A novel approach to investigate the combinatorial effects of TLK1 (Tousled-Like Kinase1) inhibitors with Temozolomide for glioblastoma therapy.

ABSTRACT

Glioblastoma multiforme (GBM) is an aggressive, incurable brain tumor with poor prognosis and limited treatment options. Temozolomide (TMZ) is the standard chemotherapeutic treatment for GBM, but its efficacy has drawn strong criticism from clinicians due to short survival gains and frequent relapses. One critical limitation of TMZ therapy is the hyperactivation of DNA repair pathways, which over time neutralizes the cytotoxic effects of TMZ, thus highlighting the urgent need for new treatment approaches. Addressing this, our study explores the therapeutic potential of in-house-designed phenothiazine-based Tousled-like kinase-1 (TLK1) inhibitors for GBM treatment. TLK1, overexpressed in GBM, plays a role in DNA repair. Phenothiazines are known to cross the blood-brain barrier (BBB). Among all molecules, J54 was identified as a potential lead molecule with improved cytotoxicity. In the context of O6-methylguanine-DNA methyltransferase (MGMT)-deficient GBM cells, the combined administration of phenothiazines and TMZ exhibited a collective reduction in clonogenic growth, coupled with anti-migratory and anti-invasion effects. Conversely, in MGMT-proficient cells, phenothiazine monotherapy alone showed reduced clonogenic growth, along with anti-migratory and anti-invasion effects. Notably, a synergistic increase in γH2AX levels and concurrent attenuation of DNA repair upon combinatorial exposure to TMZ and J54 were observed, implying increased cytotoxicity due to sustained DNA strand breaks. Overall, this study provides new insights into TLK1 inhibition for GBM therapy. Collectively, these findings indicate that TLK1 is one of the upregulated kinases in GBM and phenothiazine-based TLK1 inhibitors could be a promising treatment option for GBM patients.