-
Nature Jun 2024DNA base damage is a major source of oncogenic mutations. Such damage can produce strand-phased mutation patterns and multiallelic variation through the process of...
DNA base damage is a major source of oncogenic mutations. Such damage can produce strand-phased mutation patterns and multiallelic variation through the process of lesion segregation. Here we exploited these properties to reveal how strand-asymmetric processes, such as replication and transcription, shape DNA damage and repair. Despite distinct mechanisms of leading and lagging strand replication, we observe identical fidelity and damage tolerance for both strands. For small alkylation adducts of DNA, our results support a model in which the same translesion polymerase is recruited on-the-fly to both replication strands, starkly contrasting the strand asymmetric tolerance of bulky UV-induced adducts. The accumulation of multiple distinct mutations at the site of persistent lesions provides the means to quantify the relative efficiency of repair processes genome wide and at single-base resolution. At multiple scales, we show DNA damage-induced mutations are largely shaped by the influence of DNA accessibility on repair efficiency, rather than gradients of DNA damage. Finally, we reveal specific genomic conditions that can actively drive oncogenic mutagenesis by corrupting the fidelity of nucleotide excision repair. These results provide insight into how strand-asymmetric mechanisms underlie the formation, tolerance and repair of DNA damage, thereby shaping cancer genome evolution.
Topics: DNA Damage; DNA Repair; Mutagenesis; DNA Replication; Mutation; Humans; Animals; DNA Adducts; Ultraviolet Rays; DNA; Alkylation; DNA-Directed DNA Polymerase
PubMed: 38867042
DOI: 10.1038/s41586-024-07490-1 -
Nature Communications Jun 2024Site-directed insertion is a powerful approach for generating mutant alleles, but low efficiency and the need for customisation for each target has limited its...
Site-directed insertion is a powerful approach for generating mutant alleles, but low efficiency and the need for customisation for each target has limited its application. To overcome this, we developed a highly efficient targeted insertional mutagenesis system, CRIMP, and an associated plasmid toolkit, CRIMPkit, that disrupts native gene expression by inducing complete transcriptional termination, generating null mutant alleles without inducing genetic compensation. The protocol results in a high frequency of integration events and can generate very early targeted insertions, during the first cell division, producing embryos with expression in one or both halves of the body plan. Fluorescent readout of integration events facilitates selection of successfully mutagenized fish and, subsequently, visual identification of heterozygous and mutant animals. Together, these advances greatly improve the efficacy of generating and studying mutant lines. The CRIMPkit contains 24 ready-to-use plasmid vectors to allow easy and complete mutagenesis of any gene in any reading frame without requiring custom sequences, modification, or subcloning.
Topics: Mutagenesis, Insertional; CRISPR-Cas Systems; Animals; Plasmids; Zebrafish; Genetic Vectors; Gene Editing; Alleles
PubMed: 38866742
DOI: 10.1038/s41467-024-49341-7 -
Scientific Reports Jun 2024Despite limited research on refractory and/or endocrine therapy failure in elderly metastatic breast cancer (MBC) patients, a prior study showed that low-dose oral...
Despite limited research on refractory and/or endocrine therapy failure in elderly metastatic breast cancer (MBC) patients, a prior study showed that low-dose oral cyclophosphamide (CY) can improve the overall survival rate of MBC patients, possibly through the immunoregulation of regulatory T cells (Tregs). We preliminarily investigated the combination of endocrine therapy (ET) with oral low-dose CY as salvage therapy in elderly patients via peripheral blood regulatory T-cell analyses. In addition, we evaluated the associations of tumor tertiary lymphoid structures (TLSs) with therapeutic outcomes. HR+/HER2- advanced breast cancer patients who received low-dose CY combined with ET or ET only from April 2015 to August 2021 were enrolled in this retrospective study. The primary outcome was the clinical control rate (CCR), and the secondary outcome was progression-free survival (PFS). Circulating T lymphocyte subpopulations represented by Tregs were monitored during treatment by flow cytometry methods. TLSs wereconfirmed by hematoxylin-eosin staining of pretreatment specimens, and CD3, CD4, and Foxp3 were detected using Opal multicolor immunofluorescence. A total of 85 patients who received CY + ET and 50 patients who received ET only were enrolled, the percentage of patients who received CCR was 73% (62/85) vs. 70% (45/50), and the objective response rate (ORR) was 28% (24/85) vs. 24% (12/50). No deaths occurred during the study period. The mean PFS time was 13 vs. 11 months (P = 0.03). In the CY + ET group, decreases in CD4+/CD25+/Foxp3+ T cells (P < 0.001) were favorable for both clinical control and prolonged PFS (P < 0.001). Compared with patients without TLSs, those with TLSs were more likely to have better clinical control and PFS (mean time = 6 months), and a greater number of Treg cells during TLS pretreatment correlated with longer PFS (P = 0.043). Oral low-dose CY combined with standard ET exerts immunological effects by decreasing Treg levels to achieve improved clinical responses. Moreover, patients with TLSs might benefit more from such therapy than those without TLSs, and a high Treg cell count in TLSs before treatment predicts better therapeutic efficacy.
Topics: Humans; Cyclophosphamide; T-Lymphocytes, Regulatory; Female; Breast Neoplasms; Aged; Retrospective Studies; Administration, Oral; Middle Aged; Aged, 80 and over; Antineoplastic Combined Chemotherapy Protocols; Neoplasm Metastasis; Treatment Outcome
PubMed: 38862586
DOI: 10.1038/s41598-024-64042-3 -
The Journal of Organic Chemistry Jun 2024A comprehensive quantum mechanical investigation delved into the antioxidative activity of galangin (). Thermochemical and kinetic data were used to assess antiradical,...
A comprehensive quantum mechanical investigation delved into the antioxidative activity of galangin (). Thermochemical and kinetic data were used to assess antiradical, chelating, and renewal potential under physiological conditions. A brief comparison with reference antioxidants and other flavonoids characterized as a moderate antioxidative agent. The substance showed significantly lower performance in lipid compared to aqueous solvent─the reaction rates for scavenging OOH in both media were established at 3.77 × 10 M s and 6.21 × 10 M s, respectively, accounting for the molar fraction of both interacting molecules at the given pH. The impact of pH value on the kinetics was assessed. Although efficient at chelating Cu(II) ions, the formed complexes can still undergo the Fenton reaction. On the other hand, they persistently scavenge OH . The flavonoid effectively repairs oxidatively damaged biomolecules except model lipid acids. All radicals are readily restored by physiologically prevailing O. Given this, the polyphenol is expected to participate in antiradical and regenerating activities multiple times, amplifying its antioxidative potential.
Topics: Flavonoids; Antioxidants; Density Functional Theory; Molecular Structure; Kinetics; Hydrogen-Ion Concentration
PubMed: 38861646
DOI: 10.1021/acs.joc.4c00611 -
Cell Transplantation 2024Posttransplant lymphoproliferative disorder (PTLD) is a rare lymphoid and/or plasmocytic proliferation that occurs after allogeneic hematopoietic stem cell...
Posttransplant lymphoproliferative disorder (PTLD) is a rare lymphoid and/or plasmocytic proliferation that occurs after allogeneic hematopoietic stem cell transplantation (allo-HSCT). We aimed to identify the pathologic features and clinical outcomes of T-cell PTLD, an extremely rare subtype of PTLD, after allo-HSCT. In this study, six allo-HSCT recipients with T-cell PTLD from five transplant centers in China were enrolled. All the T-cell PTLD were donor-derived, and three patients were with monomorphic and three with polymorphic types, respectively. All patients received cyclophosphamide, doxorubicin, vincristine, and prednisone (CHOP)-based chemotherapy. Five patients achieved complete response (CR), and one experienced progressive disease (PD). The median time from HSCT to onset was 4 (range: 0.6-72) months, analyzed in combination with the other 16 patients with T-cell PTLD identified from previous reports. About 56.3% of the T-cell samples (9/16) were positive for in situ hybridization with an Epstein-Barr virus (EBV)-encoded small nuclear early region (EBER ISH). CHOP-based chemotherapy might be the optimal strategy for patients who showed no response to empiric therapy with a CR rate of 87.5%. In conclusion, our study observed that T-cell PTLD has distinct clinical manifestations and morphological features, which characterized by less relation to EBV, later occurrence, and poorer prognosis when compared with B-cell PTLD.
Topics: Humans; Hematopoietic Stem Cell Transplantation; Lymphoproliferative Disorders; Male; Female; Adult; T-Lymphocytes; Transplantation, Homologous; Adolescent; Child; Middle Aged; Young Adult; Cyclophosphamide
PubMed: 38856035
DOI: 10.1177/09636897241259722 -
NAR Cancer Jun 2024Extrachromosomal circular DNAs (eccDNAs) are produced from all regions of the eucaryotic genome. We used inverse PCR of non-B microsatellites capable of forming hairpin,...
Extrachromosomal circular DNAs (eccDNAs) are produced from all regions of the eucaryotic genome. We used inverse PCR of non-B microsatellites capable of forming hairpin, triplex, quadruplex and AT-rich structures integrated at a common ectopic chromosomal site to show that these non-B DNAs generate highly mutagenized eccDNAs by replication-dependent mechanisms. Mutagenesis occurs within the non-B DNAs and extends several kilobases bidirectionally into flanking and nonallelic DNA. Each non-B DNA exhibits a different pattern of mutagenesis, while sister clones containing the same non-B DNA also display distinct patterns of recombination, microhomology-mediated template switching and base substitutions. Mutations include mismatches, short duplications, long nontemplated insertions, large deletions and template switches to sister chromatids and nonallelic chromosomes. Drug-induced replication stress or the depletion of DNA repair factors Rad51, the COPS2 signalosome subunit or POLη change the pattern of template switching and alter the eccDNA mutagenic profiles. We propose an asynchronous capture model based on break-induced replication from microsatellite-induced DNA double strand breaks to account for the generation and circularization of mutagenized eccDNAs and the appearance of genomic homologous recombination deficiency (HRD) scars. These results may help to explain the appearance of tumor eccDNAS and their roles in neoantigen production, oncogenesis and resistance to chemotherapy.
PubMed: 38854437
DOI: 10.1093/narcan/zcae027 -
BioRxiv : the Preprint Server For... May 2024In alginate biosynthesis gene expression is inhibited by the transmembrane anti-sigma factor MucA, which sequesters the AlgU sigma factor. Cell envelope stress...
UNLABELLED
In alginate biosynthesis gene expression is inhibited by the transmembrane anti-sigma factor MucA, which sequesters the AlgU sigma factor. Cell envelope stress initiates cleavage of the MucA periplasmic domain by site-1 protease AlgW, followed by further MucA degradation to release AlgU. However, after colonizing the lungs of people with cystic fibrosis, converts to a mucoid form that produces alginate constitutively. Mucoid isolates often have mutations, with the most common being , which truncates the periplasmic domain. MucA22 is degraded constitutively, and genetic studies suggested that the Prc protease is responsible. Some studies also suggested that Prc contributes to induction in strains with wild type MucA, whereas others suggested the opposite. However, missing from all previous studies is a demonstration that Prc cleaves any protein directly, which leaves open the possibility that the effect of a null mutation is indirect. To address the ambiguities and shortfalls, we reevaluated the roles of AlgW and Prc as MucA and MucA22 site-1 proteases. analyses using three different assays, and two different inducing conditions, all suggested that AlgW is the only site-1 protease for wild type MucA in any condition. In contrast, genetics suggested that AlgW or Prc act as MucA22 site-1 proteases in inducing conditions, whereas Prc is the only MucA22 site-1 protease in non-inducing conditions. For the first time, we also show that Prc is unable to degrade the periplasmic domain of wild type MucA, but does degrade the mutated periplasmic domain of MucA22 directly.
IMPORTANCE
After colonizing the lungs of individuals with cystic fibrosis, undergoes mutagenic conversion to a mucoid form, worsening the prognosis. Most mucoid isolates have a truncated negative regulatory protein MucA, which leads to constitutive production of the extracellular polysaccharide alginate. The protease Prc has been implicated, but not shown, to degrade the most common MucA variant, MucA22, to trigger alginate production. This work provides the first demonstration that the molecular mechanism of Prc involvement is direct degradation of the MucA22 periplasmic domain, and perhaps other truncated MucA variants as well. MucA truncation and degradation by Prc might be the predominant mechanism of mucoid conversion in cystic fibrosis infections, suggesting that Prc activity could be a useful therapeutic target.
PubMed: 38854061
DOI: 10.1101/2024.05.28.596254 -
Chemosphere Aug 2024This study aims the characterization of several tianeptine transformation products in ultrapure water by simulated sunlight irradiation. Tianeptine was completely...
This study aims the characterization of several tianeptine transformation products in ultrapure water by simulated sunlight irradiation. Tianeptine was completely degraded after 106 h of exposition following pseudo-first-order kinetics (half-life time = 12.0 ± 2.4 h). Furthermore, an ultra-high-performance liquid chromatography coupled with a high-resolution quadrupole time-of-flight-mass spectrometry method was developed and fully validated taking into account different method performance parameters for the quantification of tianeptine in river water up to a concentration of 400 pg L. Following a non-targeted approach based on mass data-independent acquisition, eight different transformation products not previously reported in the literature were identified and accordingly elucidated, proposing a photodegradation mechanism based on the accurate tandem mass spectrometry information acquired. Irradiation experiments were replicated for a tianeptine solution prepared in a blank river water sample, resulting in the formation of the same transformation products and similar degradation kinetics. In addition, a toxicity assessment of the photoproducts was performed by in silico method, being generally all TPs of comparable toxicity to the precursor except for TP1, and showing a similar persistence in the environment except for TP2 and TP6, while TP4 was the only TP predicted as mutagenic. The developed method was applied for the analysis of four river water samples.
Topics: Water Pollutants, Chemical; Tandem Mass Spectrometry; Chromatography, High Pressure Liquid; Photolysis; Thiazepines; Rivers; Kinetics; Sunlight
PubMed: 38849097
DOI: 10.1016/j.chemosphere.2024.142534 -
Environmental Pollution (Barking, Essex... Jun 2024Chemical pollution is one of the most important threats to freshwater ecosystems. The plethora of potentially occurring chemicals and their effects in complex mixtures...
Chemical pollution is one of the most important threats to freshwater ecosystems. The plethora of potentially occurring chemicals and their effects in complex mixtures challenge standard monitoring methods. Effect-based methods (EBMs) are proposed as complementary tools for the assessment of chemical pollution and toxic effects. To investigate the effects of chemical pollution, the ecological relevance of EBMs and the potential of macroinvertebrates as toxicity-specific bioindicators, ecological and ecotoxicological data were linked. Baseline toxicity, mutagenicity, dioxin-like and estrogenic activity of water and sediment samples from 30 river sites in central Germany were quantified with four in vitro bioassays. The responses of macroinvertebrate communities at these sites were assessed by calculating 16 taxonomic and functional metrics and by investigating changes in the taxonomic and trait composition. Principal component analysis revealed an increase in toxicity along a joint gradient of chemicals with different modes of action. This toxicity gradient was associated with a decrease in biodiversity and ecological quality, as well as significant changes in taxonomic and functional composition. The strength of the effects suggested a strong impact of chemical pollution and underlined the suitability of EBMs in detecting ecological relevant effects. However, the metrics, taxa, and traits associated with vulnerability or tolerance to toxicity were found to also respond to other stressors in previous studies and thus may have only a low potential as toxicity-specific bioindicators. Because macroinvertebrates respond integratively to all present stressors, linking both ecological and environmental monitoring is necessary to investigate the overall effects but also isolate individual stressors. EBMs have a high potential to separate the toxicity of chemical mixtures from other stressors in a multiple stressor scenario, as well as identifying the presence of chemical groups with specific modes of action.
PubMed: 38848961
DOI: 10.1016/j.envpol.2024.124330 -
Medicine Jun 2024Intestinal T-cell lymphomas are exceedingly rare diseases. Intestinal T-cell lymphoma NOS, as a "wastebasket" category, is difficult to diagnosis. Endoscopy can identify...
RATIONALE
Intestinal T-cell lymphomas are exceedingly rare diseases. Intestinal T-cell lymphoma NOS, as a "wastebasket" category, is difficult to diagnosis. Endoscopy can identify abnormal mucosa in most patients at a reasonably early stage. Therefore, it is crucial to increase the understanding of endoscopists in terms of the endoscopic characteristics of ITCL.
PATIENT CONCERNS
A 74-year-old male alone with wasting as the major complaint, had multiple polypoid lesions in the large intestine. The patient then had endoscopic care.
DIAGNOSES
Only 1 polypoid lesion on white-light endoscopy in the sigmoid colon was pathologically diagnosed as intestinal T-cell lymphomas, not otherwise specified (ITCL-NOS).
INTERVENTIONS
The patient underwent intensity-reduced CHOP therapy.
OUTCOMES
The patient is still with controlled disease but developed chemotherapy-related side effects.
LESSONS
In the individual with unexplained anemia and waste, endoscopy should not be delayed. For each of polypoid lesion on white-light endoscopy, the endoscopist need to remain cautious, because every lesion in the same patient can exhibit the independence of histopathological features. Meanwhile, we suggest that endoscopists should routinely observe the terminal ileum, even take biopsy samples if necessary.
Topics: Humans; Aged; Male; Antineoplastic Combined Chemotherapy Protocols; Lymphoma, T-Cell; Doxorubicin; Vincristine; Intestinal Neoplasms; Cyclophosphamide; Prednisone; Colonoscopy
PubMed: 38847694
DOI: 10.1097/MD.0000000000038465