Alkylpurine–DNA–N-glycosylase confers resistance to temozolomide in xenograft models of glioblastoma multiforme and is associated with poor survival in patients

Abstract

Glioblastoma multiforme (GBM) is the most common and lethal of all gliomas. The current standard of care includes surgery followed by concomitant radiation and chemotherapy with the DNA alkylating agent temozolomide (TMZ). O 6 -methylguanine-DNA methyltransferase (MGMT) repairs the most cytotoxic of lesions generated by TMZ, O 6 -methylguanine. Methylation of the MGMT promoter in GBM correlates with increased therapeutic sensitivity to alkylating agent therapy. However, several aspects of TMZ sensitivity are not explained by MGMT promoter methylation. Here, we investigated our hypothesis that the base excision repair enzyme alkylpurine-DNA-N-glycosylase (APNG), which repairs the cytotoxic lesions N 3 -methyladenine and N 7 -methylguanine, may contribute to TMZ resistance. Silencing of APNG in established and primary TMZresistant GBM cell lines endogenously expressing MGMT and APNG attenuated repair of TMZ-induced DNA damage and enhanced apoptosis. Reintroducing expression of APNG in TMZ-sensitive GBM lines conferred resistance to TMZ in vitro and in orthotopic xenograft mouse models. In addition, resistance was enhanced with coexpression of MGMT. Evaluation of APNG protein levels in several clinical datasets demonstrated that in patients, high nuclear APNG expression correlated with poorer overall survival compared with patients lacking APNG expression. Loss of APNG expression in a subset of patients was also associated with increased APNG promoter methylation. Collectively, our data demonstrate that APNG contributes to TMZ resistance in GBM and may be useful in the diagnosis and treatment of the disease.

Theoretical considerations for thresholds in chemical carcinogenesis

“…As an example, mass 62 spectrometry data quantified N7-methylguanine (N7MeG) adducts 63 as biomarkers of exposure. A dose of 0.025 mg/ml N-methyl-N- 64 nitrosourea (MNU) caused 9.1 N7MeG adducts/10 8 nucleotides 65 and a significant increase in mutant frequency in human 66 lymphoblastoid cells, which can be considered as greater than 67 the PoD for MNU-induced point mutations [5]. 299 Current thinking pertains that PoDs for mutations are 300 dependent upon the enzymology of the specific repair process 301 required to remove the respective adducts following treatment, 302 whereby the dose that causes saturation of repair function is the 303 PoD, and mutations will then accrue linearly at higher doses.…”

DNA repair mechanisms and their clinical impact in glioblastoma

“…<H2>5.2. Strategies to inhibit base excision repair In 2012, Agnihotri et al reported a positive correlation between the expression levels of the BER factor MPG and the TMZ IC50 in adult GBM cells, where MPG was found topromote TMZ resistance(99). Later, the authors identified a similar correlation in pediatric GBM lines and also uncovered MPG and the apurinic/apyrimidinic (AP) endonuclease 1 (APEX1) amongst TMZ sensitizers in a siRNA screen with TMZ-resistant pediatric GBM cell lines(98).…”

“…This small molecule inhibitor binds the abasic sites generated by MPG, thereby preventing further processing. Compared to treatment with either molecule alone, the combination of methoxyamine plus TMZ dramatically increased survival of mice injected intracranially with SJG2 cells, as well as in an orthotopic PDX mouse model generated from pediatric GBM primary cells (98,99).…”

miR-20a mediates temozolomide-resistance in glioblastoma cells via negatively regulating LRIG1 expression

“…Many researches on TMZ resistance have identified several differential mechanisms involved in the step of obtaining the resistance, such as promoter methylation of the O6-methylguanine-DNA methyltransferase (MGMT) gene, activation of the base excision DNA repair pathway [31,32] and phospho-inostide-3-kinase mediated up-regulation of the HOXA9/HOXA10 genes [33]. However, several aspects of TMZ sensitivity still cannot be explained and most patients eventually recur often due to resistance to TMZ.…”