Temozolomide-associated inflammatory-repair remodeling in recurrent glioblastoma exposes a ROCK-linked therapeutic vulnerability
- 1Department of Neurological Surgery, Northwestern University Feinberg School of Medicine, Chicago, IL 60611, USA;Lou and Jean Malnati Brain Tumor Institute of th
Abstract
Background: Glioblastoma (GBM) adapts to therapy through coordinated malignant-cell and microenvironmental responses, but the mechanisms linking treatment-associated inflammation to tumor-cell phenotypic plasticity remain poorly defined. Here, we investigated whether preoperative temozolomide exposure is associated with inflammatory-repair remodeling and a ROCK-linked therapeutic vulnerability in recurrent GBM using a window-of-opportunity clinical trial cohort. Methods: We integrated patient-resolved single-cell transcriptomics, spatial RNA profiling and multiplex protein imaging with experimental perturbations and orthotopic glioblastoma models. In the presurgical window-of-opportunity subgroup of NCT05236036, four patients with recurrent glioblastoma received additional preoperative temozolomide (TMZ; 150 mg/m2/day for 5 days) before resection and were compared with three recurrent comparators who did not receive additional preoperative TMZ. Results: Across seven tumors, malignant-cell inflammatory activity covaried with integrin-binding and wound-healing programs (median partial {rho} = 0.45 and 0.42, respectively). Tumors exposed to preoperative TMZ showed enrichment of inflammatory, chemokine, adhesion and wound-healing programs (GSEA q < 0.05), with the direction of enrichment preserved in all leave-one-patient-out analyses. In two tumors exposed to preoperative TMZ, CXCL12 localized to vascular territories, whereas chemokine and wound-repair programs increased near injury-reference regions. Across seven specimens analyzed by multiplex protein imaging, MYL9 abundance correlated with local CXCL12 ({rho} = 0.61), phosphorylated MYPT1 ({rho} = 0.48) and nuclear phosphorylated STAT3 ({rho} = 0.45), with positive associations in every specimen. Experimentally, TMZ increased CXCL8 and reactive-state markers and enhanced subsequent scratch closure, while CXCL8 and CXCL12 increased MLC2 phosphorylation. Fasudil attenuated TNF-NF-{kappa}B, inflammatory-response and IFN-{gamma}-response programs in TMZ-treated cells and reduced scratch closure and p-MLC2 in complementary assays. Across three orthotopic models, fasudil plus TMZ prolonged survival versus TMZ alone (model-stratified HR, 0.21; 95% CI, 0.07-0.61; P = 0.001). Conclusions: These findings link treatment-associated inflammatory-repair programs with cytoskeletal remodeling and support further translational evaluation of fasudil plus TMZ in glioblastoma.
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BibTeX
@misc{chojak2026temozolomideassociated,
title = {{Temozolomide-associated inflammatory-repair remodeling in recurrent glioblastoma exposes a ROCK-linked therapeutic vulnerability}},
author = {R. Chojak and U. H. Faisal and P. U. Kumthekar and N. B. Drewes and L. Koutah and J. Jia and J. R. Turunen and M. C. O'Shea and V. Falkowski and A. Akhtar and S. Wang and J. Chen and S. Ahmed and H. Kazi and B. Auffinger and P. Jamshidi and K. McCortney and A. M. Catezone and M. C. Tate and R. Stupp and M. S. Lesniak and J. Miska and K. S. Dixit and D. Primdahl and D. H. Heiland and A. M. Sonabend and A. U. Ahmed},
howpublished = {bioRxiv},
year = {2026},
month = sep,
doi = {10.64898/2026.09.20.752931},
url = {https://www.biorxiv.org/content/10.64898/2026.09.20.752931v1},
note = {CC BY},
}
RIS
TY - UNPB
TI - Temozolomide-associated inflammatory-repair remodeling in recurrent glioblastoma exposes a ROCK-linked therapeutic vulnerability
AU - R. Chojak
AU - U. H. Faisal
AU - P. U. Kumthekar
AU - N. B. Drewes
AU - L. Koutah
AU - J. Jia
AU - J. R. Turunen
AU - M. C. O'Shea
AU - V. Falkowski
AU - A. Akhtar
AU - S. Wang
AU - J. Chen
AU - S. Ahmed
AU - H. Kazi
AU - B. Auffinger
AU - P. Jamshidi
AU - K. McCortney
AU - A. M. Catezone
AU - M. C. Tate
AU - R. Stupp
AU - M. S. Lesniak
AU - J. Miska
AU - K. S. Dixit
AU - D. Primdahl
AU - D. H. Heiland
AU - A. M. Sonabend
AU - A. U. Ahmed
PY - 2026
DA - 2026/09/20
PB - bioRxiv
DO - 10.64898/2026.09.20.752931
UR - https://www.biorxiv.org/content/10.64898/2026.09.20.752931v1
AB - Background: Glioblastoma (GBM) adapts to therapy through coordinated malignant-cell and microenvironmental responses, but the mechanisms linking treatment-associated inflammation to tumor-cell phenotypic plasticity remain poorly defined. Here, we investigated whether preoperative temozolomide exposure is associated with inflammatory-repair remodeling and a ROCK-linked therapeutic vulnerability in recurrent GBM using a window-of-opportunity clinical trial cohort. Methods: We integrated patient-resolved single-cell transcriptomics, spatial RNA profiling and multiplex protein imaging with experimental perturbations and orthotopic glioblastoma models. In the presurgical window-of-opportunity subgroup of NCT05236036, four patients with recurrent glioblastoma received additional preoperative temozolomide (TMZ; 150 mg/m2/day for 5 days) before resection and were compared with three recurrent comparators who did not receive additional preoperative TMZ. Results: Across seven tumors, malignant-cell inflammatory activity covaried with integrin-binding and wound-healing programs (median partial {rho} = 0.45 and 0.42, respectively). Tumors exposed to preoperative TMZ showed enrichment of inflammatory, chemokine, adhesion and wound-healing programs (GSEA q < 0.05), with the direction of enrichment preserved in all leave-one-patient-out analyses. In two tumors exposed to preoperative TMZ, CXCL12 localized to vascular territories, whereas chemokine and wound-repair programs increased near injury-reference regions. Across seven specimens analyzed by multiplex protein imaging, MYL9 abundance correlated with local CXCL12 ({rho} = 0.61), phosphorylated MYPT1 ({rho} = 0.48) and nuclear phosphorylated STAT3 ({rho} = 0.45), with positive associations in every specimen. Experimentally, TMZ increased CXCL8 and reactive-state markers and enhanced subsequent scratch closure, while CXCL8 and CXCL12 increased MLC2 phosphorylation. Fasudil attenuated TNF-NF-{kappa}B, inflammatory-response and IFN-{gamma}-response programs in TMZ-treated cells and reduced scratch closure and p-MLC2 in complementary assays. Across three orthotopic models, fasudil plus TMZ prolonged survival versus TMZ alone (model-stratified HR, 0.21; 95% CI, 0.07-0.61; P = 0.001). Conclusions: These findings link treatment-associated inflammatory-repair programs with cytoskeletal remodeling and support further translational evaluation of fasudil plus TMZ in glioblastoma.
ER -