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MGMT-inhibitor in combination with TGF-βRI inhibitor or CDK 4/6 inhibitor increases temozolomide sensitivity in temozolomide-resistant glioblastoma cells

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Abstract

Background

Glioblastoma (GB) remains an incurable and deadly brain malignancy that often proves resistant to upfront treatment with temozolomide. Nevertheless, temozolomide remains the most commonly prescribed FDA-approved chemotherapy for GB. The DNA repair protein methylguanine-DNA methyl transferase (MGMT) confers resistance to temozolomide. Unsurprisingly temozolomide-resistant tumors tend to possess elevated MGMT protein levels or lack inhibitory MGMT promotor methylation. In this study, cultured human temozolomide resistance GB (43RG) cells were introduced to the MGMT inhibitor O6-benzylguanine combined with temozolomide and either LY2835219 (CDK 4/6 inhibitor) or LY2157299 (TGF-βRI inhibitor) seeking to overcome GB treatment resistance.

Methods

Treatment effects were assessed using MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assay, western blot, cell viability, and cell cycle progression.

Results

Our in vitro study demonstrated that sequential treatment of O6-Benzylguanine with either LY2385219 or LY2157299-enhanced temozolomide enhanced sensitivity in MGMT+ 43RG cells. Importantly, normal human neurons and astrocytes remained impervious to the drug therapies under these conditions. Furthermore, LY2835219 has additional anti-proliferative effects on cell cycling, including induction of an RB-associated G (1) arrest via suppression of cyclin D-CDK4/6-Rb pathway. LY2157299 enhances anti-tumor effect by disrupting TGF-β–dependent HIF-1α signaling and by activating both Smad and PI3K-AKT pathways towards transcription of S/G2 checkpoints.

Conclusion

This study establishes the groundwork for the development of a combinatorial pharmacologic approach by using either LY2385219 or LY2157299 inhibitor plus O6-Benzylguanine to augment temozolomide response in temozolomide-resistant GB cells.

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Acknowledgements

Eli Lilly and Company supplied LY2157299 (TGF-β kinase inhibitor) and LY2835219 (CDK4/6 inhibitor) for this in vitro study. The results of this research and any intellectual property arising from this research are subject to existing rights and obligations to a third party, Eli Lilly and Company.

Funding

Completion of this project was made possible in part by the Funds from the Department of Neurosurgery and partial support to MA from AANS/CNS Section on Pediatric Neurological Surgery & NREF 2020-21 Research Fellowship Grant.

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Author notes

  1. F. C. Henderson Jr and M. Alshareef have contributed equally.

Authors and Affiliations

  1. Department of Neurosurgery (Divisions of Neuro-Oncology) and MUSC Brain and Spine Tumor Program CSB 310, Medical University of South Carolina at Charleston, Charleston, SC, 29425, USA

    A. Das, F. C. Henderson Jr, M. Alshareef, G. B. F. Porto, I. Kanginakudru, L. K. Infinger, W. A. Vandergrift III, S. M. Lindhorst, A. K. Varma, S. J. Patel & D. Cachia

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Correspondence to A. Das.

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All used human cell cultures have been preserved because they have characteristics that are vital to the advancement of scientific research and development. These are freely available as public resources by Mayo Clinic and ScienCell Research Laboratories. Therefore, local ethics approval was not needed.

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Das, A., Henderson, F.C., Alshareef, M. et al. MGMT-inhibitor in combination with TGF-βRI inhibitor or CDK 4/6 inhibitor increases temozolomide sensitivity in temozolomide-resistant glioblastoma cells. Clin Transl Oncol 23, 612–619 (2021). https://doi.org/10.1007/s12094-020-02456-x

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