant neuroblastoma, (131)I-MIBG uptake, and cryopreserved hematopoietic stem cells. A diagnostic dose of NCA (131)I-MIBG was followed by 3 dosimetry scans to assess radiation dose to critical organs and soft-tissue tumors. The treatment dose of NCA (131)I-MIBG (specific activity, 165 MBq/μg) was adjusted as necessary on the basis of critical organ tolerance limits. Autologous hematopoietic stem cells were infused 14 d after therapy to abrogate prolonged myelosuppression. Response and toxicity were evaluated on day 60. The NCA (131)I-MIBG was escalated from 444 to 777 MBq/kg (12-21 mCi/kg) using a 3 + 3 design. Dose-limiting toxicity (DLT) was failure to reconstitute neutrophils to greater than 500/μL within 28 d or platelets to greater than 20,000/μL within 56 d, or grade 3 or 4 nonhematologic toxicity by Common Terminology Criteria for Adverse Events (version 3.0) except for predefined exclusions.Three patients each were evaluable at 444, 555, and 666 MBq/kg without DLT. The dose of 777 MBq/kg dose was not feasible because of organ dosimetry limits; however, 3 assigned patients were evaluable for a received dose of 666 MBq/kg, providing a total of 6 patients evaluable for toxicity at 666 MBq/kg without DLT. Mean whole-body radiation was 0.23 mGy/MBq, and mean organ doses were 0.92, 0.82, and 1.2 mGy/MBq of MIBG for the liver, lung, and kidney, respectively. Eight patients had 13 soft-tissue lesions with tumor-absorbed doses of 26-378 Gy. Four of 15 patients had a complete (n = 1) or partial (n = 3) response, 1 had a mixed response, 4 had stable disease, and 6 had progressive disease.NCA (131)I-MIBG with autologous peripheral blood stem cell transplantation is feasible at 666 MBq/kg without significant nonhematologic toxicity and with promising activity.

    View details for DOI 10.2967/jnumed.111.098624

    View details for Web of Science ID 000306164600033

    View details for PubMedID 22700000

  • Outcome analysis of non-high-risk neuroblastoma patients enrolled on Children's Oncology Group trials P9641 and A3961 Meany, H., Attiyeh, E. F., Naranjo, A., Twist, C., London, W. B., Villablanca, J., Schmidt, M., Baker, D., Strother, D. R., Shimada, H., Matthay, K. K., Cohn, S., Maris, J. M., Park, J. R. AMER SOC CLINICAL ONCOLOGY. 2012
  • Current Treatment Protocols Have Eliminated the Prognostic Advantage of Type 1 Fusions in Ewing Sarcoma: A Report From the Children's Oncology Group JOURNAL OF CLINICAL ONCOLOGY van Doorninck, J. A., Ji, L., Schaub, B., Shimada, H., Wing, M. R., Krailo, M. D., Lessnick, S. L., Marina, N., Triche, T. J., Sposto, R., Womer, R. B., Lawlor, E. R. 2010; 28 (12): 1989-1994


    PURPOSE Ewing sarcoma family tumors (ESFTs) exhibit chromosomal translocations that lead to the creation of chimeric fusion oncogenes. Combinatorial diversity among chromosomal breakpoints produces varying fusions. The type 1 EWS-FLI1 transcript is created as a result of fusion between exons 7 of EWS and 6 of FLI1, and retrospective studies have reported that type 1 tumors are associated with an improved outcome. We have re-examined this association in a prospective cohort of patients with ESFT treated according to current Children's Oncology Group (COG) treatment protocols. METHODS Frozen tumor tissue was prospectively obtained from patients diagnosed with ESFT, and reverse transcriptase polymerase chain reaction (RT-PCR) was used to determine translocation status. Analysis was confined to patients with localized tumors who were diagnosed after 1994 and treated according to COG protocols. Translocation status was correlated with disease characteristics, event-free survival (EFS), and overall survival (OS). Results RT-PCR identified chimeric fusion oncogenes in 119 of 132 ESFTs. Eighty-nine percent of identified transcripts were EWS-FLI1, and of these, 58.8% were type 1. Five-year EFS and OS rates for patients with type 1 and non-type 1 fusions diagnosed between 2001 and 2005 were equivalent (type 1: EFS, 63% +/- 7%; OS, 83% +/- 6%; non-type 1: EFS, 71% +/- 9%; OS, 79% +/- 8%). CONCLUSION Current intensive treatment protocols for localized ESFT have erased the clinical disadvantage that was formerly observed in patients with non-type 1 EWS-FLI1 fusions.

    View details for DOI 10.1200/JCO.2009.24.5845

    View details for Web of Science ID 000276764000006

    View details for PubMedID 20308669

    View details for PubMedCentralID PMC2860404

  • Mouse mesenchymal stem cells expressing PAX-FKHR form alveolar rhabdomyosarcomas by cooperating with secondary mutations CANCER RESEARCH Ren, Y., Finckenstein, F., Abdueva, D. A., Shahbazian, V., Chung, B., Weinberg, K. I., Triche, T. J., Shimada, H., Anderson, M. J. 2008; 68 (16): 6587–97


    Alveolar rhabdomyosarcomas (ARMS) are highly malignant soft-tissue sarcomas that arise in children, adolescents, and young adults. Although formation and expression of the PAX-FKHR fusion genes is thought to be the initiating event in this cancer, the role of PAX-FKHR in the neoplastic process remains largely unknown in a progenitor cell that is undefined. We hypothesize that PAX-FKHR determine the ARMS progenitor to the skeletal muscle lineage, which when coupled to the inactivation and/or activation of critical cell signaling pathways leads to the formation of ARMS. Because a number of studies have proposed that mesenchymal stem cells (MSC) are the progenitor for several of the sarcomas, we tested this hypothesis in MSCs. We show that PAX-FKHR induce skeletal myogenesis in MSCs by transactivating MyoD and myogenin. Despite exhibiting enhanced growth in vitro, the PAX-FKHR-expressing populations do not form colonies in soft agar or tumors in mice. Expression of dominant-negative p53, or the SV40 early region, elicits tumor formation in some of the PAX-FKHR-expressing populations. Additional activation of the Ras signaling pathway leads to highly malignant tumor formation for all of the populations. The PAX-FKHR-expressing tumors were shown to have histologic, immunohistochemical, and gene expression profiles similar to human ARMS. Our results show the critical role played by PAX-FKHR in determining the molecular, myogenic, and histologic phenotype of ARMS. More importantly, we identify MSCs as a progenitor that can give rise to ARMS.

    View details for DOI 10.1158/0008-5472.CAN-08-0859

    View details for Web of Science ID 000258548200015

    View details for PubMedID 18701482

  • Primary and metastatic rhabdomyosarcoma in the breast: Neoplasms of adolescent females, a report from the intergroup rhabdomyosarcoma study MEDICAL AND PEDIATRIC ONCOLOGY Hays, D. M., Donaldson, S. S., Shimada, H., Crist, W. M., Newton, W. A., Andrassy, R. J., Wiener, E., Green, J., Triche, T., Maurer, H. M. 1997; 29 (3): 181-189


    The occurrence of rhabdomyosarcoma (RMS) primary in or metastatic to breast has been regarded as an uncommon event, associated with an unfavorable outcome. Records of 26 patients with diagnoses of breast RMS, either primary or secondary, entered in the Intergroup Rhabdomyosarcoma Study (IRS) (1972-1992) were reviewed and compared with data regarding 47 similar patients in published reports. Of the 26 IRS cases, the histologic subtype was alveolar in 24, embryonal in 1, and not determined in 1. All were female with ages ranging from 11.5 to 20.2 years (median, 15.2 years; mode, 14-16 years). This compact age distribution of both primary (n = 7) and metastatic (n = 19) breast RMS was seen in previously reported series. Among the 19 cases of RMS with initial dissemination to breast, primary tumor sites, were extremity (n = 8), nasopharynx/paranasal sinuses (n = 7), and trunk (n = 4). IRS treatment was risk-based according to site and extent of disease. Four of 7 patients with primary RMS remain disease free 2.9 to 7 years post diagnosis. Among 19 patients with RMS initially metastatic to breast, including 7 in IRS clinical group IV at original diagnosis, three are disease free at 7.6, 15.7 and 17.0 years. Conclusions: primary or metastatic RMS in breast is almost confined to adolescent females having tumors with alveolar histology. Approximately one-half of the patients with primary breast disease and 15% of those with metastatic breast disease as an initial recurrence are long-term survivors.

    View details for Web of Science ID A1997XH25700004

    View details for PubMedID 9212842