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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 -
Current Protocols in Nucleic Acid... Jun 2019Immobilization of DNA is an important step in relation to DNA-based biosensors and bioassays with multiple applications. This unit describes synthesis and applications...
Immobilization of DNA is an important step in relation to DNA-based biosensors and bioassays with multiple applications. This unit describes synthesis and applications of novel bifunctional linker molecules containing nitrogen mustard and one of two types of functional groups: cyclic disulfide or biotin. Two ways of immobilizing DNA on a surface are described. With the first method, a bifunctional alkylating linker molecule is first reacted with the target DNA to form alkylated DNA and then immobilized on a specific surface. With the second method, the bifunctional alkylating linker molecule is first attached to the surface, and then the target DNA is immobilized through an alkylating reaction with a nitrogen mustard moiety. We have also achieved immunochemical detection and quantification of 5-methylcytosine in a target DNA immobilized by the above methods. The methods for immobilization of intact DNA using novel bifunctional linker molecules are applicable to a wide range of biological analysis techniques. © 2019 by John Wiley & Sons, Inc.
Topics: Alkylation; Animals; Biosensing Techniques; Biotin; DNA; Denaturing Gradient Gel Electrophoresis; Disulfides; Gold; Mechlorethamine; Mice
PubMed: 31038292
DOI: 10.1002/cpnc.85 -
Nucleosides, Nucleotides & Nucleic Acids 2021We report herein comprehensive investigations of alkylation/sulfur exchange reactions of sulfur-containing substrates including nucleosides such as sU, msU, sU, sA and...
We report herein comprehensive investigations of alkylation/sulfur exchange reactions of sulfur-containing substrates including nucleosides such as sU, msU, sU, sA and sT-incorporated DNA enable by comprehensive screenings of the reagents (). It has been proven that iodoacetamide () displays the most promising feasibility toward sulfur-containing substrates including sT, sU, msU, sU and sA. In sharp contrast, the alkylation process with S-benzyl methanethiosulfonate (BMTS, ) displays the best application potential only for sU. Based on these results, the fluorescent labeling of sT-incorporated DNA and msU-modified RNA has been achieved.Supplemental data for this article is available online at https://doi.org/10.1080/15257770.2021.1942044 .
Topics: Alkylation; DNA, Bacterial; RNA
PubMed: 34180339
DOI: 10.1080/15257770.2021.1942044 -
Chemical Research in Toxicology Dec 2019Nitrogen mustards have long been used in cancer chemotherapy, and their cytotoxicity has traditionally been attributed to the formation of DNA interstrand cross-links...
Nitrogen mustards have long been used in cancer chemotherapy, and their cytotoxicity has traditionally been attributed to the formation of DNA interstrand cross-links and DNA monoalkylation. Recent studies have shown that exposure to nitrogen mustards also induces the formation of DNA-protein cross-links (DPCs) via bridging between N7 of a deoxyguanosine residue in the DNA and the side chain of a Cys residue in the protein. However, the formation of nitrogen mustard-induced DNA-histone cross-links has never been observed. Herein, we demonstrate that treating reconstituted nucleosome core particles (NCPs) with the nitrogen mustard mechlorethamine results in the formation of DNA-histone cross-links in addition to DNA monoalkylation and interstrand cross-link formation. The yields of these three types of DNA lesions in the NCPs decreased in the following order: DNA monoalkylation ≫ DNA interstrand cross-links > DNA-histone cross-links. Mechanistic studies involving tailless histones and competitive inhibition by a polyamine demonstrated that Lys residues in the N- and C-terminal tails of the histones were the predominant sites involved in DNA-histone cross-link formation. Given that NCPs are the fundamental repeating units of chromatin in eukaryotes, our findings suggest that nitrogen mustard-induced formation of DNA-histone cross-links may occur in living cells and that DPC formation may contribute to the cytotoxicity of nitrogen mustards.
Topics: Alkylating Agents; Amino Acid Sequence; Animals; Cross-Linking Reagents; DNA; Histones; Male; Mechlorethamine; Models, Chemical; Nucleosomes; Salmon; Spermatozoa
PubMed: 31726825
DOI: 10.1021/acs.chemrestox.9b00354 -
Future Oncology (London, England) Oct 2017Trabectedin presents a complex mode of action affecting key cell biology processes in tumor cells and in the tumor microenvironment. In ovarian cancer patients with a... (Review)
Review
Trabectedin presents a complex mode of action affecting key cell biology processes in tumor cells and in the tumor microenvironment. In ovarian cancer patients with a platinum treatment-free interval of 6-12 months treated with trabectedin + pegylated liposomal doxorubicin (PLD) or single-agent PLD, and retreated with platinum after relapse, overall survival was significantly prolonged in the trabectedin + PLD group. Mechanisms by which trabectedin restores tumor sensitivity to platinum include its interaction with components of the nucleotide excision repair machinery in tumor cells and inhibition of inflammatory mediators such as IL-6 in the tumor microenvironment. Additionally, BRCA mutations and associated homologous recombination repair deficiency may contribute to enhanced sensitivity to trabectedin observed in BRCA-mutated patients with ovarian cancer.
Topics: Animals; Antineoplastic Agents, Alkylating; Biomarkers, Tumor; DNA; Dioxoles; Drug Resistance, Neoplasm; Genes, BRCA1; Genes, BRCA2; Humans; Mutation; Neoplasms; Platinum; Prognosis; Tetrahydroisoquinolines; Trabectedin; Treatment Outcome
PubMed: 29020822
DOI: 10.2217/fon-2017-0318 -
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 -
European Journal of Medicinal Chemistry May 2018Cancer is considered as one of the most serious health problems today. The discovery of nitrogen mustard as an alkylating agent in 1942, opened a new era in the cancer... (Review)
Review
Cancer is considered as one of the most serious health problems today. The discovery of nitrogen mustard as an alkylating agent in 1942, opened a new era in the cancer chemotherapy. This valuable class of alkylating agent exerts its biological activity by binding to DNA, cross linking two strands, preventing DNA replication and ultimate cell death. At the molecular level, nitrogen lone pairs of nitrogen mustard generate a strained intermediate "aziridinium ion" which is very reactive towards DNA of tumor cell as well as normal cell resulting in various adverse side effects alogwith therapeutic implications. Over the last 75 years, due to its high reactivity and peripheral cytotoxicity, numerous modifications have been made in the area of nitrogen mustard to improve its efficacy as well as enhancing drug delivery specifically to tumor cells. This review mainly discusses the medicinal chemistry aspects in the development of various classes of nitrogen mustards (mechlorethamine, chlorambucil, melphalan, cyclophosphamide and steroidal based nitrogen mustards). The literature collection includes the historical and the latest developments in these areas. This comprehensive review also attempted to showcase the recent progress in the targeted delivery of nitrogen mustards that includes DNA directed nitrogen mustards, antibody directed enzyme prodrug therapy (ADEPT), gene directed enzyme prodrug therapy (GDEPT), nitrogen mustard activated by glutathione transferase, peptide based nitrogen mustards and CNS targeted nitrogen mustards.
Topics: Animals; Antineoplastic Agents, Alkylating; Drug Delivery Systems; Humans; Neoplasms; Nitrogen Mustard Compounds
PubMed: 29649739
DOI: 10.1016/j.ejmech.2018.04.001 -
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 -
MBio Apr 2022Unique DNA repair enzymes that provide self-resistance against therapeutically important, genotoxic natural products have been discovered in bacterial biosynthetic gene...
Unique DNA repair enzymes that provide self-resistance against therapeutically important, genotoxic natural products have been discovered in bacterial biosynthetic gene clusters (BGCs). Among these, the DNA glycosylase AlkZ is essential for azinomycin B production and belongs to the HTH_42 superfamily of uncharacterized proteins. Despite their widespread existence in antibiotic producers and pathogens, the roles of these proteins in production of other natural products are unknown. Here, we determine the evolutionary relationship and genomic distribution of all HTH_42 proteins from and use a resistance-based genome mining approach to identify homologs associated with known and uncharacterized BGCs. We find that AlkZ-like (AZL) proteins constitute one distinct HTH_42 subfamily and are highly enriched in BGCs and variable in sequence, suggesting each has evolved to protect against a specific secondary metabolite. As a validation of the approach, we show that the AZL protein, HedH4, associated with biosynthesis of the alkylating agent hedamycin, excises hedamycin-DNA adducts with exquisite specificity and provides resistance to the natural product in cells. We also identify a second, phylogenetically and functionally distinct subfamily whose proteins are never associated with BGCs, are highly conserved with respect to sequence and genomic neighborhood, and repair DNA lesions not associated with a particular natural product. This work delineates two related families of DNA repair enzymes-one specific for complex alkyl-DNA lesions and involved in self-resistance to antimicrobials and the other likely involved in protection against an array of genotoxins-and provides a framework for targeted discovery of new genotoxic compounds with therapeutic potential. Bacteria are rich sources of secondary metabolites that include DNA-damaging genotoxins with antitumor/antibiotic properties. Although produce a diverse number of therapeutic genotoxins, efforts toward targeted discovery of biosynthetic gene clusters (BGCs) producing DNA-damaging agents is lacking. Moreover, work on toxin-resistance genes has lagged behind our understanding of those involved in natural product synthesis. Here, we identified over 70 uncharacterized BGCs producing potentially novel genotoxins through resistance-based genome mining using the azinomycin B-resistance DNA glycosylase AlkZ. We validate our analysis by characterizing the enzymatic activity and cellular resistance of one AlkZ ortholog in the BGC of hedamycin, a potent DNA alkylating agent. Moreover, we uncover a second, phylogenetically distinct family of proteins related to Escherichia coli YcaQ, a DNA glycosylase capable of unhooking interstrand DNA cross-links, which differs from the AlkZ-like family in sequence, genomic location, proximity to BGCs, and substrate specificity. This work defines two families of DNA glycosylase for specialized repair of complex genotoxic natural products and generalized repair of a broad range of alkyl-DNA adducts and provides a framework for targeted discovery of new compounds with therapeutic potential.
Topics: Alkylating Agents; Anti-Bacterial Agents; Biological Products; DNA; DNA Adducts; DNA Glycosylases; Mutagens; Streptomyces
PubMed: 35311535
DOI: 10.1128/mbio.03297-21 -
Genes Jun 2023The inhibition of histone deacetylases (HDACs) holds promise as a potential anti-cancer therapy as histone and non-histone protein acetylation is frequently disrupted in...
The inhibition of histone deacetylases (HDACs) holds promise as a potential anti-cancer therapy as histone and non-histone protein acetylation is frequently disrupted in cancer, leading to cancer initiation and progression. Additionally, the use of a histone deacetylase inhibitor (HDACi) such as the class I HDAC inhibitor-valproic acid (VPA) has been shown to enhance the effectiveness of DNA-damaging factors, such as cisplatin or radiation. In this study, we found that the use of VPA in combination with talazoparib (BMN-673-PARP1 inhibitor-PARPi) and/or Dacarbazine (DTIC-alkylating agent) resulted in an increased rate of DNA double strand breaks (DSBs) and reduced survival (while not affecting primary melanocytes) and the proliferation of melanoma cells. Furthermore, the pharmacological inhibition of class I HDACs sensitizes melanoma cells to apoptosis following exposure to DTIC and BMN-673. In addition, the inhibition of HDACs causes the sensitization of melanoma cells to DTIV and BMN-673 in melanoma xenografts in vivo. At the mRNA and protein level, the histone deacetylase inhibitor downregulated RAD51 and FANCD2. This study aims to demonstrate that combining an HDACi, alkylating agent and PARPi could potentially enhance the treatment of melanoma, which is commonly recognized as being among the most aggressive malignant tumors. The findings presented here point to a scenario in which HDACs, via enhancing the HR-dependent repair of DSBs created during the processing of DNA lesions, are essential nodes in the resistance of malignant melanoma cells to methylating agent-based therapies.
Topics: Humans; Histone Deacetylase Inhibitors; Valproic Acid; Poly(ADP-ribose) Polymerase Inhibitors; Dacarbazine; Histone Deacetylases; Antineoplastic Agents; Melanoma; DNA; Alkylating Agents
PubMed: 37372475
DOI: 10.3390/genes14061295