Inhibition of DNA-PK potentiates the synergistic effect of NK314 and etoposide combination on human glioblastoma cells

Abstract

Etoposide (VP-16) is the topoisomerase 2 (Top2) inhibitor used for treating of glioma patients however at high dose with serious side effects. It induces DNA double-strand breaks (DSBs). These DNA lesions are repaired by non-homologous DNA end joining (NHEJ) mediated by DNA-dependent protein kinase (DNA-PK). One possible approach to decrease the toxicity of etoposide is to reduce the dose while maintaining the anticancer potential. It could be achieved through combined therapy with other anticancer drugs. We have assumed that this objective can be obtained by (1) a parallel topo2 α inhibition and (2) sensitization of cancer cells to DSBs. In this work we investigated the effect of two Top2 inhibitors NK314 and VP-16 in glioma cell lines (MO59 K and MO59 J) sensitized by DNA-PK inhibitor, NU7441. Cytotoxic effect of VP-16, NK314 alone and in combination on human glioblastoma cell lines, was assessed by a colorimetric assay. Genotoxic effect of anticancer drugs in combination with NU7441 was assessed by comet assay. Cell cycle distribution and apoptosis were analysed by flow cytometry. Compared with VP-16 or NK314 alone, the combined treatment significantly inhibited cell proliferation. Combination treatment was associated with a strong accumulation of DSBs, modulated cell cycle phases distribution and apoptotic cell death. NU7441 potentiated these effects and additionally postponed DNA repair. Our findings suggest that NK314 could overcome resistance of MO59 cells to VP-16 and NU7441 could serve as sensitizer to VP-16/ NK314 combined treatment. The combined tripartite approach of chemotherapy could reduce the overall toxicity associated with each individual therapy, while concomitantly enhancing the anticancer effect to treat human glioma cells. Thus, the use of a tripartite combinatorial approach could be promising and more efficacious than mono therapy or dual therapy to treat and increase the survival of the glioblastoma patients.

The anterior gradient homologue 2 (AGR2) co-localises with the glucose-regulated protein 78 (GRP78) in cancer stem cells, and is critical for the survival and drug resistance of recurrent glioblastoma: in situ and in vitro analyses

“…The expression of AGR2 remained high in drug-resistant cells following treatments with a range of clinically applied drugs that utilize different modes of action [ 76 – 80 ]. This suggests that the presence of AGR2 is critical for glioblastoma cell survival, regardless of drug type.…”

Inhibition of Protein Disulfide Isomerase (PDIA1) Leads to Proteasome-Mediated Degradation of Ubiquitin-like PHD and RING Finger Domain-Containing Protein 1 (UHRF1) and Increased Sensitivity of Glioblastoma Cells to Topoisomerase II Inhibitors

“…These results indicate that PDIA1 inhibition impacts DNA damage response pathways that are required to repair lesions induced predominantly by Top-II poisons such as DSBs. Similar to radiation, Top-II poisons induce replication-independent DSBs that are repaired by two pathways, homologous recombination (HR) or NHEJ, where NHEJ is the predominant repair pathway. , In contrast, Top-I poisons result primarily in single-strand breaks (SSBs) that are converted to DSBs after DNA replication, so Top-I-induced DNA damage is primarily repaired through SSB repair pathways in addition to DSB repair pathways if DSBs were generated. , On the other hand, alkylating and crosslinking agents such as TMZ and oxaliplatin are mainly repaired through SSB repair pathways such as direct repair, BER, mismatch repair (MMR), and NER. , Interestingly, bepristat-2a was synergistic with NU7441, an inhibitor of DNA-PK which plays an essential role in the NHEJ pathway. − Previous reports showed that DNA-PK inhibition increases sensitivity of tumor cells to Top-II inhibitors. − In contrast, bepristat-2a was not synergistic with inhibitors of PARP enzymes which are best known for their function in SSB repair pathways such as BER and NER. , Overall, our data support the hypothesis that PDIA1 inhibition impacts DSB repair pathways mainly the NHEJ arm leading to increased sensitivity to Top-II inhibitors.…”

Perspective on the Use of DNA Repair Inhibitors as a Tool for Imaging and Radionuclide Therapy of Glioblastoma

“…Finally, based on preclinical data alone, SU11752, KU0060648, NU7026, and NU7441 have been put forward as glioma targeted drugs, either as single agents or in combination regimens (RT/TMZ/topoisomerase II inhibitors) [ 261 , 262 , 263 , 264 , 265 ].…”