Nucleosides, Nucleotides & Nucleic Acids最新文献

Oral lichen planus (OLP) is a chronic inflammatory condition that has been clinically linked with the risk of developing oral cancer. The present study aimed to determine the oxidative stress in oral lichen planus (OLP) by assessing the immunohistochemical (IHC) expression of 8-hydroxydeoxyguanosine (8-OHdG) in OLP tissue samples and comparing it with that of normal oral mucosa. The study group consisted of 30 formalin-fixed, paraffin-embedded (FFPE) tissue blocks from oral lichen planus (OLP) cases, while 10 normal oral mucosa samples served as the control group. Tissue sections of 5-micron thickness were prepared and immunostained with the 8-hydroxydeoxyguanosine (8-OHdG) antibody. The stained slides were examined under a light microscope, and statistical analysis was conducted using Fisher's exact test. A significant increase in oxidative DNA damage marker 8-OHdG expression (p < 0.05) was observed within the basal and suprabasal epithelial layers of lichen planus tissue samples. In contrast, no immunoreactivity was detected in the normal oral mucosa. Excessive production of reactive oxygen species (ROS) during chronic inflammation is believed to play a crucial role in inducing DNA damage. The mutagenic marker 8-hydroxydeoxyguanosine (8-OHdG) in oral lichen planus tissue highlights its potential as a biomarker for assessing the risk of inflammation-driven carcinogenesis. Further research, incorporating various oxidative stress markers and clinicopathological correlations, is essential to enhance early prediction of malignant transformation in oral lichen planus.

Extracellular ATP (eATP) is a major component of the tumor microenvironment which has been shown to play an important role in inflammation and cancer. Previously, we have shown that eATP, through P2Y12 receptors, increased the levels of the pro-inflammatory enzyme cyclooxygenase 2 (COX-2) in tumor cells, which in turn imparted metastatic property to cancer cells. In a mouse model of lymphoma, we further showed that both the P2Y12 receptor-specific antagonist, AR-C 69931, and the P2Y6 receptor-specific antagonist, MRS 2578, significantly arrested tumor progression. In the case of tumor-bearing mice treated with AR-C 69931, a strong reduction in COX-2 expression was observed which in turn reduced metastasis. However, COX-2 expression was not altered in mice treated with MRS 2578. In this paper, we report that inhibition of the P2Y6 receptor acts through modulation of cell cycle proteins leading to cell cycle arrest. Our results show that blocking of P2Y6 receptors is a therapeutic alternative to arrest tumor growth and metastasis.

The sodium salts of 2-formyl-1-cycloalkanones and α-(hydroxymethylene)alkanones, when reacted with cyanothioacetamide, produced equivalent cycloalkane ring fused pyridine-2(1H)-thiones and substituted pyridone-2(1H)-thiones. The corresponding pyridine S-glycosides were obtained in good yields from the later compounds by a successful coupling reaction with protected bromoarabinose or bromoxylose in KOH-acetone at room temperature and/or HDMS/(NH₄)₂SO₄. After ten minutes of room temperature reaction between the later compounds and NH₃-MeOH, excellent yields of the deprotected derivatives were produced. Using spectral data and fundamental analyses, the structure of the newly synthesized compounds were proved (IR, ¹H NMR, and ¹³C NMR). All prepared thioglycoside substances had their antibacterial properties assessed and screened. Some thioglycoside derivatives turned out to be potent antibacterial agents.

A series of conjugates of diterpenoid isosteviol (16-oxo-ent-beyran-19-oic acid) and uracil (nucleoterpenoids) was synthesized and examined for their in vitro cytotoxicity against 9 human cancer cell lines. Nucleoterpenoids 13c,f,15 exhibited the best in vitro cytotoxic activity against cancer cell lines M-HeLa, MCF-7, and PC3 (IC₅₀ = 12.7-21.3 µM) among the synthesized compounds. The mechanisms of the in vitro cytotoxic effect of nucleoterpenoids 13f and 15 against M-HeLa cancer cell line (cervical carcinoma) were studied using flow cytofluorometry and enzyme-linked immunosorbent assay (ELISA). The results obtained indicated that 13f and 15 by acting on M-HeLa cancer cells reduced the mitochondrial membrane potential, caused oxidative stress, blocked anti-apoptotic protein Bcl-2, activated apoptosis-initiating caspase-9, thus inducing apoptosis occurring along the mitochondrial pathway. It should be emphasized that although isosteviol and uracil do not have cytotoxicity against human cancer cells, nucleoterpenoids 13c,f, and 15 containing isosteviol and uracil as fragments exhibited good cytotoxicity against M-HeLa, MCF-7, and PC3 cancer cells. Thus, it can be considered that conjugation of diterpenoid isosteviol and nucleic bases is a promising way to search for new cytotoxic agents of unusual structure.

The stability of CRISPR-Cas9 endonuclease activity is essential for its effectiveness in molecular diagnostics and gene editing. Target sequences of Human Papillomavirus (HPV) types 16 and 18 were selected by generating a consensus sequence following multiple sequence alignment, to ensure high specificity. The single guide RNAs (sgRNAs) were designed by identifying Protospacer Adjacent Motif (PAM) sequences within the E6 gene of HPV-16 and HPV-18. High-fidelity DNA oligonucleotides were synthesized from Integrated DNA Technologies (IDT) and transcribed in vitro into single guide RNAs (sgRNAs). These sgRNAs were then assembled with Cas9 protein to form CRISPR-Cas9 ribonucleoprotein (RNP) complexes, which were subsequently lyophilized to enhance storage stability. Functional validation of the RNP complexes was performed over a period of up to 18 months using polymerase chain reaction (PCR), agarose gel electrophoresis (AGE), and SYBR Green-based real-time PCR (RT-PCR) to confirm endonuclease activity and cleavage efficiency. This study assessed the activity of lyophilized HPV-16 and HPV-18 CRISPR-Cas9-RNPs stored at 4 °C for up to 18 months. Results demonstrated that the ribonucleoprotein (RNP) complex consisting sgRNA retained significant endonuclease activity, supporting lyophilization as a viable strategy for enhancing the stability and shelf life of CRISPR-Cas9 complexes for long-term applications.

N⁶-methyladenosine (m6A) is the most plentiful internal modification event in eukaryotic RNAs and regulated dynamically by a variety of its cognate functional partners in a context-dependent manner, namely writers, erasers and readers. The YTHDF1 is a well-documented cytoplasmic m6A reader that can specifically recognize and interact with m6A via its YT521-B homology (YTH) domain. In this study, the three methyl hydrogens of m6A mononucleotide were systematically swapped with four halogens (X = F, Cl, Br and I), totally resulting in the 4 single-, 10 double- and 20 triple-halogen modified counterparts of m6A (i.e. N⁶-halomethyladenosine, Xm6A). Energetic analyses and affinity assays indicated a moderate binding potency of m6A mononucleotide to YTHDF1 YTH domain, which can be improved by different single-halogen modifications. The electron-correlation calculations revealed that the affinity improvement is primarily donated from formation of additional halogen-π interactions with YTH aromatic cage upon the halogenations. Bromine, but not the bulky iodine, modification reaches the affinity maximum at Brm6A - this can be considered as a compromise between the favorable halogen-π interactions and the unfavorable steric effects caused by halogen modifications. For the same reason, single- and double-modifications generally contribute more favorably to YTH-Xm6A binding than triple-modifications. Further competition assays observed that the m6A mononucleotide and its single-modified Brm6A counterpart can competitively displace a m6A-containing RNA bound in the binding site of YTHDF1 YTH domain in moderate and high potencies, respectively, implying that halogen-modified Xm6A mononucleotides could be exploited as therapeutic strategy by competitively disrupting the cognate intermolecular interaction between YTH domain and RNA m6A.

The role of oleuropein (OLE) in K562 leukemia progression was investigated by evaluating its antiproliferative effect via the XTT assay, oxidative damage through 8-OHdG and TAS/TOS measurements, and apoptotic activity using Caspase-3 assays. Total RNA was isolated from control and treated cells, followed by cDNA synthesis and RT-PCR analysis to assess mRNA expression of DNA repair-related genes. OLE reduced K562 leukemia cell viability in a time- and dose-dependent manner, with an IC₅₀ of 244 µM at 72 h. Treatment with OLE decreased both total oxidant and antioxidant status, while increasing the oxidative stress index. However, changes in 8-OHdG and Caspase-3 levels were not statistically significant. Additionally, no significant differences were observed in the expression of XRCC1, APE1, NEIL1, and PARP1 genes after 72 h of treatment with 244 µM OLE. These findings suggest that OLE may possess anticancer potential and could contribute to the development of therapeutic strategies for hematological malignancies such as leukemia.

Hepatitis C Virus (HCV) is highly a mutable RNA virus, primarily due to lack of proofreading activity in RNA-dependent RNA polymerase. This intrinsic feature leads to the generation of substantial intra-host variation, resulting in the formation of viral quasispecies. The 5' untranslated region (5'UTR) of the HCV genome is highly conserved across different genotypes and serves as a widely adopted target for genotyping in clinical practice. In this study, the performance of targeted short-read next-generation sequencing of the HCV 5' UTR using Ion Torrent technology was evaluated, with the dual aim of assigning genotypes and detecting low-frequency intra-host variants. The 5'UTR-based nested RT-PCR products were amplified for five clinical plasma samples and sequenced on the Ion Gene Studio S5 platform. The resulting reads were assembled to generate consensus sequences and subjected to comprehensive analysis for iSNVs, SNPs, and indels. Sequencing of the amplicons yielded short-read sequences of 200-300 nucleotides. Phylogenetic analyses using MEGA and Interactive Tree of Life demonstrated that all isolates clustered within genotype 3, with sub-clustering indicative of subtype 3a and other intra-genotypic variants. Fourteen SNPs and several low-frequency iSNVs most notably at positions 72, 78, and 128, with variant frequencies reaching up to ˜25% were detected. These findings underscore the method's ability to discriminate both genotype and sub-variant structure. Overall, this strategy enhances molecular epidemiology, investigations, enables robust transmission tracking, and supports individualized therapeutic decision-making, particularly in resource-limited diagnostic settings.

Background: Triple-negative breast cancer (TNBC) is an aggressive subtype characterized by a poor prognosis. MicroRNAs (miRNAs) play a crucial regulatory role in tumorigenesis, but the specific function and mechanism of miR-1266-5p in TNBC remain unclear.

Methods: This study included 118 TNBC patients, from whom both tumor and adjacent normal tissues were collected. The expression of miR-1266-5p was determined, and its association with clinicopathological features and patient prognosis was evaluated. Functional experiments were conducted using TNBC cell lines (MDA-MB-231, MDA-MB-468), assessing the impact of miR-1266-5p on cellular processes. Bioinformatics tools and dual-luciferase reporter assays were employed to identify and validate the target gene of miR-1266-5p.

Results: miR-1266-5p was upregulated in TNBC tissues and cells (p < 0.05), and its high expression was associated with lymph node metastasis, higher histological grade, and advanced TNM stage. Kaplan-Meier analysis revealed that patients with high miR-1266-5p expression had shorter overall survival, and it was an independent prognostic factor. Functional experiments demonstrated that overexpression of miR-1266-5p significantly promoted TNBC cell proliferation, migration, and invasion (p < 0.05), while knockdown suppressed these phenotypes. Bioinformatics analysis and dual-luciferase assays identified MKRN1 as a direct target of miR-1266-5p.

Conclusion: miR-1266-5p promotes tumor progression in TNBC by targeting MKRN1. Its high expression correlates with poor patient outcomes, suggesting that it may serve as a promising biomarker for prognosis and a potential therapeutic target in TNBC.