MEK inhibitor sensitivity and resistance in MAPK-altered diffuse midline gliomas
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Embargo End Date
2025-12-02
ICR Authors
Authors
Das, M
Document Type
Thesis or Dissertation
Date
2025-06-02
Date Accepted
Abstract
Targeted therapies inhibiting MAPK pathway activation have recently been approved for
use in paediatric low-grade glioma. Pre-clinical evidence suggests these therapies could
be translated in MAPK-altered diffuse midline glioma (DMG). In this thesis, I determine the
frequency of MAPK pathway alterations in DMG and assess the utility of the MEK inhibitor
trametinib in patient-derived models. Both PIK3R1-mutant B181-3D and the NF1-mutant
B184-3D models show ~500-fold increased trametinib sensitivity compared to their
isogenic non-MAPK-altered counterparts, suggesting MAPK pathway alterations
specifically confer sensitivity to trametinib. Furthermore, drug screening revealed antiapoptosis
inhibitors could sensitize non-MAPK-altered B181-2D to trametinib. Three
independent trametinib resistant cell lines were also generated from parental B181-3D
cells, drug screening of which exhibited 100% increased sensitivity to progesterone
compared to parental cells. Trametinib resistance was also investigated in the BRAFmutant
B169-2D model using coupled lineage tracing and single cell RNA sequencing
(scRNA-seq). A subset of clones was reproducibly enriched across several technical
replicate experiments, suggesting deterministic patterns of selection and expansion.
Linking clonal identity with transcriptional output suggests resistant cells are phenotypically
plastic and shuttle between astrocyte-like and neural precursor-like cell states. Previously
reported trametinib resistance-conferring MAP2K1 mutations were also detected in this
model and seem to arise from phenotypic plasticity that is reminiscent of drug tolerant
persisters (DTP). In the same model, a subpopulation of resistant cells were distinctly
characterized by epithelial to mesenchymal transition, suggesting two transcriptionally and
clonally disparate resistance mechanisms occur simultaneously. Optimization experiments
were also conducted to allow for downstream retrieval of resistant clones from pretreatment
barcoded cells to determine whether resistance was pre-existing. Overall, I
investigated trametinib resistance in two MAPK-altered DMG models by utilizing
conventional resistance modelling techniques and development of novel tools that shed
light on the role of clonal evolution in resistance. To our best knowledge, this coupling of
lineage tracing and scRNA-seq is the first time this technique has been used to study
targeted therapy resistance in DMG.
Citation
2025
DOI
Source Title
Publisher
Institute of Cancer Research (University Of London)
ISSN
eISSN
Collections
Research Team
Glioma Team