Poster Presentation Advances in Neuroblastoma Research Congress 2016

Human stem cell models for relapse neuroblastoma (#376)

Tina Zheng 1 , Miller Huang 1 , Max A Horlbeck 1 , Luke A Gilbert 1 , Jonathan S Weissman 1 , William A Weiss 1
  1. University of California San Francisco, San Francisco, CA, United States

High-risk neuroblastoma is enigmatic among aggressive tumors. Initially treatment-sensitive, these tumors inevitably acquire therapy-resistance. Relapse neuroblastoma demonstrates frequent activation of mitogen activated protein kinase (MAPK) signaling1. Relapse tumors mutate NF1, a tumor suppressor also frequently silenced epigenetically in neuroblastoma1,2. NF1 is a GTPase activating protein that negatively regulates the GTPase RAS. Loss of NF1 activates MAPK in hematopoietic tumors3, and phosphatidylinositol 3’ kinase (PI3K) in malignant neural sheath tumors4,5. How NF1 loss affects signaling in relapse neuroblastoma, and how to best treat these patients, remains poorly understood. To model epigenetic silencing of NF1 in neuroblastoma, we used CRISPR interference (CRISPRi), blocking transcription of NF1 reversibly in human induced pluripotent stem cells (iPSC). Human iPSC were then differentiated to trunk neural crest cells (NCC), presumed cells of origin for neuroblastoma6. We combined NF1 CRISPRi with misexpression of MYCN. Transduced NCC were subsequently introduced orthotopically into mice, to generate MYCN-driven, NF1 silenced tumors. Characterization of these tumors promises to clarify signaling pathways activated in response to silencing of NF1, to determine how silencing of NF1, when combined with misexpression of MYCN, can inform treatment of relapse, high-risk, therapy resistant neuroblastoma in vivo. We demonstrate that NF1 CRISPRi leads to decreased levels of NF1 protein. We are currently combining CRISPi and mis-expression of NF1, to generate a “human in mouse” model of recurrent disease. Human-in-mouse models for MYCN amplified, NF1 silenced neuroblastoma promise to inform biology and therapy for high-risk relapse neuroblastoma.

  1. Thomas F Eleveld et al., “Relapsed Neuroblastomas Show Frequent RAS-MAPK Pathway Mutations.,” Nature Genetics 47, no. 8 (August 2015): 864–71, doi:10.1038/ng.3333.
  2. Michael Hölzel et al., “NF1 Is a Tumor Suppressor in Neuroblastoma That Determines Retinoic Acid Response and Disease Outcome,” Cell 142, no. 2 (July 2010): 218–29, doi:10.1016/j.cell.2010.06.004.
  3. Jennifer O Lauchle et al., “Response and Resistance to MEK Inhibition in Leukaemias Initiated by Hyperactive Ras.,” Nature 461, no. 7262 (September 17, 2009): 411–14, doi:10.1038/nature08279.
  4. Cory M Johannessen et al., “The NF1 Tumor Suppressor Critically Regulates TSC2 and mTOR.,” Proceedings of the National Academy of Sciences of the United States of America 102, no. 24 (June 14, 2005): 8573–78, doi:10.1073/pnas.0503224102.
  5. Clare F Malone et al., “Defining Key Signaling Nodes and Therapeutic Biomarkers in NF1-Mutant Cancers.,” Cancer Discovery 4, no. 9 (September 2014): 1062–73, doi:10.1158/2159-8290.CD-14-0159.
  6. Nai-Kong V Cheung and Michael A Dyer, “Neuroblastoma: Developmental Biology, Cancer Genomics and Immunotherapy,” June 1, 2013, 1–15, doi:10.1038/nrc3526.