Dataset Information

Targeting glioma stem cells in vivo by a G-quadruplex-stabilizing synthetic macrocyclic hexaoxazole.

ABSTRACT: G-quadruplex (G4) is a higher-order nucleic acid structure that is formed by guanine-rich sequences. G4 stabilization by small-molecule compounds called G4 ligands often causes cytotoxicity, although the potential medicinal impact of this effect has not been fully established. Here we demonstrate that a synthetic G4 ligand, Y2H2-6M(4)-oxazole telomestatin derivative (6OTD), limits the growth of intractable glioblastoma (grade IV glioma) and glioma stem cells (GSCs). Experiments involving a human cancer cell line panel and mouse xenografts revealed that 6OTD exhibits antitumor activity against glioblastoma. 6OTD inhibited the growth of GSCs more potently than it did the growth of differentiated non-stem glioma cells (NSGCs). 6OTD caused DNA damage, G1 cell cycle arrest, and apoptosis in GSCs but not in NSGCs. These DNA damage foci tended to colocalize with telomeres, which contain repetitive G4-forming sequences. Compared with temozolomide, a clinical DNA-alkylating agent against glioma, 6OTD required lower concentrations to exert anti-cancer effects and preferentially affected GSCs and telomeres. 6OTD suppressed the intracranial growth of GSC-derived tumors in a mouse xenograft model. These observations indicate that 6OTD targets GSCs through G4 stabilization and promotion of DNA damage responses. Therefore, G4s are promising therapeutic targets for glioblastoma.

SUBMITTER: Nakamura T

PROVIDER: S-EPMC5472576 | biostudies-literature | 2017 Jun

REPOSITORIES: biostudies-literature

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Targeting glioma stem cells in vivo by a G-quadruplex-stabilizing synthetic macrocyclic hexaoxazole.

Nakamura Takahiro T Okabe Sachiko S Yoshida Haruka H Iida Keisuke K Ma Yue Y Sasaki Shogo S Yamori Takao T Shin-Ya Kazuo K Nakano Ichiro I Nagasawa Kazuo K Seimiya Hiroyuki H

Scientific reports 20170615 1

G-quadruplex (G4) is a higher-order nucleic acid structure that is formed by guanine-rich sequences. G4 stabilization by small-molecule compounds called G4 ligands often causes cytotoxicity, although the potential medicinal impact of this effect has not been fully established. Here we demonstrate that a synthetic G4 ligand, Y2H2-6M(4)-oxazole telomestatin derivative (6OTD), limits the growth of intractable glioblastoma (grade IV glioma) and glioma stem cells (GSCs). Experiments involving a human ...[more]

PMID: 28620243

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Project description:Abstract The survival advantage of glioma stem cells (GSCs) represents a critical mechanism for growth, therapy resistance and recurrence in glioblastoma. So far, targeting GSCs has not been highly specific, since these cells co-opt the normal developmental signaling pathways. We have demonstrated that the activated CDK5-CREB1 signaling axis regulates GSC self-renewal and also promotes radiation-resistance. Thus targeting CDK5 signaling is highly rational, yet there are challenges. Most of the available CDK5 inhibitors also target other CDKs non-specifically. In collaboration with The Center for Molecular Evolution and Drug Discovery, we are developing novel CDK5 inhibitors that are highly potent and specific. METHODOLOGY: The CKD5-p25 crystal structure (pdb code 1UNL) was used to conduct a virtual high throughput screen (vHTS). A library of 10 million commercially available compounds which had been filtered to ensure they possessed good drug-like properties was screened against the crystal structure. The top 33 compounds based on their predicted target binding, synthetic feasibility and availability were tested in an in vitro kinase assay to measure CDK5 inhibition. RESULTS: Of the 33 potential hit, 11 compounds showed a CDK5 inhibition of < 50 µM. These 11 hits represent 4 distinct chemical scaffolds. Two of them have IC50 < 1 µM, with one compound having an IC50 < 0.4 µM. The vHTS and subsequent in vitro testing have therefore confirmed the identification of several new series of potent CDK5 hit compounds. We are now characterizing the kinase selectivity of our different hit series and evaluating their activity in cell-based assays. This will help focus efforts on the most promising 1–2 scaffolds for further medicinal chemistry optimization to improve the compounds’ potency, selectivity and brain penetration. Ultimately, our optimized compounds will be tested in GBM models to demonstrate their effectiveness in inhibiting CDK5 as a new approach for treating GBM.

Project description:BackgroundGlioblastoma is the most aggressive and most lethal primary brain tumor in the adulthood. Current standard therapies are not curative and novel therapeutic options are urgently required. Present knowledge suggests that the continued glioblastoma growth and recurrence is determined by glioblastoma stem-like cells (GSCs), which display self-renewal, tumorigenic potential, and increased radio- and chemo-resistance. The G-quadruplex ligand RHPS4 displays in vitro radiosensitizing effect in GBM radioresistant cells through the targeting and dysfunctionalization of telomeres but RHPS4 and Ionizing Radiation (IR) combined treatment efficacy in vivo has not been explored so far.MethodsRHPS4 and IR combined effects were tested in vivo in a heterotopic mice xenograft model and in vitro in stem-like cells derived from U251MG and from four GBM patients. Cell growth assays, cytogenetic analysis, immunoblotting, gene expression and cytofluorimetric analysis were performed in order to characterize the response of differentiated and stem-like cells to RHPS4 and IR in single and combined treatments.ResultsRHPS4 administration and IR exposure is very effective in blocking tumor growth in vivo up to 65 days. The tumor volume reduction and the long-term tumor control suggested the targeting of the stem cell compartment. Interestingly, RHPS4 treatment was able to strongly reduce cell proliferation in GSCs but, unexpectedly, did not synergize with IR. Lack of radiosensitization was supported by the GSCs telomeric-resistance observed as the total absence of telomere-involving chromosomal aberrations. Remarkably, RHPS4 treatment determined a strong reduction of CHK1 and RAD51 proteins and transcript levels suggesting that the inhibition of GSCs growth is determined by the impairment of the replication stress (RS) response and DNA repair.ConclusionsWe propose that the potent antiproliferative effect of RHPS4 in GSCs is not determined by telomeric dysfunction but is achieved by the induction of RS and by the concomitant depletion of CHK1 and RAD51, leading to DNA damage and cell death. These data open to novel therapeutic options for the targeting of GSCs, indicating that the combined inhibition of cell-cycle checkpoints and DNA repair proteins provides the most effective means to overcome resistance of GSC to genotoxic insults.

Dataset Information

Targeting glioma stem cells in vivo by a G-quadruplex-stabilizing synthetic macrocyclic hexaoxazole.

Publications

Targeting glioma stem cells in vivo by a G-quadruplex-stabilizing synthetic macrocyclic hexaoxazole.

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