Investigating the mode of action of NMT inhibitors in MYCN-amplified neuroblastoma
File(s)
Author(s)
Zhang, James
Type
Thesis
Abstract
The N-myristoyltransferase (NMT) proteins catalyse the irreversible fatty acylation of >200 substrates with myristate on N-terminal glycines, modulating multiple protein properties including localisation, stability, and activity. NMTs are promising targets in cancer, however an understanding of the mechanism of action of NMT inhibitors (NMTi) is still emerging. It has been shown that deregulated MYC sensitises lymphoma cells to NMT inhibition in part through effects on respiratory complex I. However, the applicability and biological consequences of NMTi in other cancer contexts is still unclear.
In this thesis, the applicability of NMTi in neuroblastoma, the most common paediatric extracranial solid tumour, was investigated. Amplification of MYCN, the neural analogue of MYC, is both prevalent and associated with poor outcome in neuroblastoma. Using a panel of neuroblastoma cell lines, MYCN amplification was found to sensitise cells to NMTi. Multiple -omics based analyses of the effects of NMT inhibition in a MYCN-regulatable isogenic cell line revealed that NMT inhibition affects multiple pathways, with effects more rapidly induced in high MYCN cells. In particular, strong depletion of respiratory complex I was identified, associated with loss of the NMT substrate NDUFAF4. To validate this association in MYCN-amplified neuroblastoma, a NDUFAF4 G2A knock-in mutant was generated, which confirmed the importance of NDUFAF4 N-myristoylation alone on oxidative phosphorylation and indicated that the effects of NMTi on oxidative phosphorylation are mediated primarily through respiratory complex I. However, this mutation did not alter sensitivity to NMTi, indicating that the cytotoxicity of NMTi in MYCN-amplified neuroblastoma is not mediated primarily by effects on NDUFAF4, but is likely instead by simultaneous modulation of multiple biological pathways.
In total, this work suggests that NMTi would be effective in MYCN-amplified neuroblastomas, expanding the possible clinical applications of this compound class, and identifies pathways through which NMTi may exert its cytotoxic effects in neuroblastoma.
In this thesis, the applicability of NMTi in neuroblastoma, the most common paediatric extracranial solid tumour, was investigated. Amplification of MYCN, the neural analogue of MYC, is both prevalent and associated with poor outcome in neuroblastoma. Using a panel of neuroblastoma cell lines, MYCN amplification was found to sensitise cells to NMTi. Multiple -omics based analyses of the effects of NMT inhibition in a MYCN-regulatable isogenic cell line revealed that NMT inhibition affects multiple pathways, with effects more rapidly induced in high MYCN cells. In particular, strong depletion of respiratory complex I was identified, associated with loss of the NMT substrate NDUFAF4. To validate this association in MYCN-amplified neuroblastoma, a NDUFAF4 G2A knock-in mutant was generated, which confirmed the importance of NDUFAF4 N-myristoylation alone on oxidative phosphorylation and indicated that the effects of NMTi on oxidative phosphorylation are mediated primarily through respiratory complex I. However, this mutation did not alter sensitivity to NMTi, indicating that the cytotoxicity of NMTi in MYCN-amplified neuroblastoma is not mediated primarily by effects on NDUFAF4, but is likely instead by simultaneous modulation of multiple biological pathways.
In total, this work suggests that NMTi would be effective in MYCN-amplified neuroblastomas, expanding the possible clinical applications of this compound class, and identifies pathways through which NMTi may exert its cytotoxic effects in neuroblastoma.
Version
Open Access
Date Issued
2024-12-17
Date Awarded
2025-06-01
Copyright Statement
Attribution-NonCommercial 4.0 International Licence (CC BY-NC)
License URL
Advisor
Tate, Edward
Sponsor
Cancer Research UK
Grant Number
C309/A31546
Publisher Department
Department of Chemistry
Publisher Institution
Imperial College London
Qualification Level
Doctoral
Qualification Name
Doctor of Philosophy (PhD)