Small Extracellular Vesicle mediated mechanisms in Neuroblastoma Metastasis
Restricted (Penn State Only)
- Author:
- Dhamdhere, Mayura
- Graduate Program:
- Biomedical Sciences
- Degree:
- Doctor of Philosophy
- Document Type:
- Dissertation
- Date of Defense:
- May 23, 2024
- Committee Members:
- Vladimir Spiegelman, Co-Chair & Dissertation Advisor
Hong-Gang Wang, Co-Chair of Committee
Todd Schell, Major Field Member
Lisa Shantz, Program Head/Chair
Jeffrey Sundstrom, Outside Unit & Field Member - Keywords:
- Neuroblastoma
Neuroblastoma Metastasis
Extracellular vesicles
immunocompetent NB models
pre-metastatic niche
NB tumor microenvironment
NB immunotherapy - Abstract:
- Neuroblastoma (NB) is the most common extracranial solid pediatric cancer, and due to being highly metastatic in nature, it is among the leading causes of cancer-related deaths in children. Approximately half of all NB patients are classified to have high-risk (HR) metastatic disease. Despite treatments with recent multi-modal radio-chemo-immunotherapy, 5-year survival in the HR group still remains <50%, due to therapeutic resistance and increased disease relapse. Thus, it is essential to identify novel therapeutic targets or design combination strategies to address these clinically resistant NBs. Among the few well-known oncogenic drivers of NB (MYCN amplification, chromosomal aberrations - 1p loss, 11q loss, 17q gain), 17q gain is the prominently observed one and is independently associated with poor survival. IGF2BP1 is a candidate oncogene located at 17q locus, and is found to be upregulated in patients with advanced-stage NBs. Our work identified IGF2BP1 as a novel oncogenic driver in NB progression, and established its role in promoting NB metastasis. Importantly, for the first time, we reveal the significance of small extracellular vesicles (EVs) in NB progression, and determine the pro-metastatic function of IGF2BP1 in an EV-mediated manner. We discovered two novel targets of IGF2BP1 – SEMA3A and SHMT2, and reveal the associated mechanisms in NB metastasis. Mechanistically, IGF2BP1 directly binds and stabilizes SHMT2/SEMA3A mRNA, thereby increasing the expression of SHMT2/SEMA3A in NB cells, and eventually increasing their protein levels in the secreted NB-EVs. IGF2BP1-governed SEMA3A and SHMT2 in the EVs, regulate the formation of pro-metastatic microenvironment at potential metastatic organs. Our work further establishes the importance of IGF2BP1-SEMA3A axis in NB, and the pro-metastatic role of SEMA3A via Nrp1-receptor activation in recipient cells/organs. We demonstrate that EV-SEMA3A induces a pro-metastatic microenvironment by promoting immunosuppressive cell infiltrate and cancer-associated/metastasis associated fibroblast activation at potential metastatic organs. Targeting SEMA3A-Nrp1 signaling prevents the formation of this pro-metastatic, tumor-supportive microenvironment, and reduces tumor growth and metastasis in HR-NB mouse model. Furthermore, we demonstrate the function of IGF2BP1 in regulating the primary tumor microenvironment in HR-NB. We show that IGF2BP1 mediates an immunosuppressive microenvironment in NB tumors by promoting the accumulation of immunosuppressive immune cells. Depletion of IGF2BP1, favorably alters the NB tumor microenvironment to an immunogenic type. Finally, we demonstrate that depleting IGF2BP1 in NB cells, along with the anti-GD2 immunotherapy further has a synergistic effect on promoting an immunogenic TME, and delays tumor growth in mice. Besides, a lack of appropriate metastasis models in NB, has limited the investigations on underlying biology of HR-NB metastasis in the past. Our study fills this gap by providing newly-developed appropriate syngeneic HR-NB cell lines that are highly metastatic in nature and feature metastasis to the prominent organs observed in HR-NB patients. The newly-developed and validated cell lines are relevant and promising tools for HR-NB metastasis and microenvironment studies, in an immunocompetent system. Together my dissertation work, apart from providing novel tools for NB studies, establish the importance of EVs in NB, determine the EV-mediated IGF2BP1-SEMA3A mechanism in NB metastasis, and identify the role of IGF2BP1 in NB TME regulation; thus, providing avenues for translating these findings for future NB treatments.