-
Neuro-oncology Apr 2023The goal was to determine whether the addition of temozolomide (TMZ) to the standard treatment of high-dose methotrexate (HD-MTX) and whole-brain radiotherapy (WBRT) for... (Randomized Controlled Trial)
Randomized Controlled Trial
BACKGROUND
The goal was to determine whether the addition of temozolomide (TMZ) to the standard treatment of high-dose methotrexate (HD-MTX) and whole-brain radiotherapy (WBRT) for primary central nervous system lymphoma (PCNSL) improves survival.
METHODS
An open-label, randomized, phase III trial was conducted in Japan, enrolling immunocompetent patients aged 20-70 years with histologically confirmed, newly diagnosed PCNSL. After administration of HD-MTX, patients were randomly assigned to receive WBRT (30 Gy) ± 10 Gy boost (arm A) or WBRT ± boost with concomitant and maintenance TMZ for 2 years (arm B). The primary endpoint was overall survival (OS).
RESULTS
Between September 29, 2014 and October 15, 2018, 134 patients were enrolled, of whom 122 were randomly assigned and analyzed. At the planned interim analysis, 2-year OS was 86.8% (95% confidence interval [CI]: 72.5-94.0%) in arm A and 71.4% (56.0-82.2%) in arm B. The hazard ratio was 2.18 (95% CI: 0.95-4.98), with the predicted probability of showing the superiority of arm B at the final analysis estimated to be 1.3%. The study was terminated early due to futility. O6-methylguanine-DNA methyltransferase (MGMT) promoter methylation status was measured in 115 tumors, and it was neither prognostic nor predictive of TMZ response.
CONCLUSIONS
This study failed to demonstrate the benefit of concomitant and maintenance TMZ in newly diagnosed PCNSL.
Topics: Humans; Temozolomide; Methotrexate; Disease-Free Survival; Lymphoma; Brain; Central Nervous System Neoplasms; Antineoplastic Agents, Alkylating
PubMed: 36334050
DOI: 10.1093/neuonc/noac246 -
Nucleic Acids Research May 2018Exposure to many endogenous and exogenous agents can give rise to DNA alkylation, which constitutes a major type of DNA damage. Among the DNA alkylation products, alkyl...
Exposure to many endogenous and exogenous agents can give rise to DNA alkylation, which constitutes a major type of DNA damage. Among the DNA alkylation products, alkyl phosphotriesters have relatively high frequencies of occurrence and are resistant to repair in mammalian tissues. However, little is known about how these lesions affect the efficiency and fidelity of DNA replication in cells or how the replicative bypass of these lesions is modulated by translesion synthesis DNA polymerases. In this study, we synthesized oligodeoxyribonucleotides containing four pairs (Sp and Rp) of alkyl phosphotriester lesions at a defined site, and examined how these lesions are recognized by DNA replication machinery in Escherichia coli cells. We found that the Sp diastereomer of the alkyl phosphotriester lesions could be efficiently bypassed, whereas the Rp counterparts moderately blocked DNA replication. Moreover, the Sp-methyl phosphotriester induced TT→GT and TT→GC mutations at the flanking TT dinucleotide site, and the induction of these mutations required Ada protein, which is known to remove efficiently the methyl group from the Sp-methyl phosphotriester. Together, our study provided a comprehensive understanding about the recognition of alkyl phosphotriester lesions by DNA replication machinery in cells, and revealed for the first time the Ada-dependent induction of mutations at the Sp-methyl phosphotriester site.
Topics: Alkylation; DNA Damage; DNA Repair; DNA Replication; DNA, Bacterial; DNA-Directed DNA Polymerase; Escherichia coli; Escherichia coli Proteins; Mutagenesis; Mutation; O(6)-Methylguanine-DNA Methyltransferase; Stereoisomerism; Transcription Factors
PubMed: 29514270
DOI: 10.1093/nar/gky140 -
Nucleic Acids Research May 2023The importance of non-canonical DNA structures such as G-quadruplexes (G4) and intercalating-motifs (iMs) in the fine regulation of a variety of cellular processes has...
The importance of non-canonical DNA structures such as G-quadruplexes (G4) and intercalating-motifs (iMs) in the fine regulation of a variety of cellular processes has been recently demonstrated. As the crucial roles of these structures are being unravelled, it is becoming more and more important to develop tools that allow targeting these structures with the highest possible specificity. While targeting methodologies have been reported for G4s, this is not the case for iMs, as evidenced by the limited number of specific ligands able to bind the latter and the total absence of selective alkylating agents for their covalent targeting. Furthermore, strategies for the sequence-specific covalent targeting of G4s and iMs have not been reported thus far. Herein, we describe a simple methodology to achieve sequence-specific covalent targeting of G4 and iM DNA structures based on the combination of (i) a peptide nucleic acid (PNA) recognizing a specific sequence of interest, (ii) a pro-reactive moiety enabling a controlled alkylation reaction, and (iii) a G4 or iM ligand orienting the alkylating warhead to the reactive residues. This multi-component system allows for the targeting of specific G4 or iM sequences of interest in the presence of competing DNA sequences and under biologically relevant conditions.
Topics: Alkylating Agents; Alkylation; DNA; G-Quadruplexes; Ligands; Light; Color
PubMed: 36971129
DOI: 10.1093/nar/gkad189 -
Biochimica Et Biophysica Acta.... Jun 2022PIMREG expression strongly correlates with cellular proliferation in both malignant and normal cells. Throughout embryo development, PIMREG expression is prominent in...
PIMREG expression strongly correlates with cellular proliferation in both malignant and normal cells. Throughout embryo development, PIMREG expression is prominent in the central nervous system. Recent studies have described elevated PIMREG expression in different types of tumors, which correlates with patient survival and tumor aggressiveness. Given the emerging significance of PIMREG in carcinogenesis and its putative role in the context of the nervous system, we investigated the expression and function of PIMREG in gliomas, the most common primary brain tumors. We performed an extensive analysis of PIMREG expression in tumors samples from glioma patients. We then assessed the effects of PIMREG silencing and overexpression on the sensitivity of glioblastoma cell lines treated with genotoxic agents commonly used for treating patients and assessed for treatment response, proliferation and migration. Our analysis shows that glioblastoma exhibits the highest levels of PIMREG expression among all cancers analyzed and that elevated PIMREG expression is a biomarker for glioma progression and patient outcome. Moreover, PIMREG is induced by genotoxic agents, and its silencing renders glioblastoma cells sensitive to temozolomide treatment and affects ATR- and ATM-dependent signaling. Our data demonstrate that PIMREG is involved in DNA damage response and temozolomide resistance of glioblastoma cells and further supports a role for PIMREG in tumorigenesis.
Topics: Antineoplastic Agents, Alkylating; Cell Line, Tumor; DNA Damage; Glioblastoma; Humans; Temozolomide
PubMed: 35301087
DOI: 10.1016/j.bbadis.2022.166382 -
The Journal of Biological Chemistry Jun 2018Endogenous metabolism, environmental exposure, and cancer chemotherapy can lead to alkylation of DNA. It has been well documented that, among the different DNA...
Endogenous metabolism, environmental exposure, and cancer chemotherapy can lead to alkylation of DNA. It has been well documented that, among the different DNA alkylation products, minor-groove -alkylthymidine (-alkyldT) lesions are inefficiently repaired. In the present study, we examined how seven -alkyldT lesions, with the alkyl group being a Me, Et, Pr, Pr, Bu, Bu, or Bu, are recognized by the DNA replication machinery in human cells. We found that the replication bypass efficiencies of these lesions decrease with increasing length of the alkyl chain, and that these lesions induce substantial frequencies of T→A and T→G mutations. Replication experiments using isogenic cells deficient in specific translesion synthesis (TLS) DNA polymerases revealed that the absence of polymerase η or polymerase ζ, but not polymerase κ or polymerase ι, significantly decreased both the bypass efficiencies and the mutation frequencies for those -alkyldT lesions carrying a straight-chain alkyl group. Moreover, the mutagenic properties of the -alkyldT lesions were influenced by the length and topology of the alkyl chain and by TLS polymerases. Together, our results provide important new knowledge about the cytotoxic and mutagenic properties of -alkyldT lesions, and illustrate the roles of TLS polymerases in replicative bypass of these lesions in human cells.
Topics: Alkylation; CRISPR-Cas Systems; DNA; DNA Damage; DNA Replication; DNA-Directed DNA Polymerase; Gene Editing; HEK293 Cells; Humans; Mutagenesis; Thymidine
PubMed: 29685891
DOI: 10.1074/jbc.RA118.003133 -
Journal of Hematology & Oncology Mar 2019Genomic instability is a feature of multiple myeloma (MM), and impairment in DNA damaging response (DDR) has an established role in disease pathobiology. Indeed, a...
BACKGROUND
Genomic instability is a feature of multiple myeloma (MM), and impairment in DNA damaging response (DDR) has an established role in disease pathobiology. Indeed, a deregulation of DNA repair pathways may contribute to genomic instability, to the establishment of drug resistance to genotoxic agents, and to the escape from immune surveillance. On these bases, we evaluated the role of different DDR pathways in MM and investigated, for the first time, the direct and immune-mediated anti-MM activity of the nucleotide excision repair (NER)-dependent agent trabectedin.
METHODS
Gene-expression profiling (GEP) was carried out with HTA2.0 Affymetrix array. Evaluation of apoptosis, cell cycle, and changes in cytokine production and release have been performed in 2D and 3D Matrigel-spheroid models through flow cytometry on MM cell lines and patients-derived primary MM cells exposed to increasing nanomolar concentrations of trabectedin. DNA-damage response has been evaluated through Western blot, immunofluorescence, and DNA fragmentation assay. Trabectedin-induced activation of NK has been assessed by CD107a degranulation. miRNAs quantification has been done through RT-PCR.
RESULTS
By comparing GEP meta-analysis of normal and MM plasma cells (PCs), we observed an enrichment in DNA NER genes in poor prognosis MM. Trabectedin triggered apoptosis in primary MM cells and MM cell lines in both 2D and 3D in vitro assays. Moreover, trabectedin induced DDR activation, cellular stress with ROS production, and cell cycle arrest. Additionally, a significant reduction of MCP1 cytokine and VEGF-A in U266-monocytes co-cultures was observed, confirming the impairment of MM-promoting milieu. Drug-induced cell stress in MM cells led to upregulation of NK activating receptors ligands (i.e., NKG2D), which translated into increased NK activation and degranulation. Mechanistically, this effect was linked to trabectedin-induced inhibition of NKG2D-ligands negative regulators IRF4 and IKZF1, as well as to miR-17 family downregulation in MM cells.
CONCLUSIONS
Taken together, our findings indicate a pleiotropic activity of NER-targeting agent trabectedin, which appears a promising candidate for novel anti-MM therapeutic strategies.
Topics: Antineoplastic Agents, Alkylating; Humans; Killer Cells, Natural; Multiple Myeloma; Trabectedin
PubMed: 30898137
DOI: 10.1186/s13045-019-0714-9 -
Scientific Reports Apr 2023DNA methylation is important for establishing and maintaining cell identity and for genomic stability. This is achieved by regulating the accessibility of regulatory and...
DNA methylation is important for establishing and maintaining cell identity and for genomic stability. This is achieved by regulating the accessibility of regulatory and transcriptional elements and the compaction of subtelomeric, centromeric, and other inactive genomic regions. Carcinogenesis is accompanied by a global loss in DNA methylation, which facilitates the transformation of cells. Cancer hypomethylation may also cause genomic instability, for example through interference with the protective function of telomeres and centromeres. However, understanding the role(s) of hypomethylation in tumor evolution is incomplete because the precise mutational consequences of global hypomethylation have thus far not been systematically assessed. Here we made genome-wide inventories of all possible genetic variation that accumulates in single cells upon the long-term global hypomethylation by CRISPR interference-mediated conditional knockdown of DNMT1. Depletion of DNMT1 resulted in a genomewide reduction in DNA methylation. The degree of DNA methylation loss was similar to that observed in many cancer types. Hypomethylated cells showed reduced proliferation rates, increased transcription of genes, reactivation of the inactive X-chromosome and abnormal nuclear morphologies. Prolonged hypomethylation was accompanied by increased chromosomal instability. However, there was no increase in mutational burden, enrichment for certain mutational signatures or accumulation of structural variation to the genome. In conclusion, the primary consequence of hypomethylation is genomic instability, which in cancer leads to increased tumor heterogeneity and thereby fuels cancer evolution.
Topics: Humans; DNA Methylation; Mutation; Genomic Instability; Carcinogenesis; DNA
PubMed: 37106015
DOI: 10.1038/s41598-023-33932-3 -
The Journal of Biological Chemistry Jul 2019-alkyl-2'-deoxyguanosine (-alkyl-dG) lesions are among the most mutagenic and prevalent alkylated DNA lesions that are associated with cancer initiation and progression....
-alkyl-2'-deoxyguanosine (-alkyl-dG) lesions are among the most mutagenic and prevalent alkylated DNA lesions that are associated with cancer initiation and progression. In this study, using a shuttle vector-based strand-specific PCR-competitive replication and adduct bypass assay in conjunction with tandem MS for product identification, we systematically assessed the repair and replicative bypass of a series of -alkyl-dG lesions, with the alkyl group being a Me, Et, Pr, Pr, Bu, Bu, or Bu, in several human cell lines. We found that the extent of replication-blocking effects of these lesions is influenced by the size of the alkyl groups situated on the position of the guanine base. We also noted involvement of distinct DNA repair pathways and translesion synthesis polymerases (Pols) in ameliorating the replication blockage effects elicited by the straight- and branched-chain -alkyl-dG lesions. We observed that -methylguanine DNA methyltransferase is effective in removing the smaller alkyl groups from the position of guanine, whereas repair of the branched-chain lesions relied on nucleotide excision repair. Moreover, these lesions were highly mutagenic during cellular replication and exclusively directed G→A mutations; Pol η and Pol ζ participated in error-prone bypass of the straight-chain lesions, whereas Pol κ preferentially incorporated the correct dCMP opposite the branched-chain lesions. Together, these results uncover key cellular proteins involved in repair and translesion synthesis of -alkyl-dG lesions and provide a better understanding of the roles of these types of lesions in the etiology of human cancer.
Topics: Alkylation; Cell Line; DNA Damage; DNA Repair; DNA Replication; Humans; Mutagenesis; O(6)-Methylguanine-DNA Methyltransferase
PubMed: 31167778
DOI: 10.1074/jbc.RA119.009054 -
Neuroendocrinology 2015Alkylating agents, such as streptozocin and dacarbazine, have been reported as active in neuroendocrine neoplasms (NENs). Temozolomide (TMZ) is an oral, potentially less... (Review)
Review
Alkylating agents, such as streptozocin and dacarbazine, have been reported as active in neuroendocrine neoplasms (NENs). Temozolomide (TMZ) is an oral, potentially less toxic derivative of dacarbazine, which has shown activity both as a single agent and in combination with other drugs. Nevertheless, its role in NENs has not been well defined. Several retrospective and prospective phase I-II studies have been published describing its use in a variety of NENs. In a retrospective series, the combination of capecitabine and TMZ was reported to be associated with a particularly high tumour response in pancreatic NENs as a first-line treatment. Although in NENs, determination of the O6-methylguanine-DNA methyltransferase (MGMT) status has been suggested as a predictive biomarker of response, its role still remains investigational, awaiting validation along with the establishment of the optimal detection method. Metronomic schedules have been reported to potentially overcome MGMT-related drug resistance. Toxicity is manageable if well monitored. We reviewed the literature regarding pharmacological and clinical aspects of TMZ, focusing on specific settings of NENs, different schedules, toxicity and safety profiles, and potential predictive biomarkers of response.
Topics: Antineoplastic Agents, Alkylating; Dacarbazine; Humans; Neuroendocrine Tumors; Temozolomide
PubMed: 25924937
DOI: 10.1159/000430816 -
Cells Jun 2023Glioblastoma is the most aggressive form of brain tumor originating from glial cells with a maximum life expectancy of 14.6 months. Despite the establishment of multiple... (Review)
Review
Glioblastoma is the most aggressive form of brain tumor originating from glial cells with a maximum life expectancy of 14.6 months. Despite the establishment of multiple promising therapies, the clinical outcome of glioblastoma patients is abysmal. Drug resistance has been identified as a major factor contributing to the failure of current multimodal therapy. Epigenetic modification, especially DNA methylation has been identified as a major regulatory mechanism behind glioblastoma progression. In addition, miRNAs, a class of non-coding RNA, have been found to play a role in the regulation as well as in the diagnosis of glioblastoma. The relationship between epigenetics, drug resistance, and glioblastoma progression has been clearly demonstrated. hypermethylation, leading to a lack of expression, is associated with a cytotoxic effect of TMZ in GBM, while resistance to TMZ frequently appears in non-methylated GBM. In this review, we will elaborate on known miRNAs linked to glioblastoma; their distinctive oncogenic or tumor suppressor roles; and how epigenetic modification of miRNAs, particularly via methylation, leads to their upregulation or downregulation in glioblastoma. Moreover, we will try to identify those miRNAs that might be potential regulators of expression and their role as predictors of tumor response to temozolomide treatment. Although we do not impact clinical data and survival, we open possible experimental approaches to treat GBM, although they should be further validated with clinically oriented studies.
Topics: Humans; Glioblastoma; Dacarbazine; Antineoplastic Agents, Alkylating; MicroRNAs; DNA Repair Enzymes; Tumor Suppressor Proteins; DNA Modification Methylases; Temozolomide; DNA Methylation; Epigenesis, Genetic
PubMed: 37371047
DOI: 10.3390/cells12121578