Evaluation of DFT methods for predicting geometries and NMR spectra of Bi(III) dithiocarbamate complexes with antitumor properties.

CONTEXT: Bismuth complexes with dithiocarbamate ligands have attracted attention because of their biological applications, such as antimicrobial, antileishmanial, and anticancer properties. These complexes have high cytotoxic activity against cancer cells, being more active than the standard drugs cisplatin, doxorubicin, and tamoxifen. In the present study, we investigated the ability of some DFT methods to reproduce the geometries and NMR spectra of the Bi(III) dithiocarbamate complexes, selected based on their proven antitumor activity. Our investigation revealed that the M06-L/def2-TZVP/ECP/CPCM method presented good accuracy in predicting geometries, while the TPSSh/def2-SVP/ECP/CPCM method proved effective in analyzing the 13C NMR spectra of these molecules. In general, all examined methods exhibited comparable performance in predicting 1H NMR signals. METHODS: Calculations were performed with the Gaussian 09 program using the def2-SVP and def2-TZVP basis sets, employing relativistic effective core potential (ECP) for Bi and using the CPCM solvent model. The exchange-correlation functionals BP86, PBE, OLYP, M06-L, B3LYP, B3LYP-D3, M06-2X, TPSSh, CAM-B3LYP, and ωB97XD were used in the study. Geometry optimizations were started from crystallographic structures available at the Cambridge Structural Database. The theoretical results were compared with experimental data using the mean root-mean-square deviation (RMSD), mean absolute deviations (MAD), and linear correlation coefficient (R2).

Anticancer Potential of Indole Phytoalexins and Their Analogues.

Indole phytoalexins, found in economically significant Cruciferae family plants, are synthesized in response to pathogen attacks or stress, serving as crucial components of plant defense mechanisms against bacterial and fungal infections. Furthermore, recent research indicates that these compounds hold promise for improving human health, particularly in terms of potential anticancer effects that have been observed in various studies. Since our last comprehensive overview in 2016 focusing on the antiproliferative effects of these substances, brassinin and camalexin have been the most extensively studied. This review analyses the multifaceted pharmacological effects of brassinin and camalexin, highlighting their anticancer potential. In this article, we also provide an overview of the antiproliferative activity of new synthetic analogs of indole phytoalexins, which were synthesized and tested at our university with the aim of enhancing efficacy compared to the parent compound.

Investigations of Ni(II)Cysteine-Tyrosine Dithiocarbamate Complex: Synthesis, Characterization, Molecular Docking, Molecular Dynamic, and Anticancer Activity on MCF-7 Breast Cancer Cell Line.

OBJECTIVE: Breast cancer ranks second in terms of the highest number of cancer deaths for women worldwide and is one of the leading causes of death from cancer in women. The drug that is often used for chemotherapy is cisplatin. However, cisplatin drugs have a number of problems, including lack of selectivity, unwanted side effects, resistance, and toxicity in the body. In this work, we investigated Ni(II) cysteine-tyrosine dithiocarbamate complex against breast cancer. METHODS: Research on the new complex compound Ni(II) cysteine-tyrosine dithiocarbamate have several stages including synthesis, characterization, in-silico and in-vitro testing of MCF-7 cells for anticancer drugs. The synthesis involved reacting cysteine, CS2, KOH and tyrosine with Mn metal. The new complex compound Ni(II) cysteine-tyrosine dithiocarbamate has been synthesized, characterized, and tested in vitro MCF-7 cells for anticancer drugs. Characterization tests such as melting point, conductivity, SEM-EDS, UV Vis, XRD, and FT-IR spectroscopy have been carried out. RESULT: The synthesis yielded a 60,16%, conversion with a melting point of 216-218 oC and a conductivity value of 0.4 mS/cm. In vitro test results showed morphological changes (apoptosis) in MCF-7 cancer cells starting at a sample concentration of 250 µg/mL and an IC50 value of 618.40 µg/mL. Molecular docking study of Ni(II) cysteine-tyrosine dithiocarbamate complex identified with 4,4',4''-[(2R)-butane-1,1,2-triyl]triphenol - Estrogen α showing active site with acidic residue amino E323, M388, L387, G390 and I389. Hydrophobic and hydrophobic bonds are seen in Ni(II) cysteine-tyrosine dithiocarbamate - Estrogen α has a binding energy of -80.9429 kJ /mol. CONCLUSION: there were 5 residues responsible for maintaining the ligand binding stable. The compound had significant Hbond contact intensity, however, it was not strong enough to make a significant anticancer effect. Though the synthesized compound shows low bioactivity, this research is expected to give valuable insight into the effect of molecular structure on anticancer activity.

Anaplastic and poorly differentiated thyroid carcinomas: genetic evidence of high-grade transformation from differentiated thyroid carcinoma.

Anaplastic thyroid carcinoma (ATC) is the most advanced and aggressive thyroid cancer, and poorly differentiated thyroid carcinoma (PDTC) lacks anaplastic histology but has lost architectural and cytologic differentiation. Only a few studies have focused on the genetic relationship between the two advanced carcinomas and coexisting differentiated thyroid carcinomas (DTCs). In the present study, we investigated clinicopathologic features and genetic profiles in 57 ATC and PDTC samples, among which 33 cases had concomitant DTC components or DTC history. We performed immunohistochemistry for BRAF V600E, p53, and PD-L1 expression, Sanger sequencing for TERT promoter and RAS mutations, and fluorescence in situ hybridization for ALK and RET rearrangements. We found that ATCs and PDTCs shared similar gene alterations to their coexisting DTCs, and most DTCs were aggressive subtypes harboring frequent TERT promoter mutations. A significantly higher proportion of ATCs expressed p53 and PD-L1, and a lower proportion expressed PAX-8 and TTF-1, than the coexisting DTCs. Our findings provide more reliable evidence that ATCs and PDTCs are derived from DTCs.

Exploring the Antiproliferative and Modulatory Effects of 1-Methoxyisobrassinin on Ovarian Cancer Cells: Insights into Cell Cycle Regulation, Apoptosis, Autophagy, and Its Interactions with NAC.

Ovarian cancer, a highly lethal malignancy among reproductive organ cancers, poses a significant challenge with its high mortality rate, particularly in advanced-stage cases resistant to platinum-based chemotherapy. This study explores the potential therapeutic efficacy of 1-methoxyisobrassinin (MB-591), a derivative of indole phytoalexins found in Cruciferae family plants, on both cisplatin-sensitive (A2780) and cisplatin-resistant ovarian cancer cells (A2780 cis). The findings reveal that MB-591 exhibits an antiproliferative effect on both cell lines, with significantly increased potency against cisplatin-sensitive cells. The substance induces alterations in the distribution of the cell cycle, particularly in the S and G2/M phases, accompanied by changes in key regulatory proteins. Moreover, MB-591 triggers apoptosis in both cell lines, involving caspase-9 cleavage, PARP cleavage induction, and DNA damage, accompanied by the generation of reactive oxygen species (ROS) and mitochondrial dysfunction. Notably, the substance selectively induces autophagy in cisplatin-resistant cells, suggesting potential targeted therapeutic applications. The study further explores the interplay between MB-591 and antioxidant N-acetylcysteine (NAC), in modulating cellular processes. NAC demonstrates a protective effect against MB-591-induced cytotoxicity, affecting cell cycle distribution and apoptosis-related proteins. Additionally, NAC exhibits inhibitory effects on autophagy initiation in cisplatin-resistant cells, suggesting its potential role in overcoming resistance mechanisms.

Comparative study between poorly differentiated thyroid cancer and anaplastic thyroid cancer: real-world pathological distribution, death attribution, and prognostic factor estimation.

Background: The clinic-pathological boundary between poorly differentiated thyroid cancer (PDTC) and anaplastic thyroid cancer (ATC) is unclear due to a wide spectrum of histopathological features and the rarity of the disease. In addition to that, with the highest mortality rate and non-standard treatment modality, the PDTC/ATC population has not been subjected to comprehensive description and comparison with the extent of histological characteristics, therapeutic response, prognostic factors, and death attribution analysis. Method: A total of 4,947 PDTC/ATC patients from 2000 to 2018 were identified from the Surveillance, Epidemiology, and End Results (SEER) database. Kaplan-Meier survival curve estimation and Cox proportional hazard regression were applied. Results: Overall, the 5- and 10-year DSS for PDTC were 71.9% and 68.0%, respectively, whereas the 5- and 10-year OS are 59.3% and 51.2%, respectively. The median survival time for ATC patients was 3 months with 1-year OS being 26.9% and 1-year DSS being 31.2%. During the follow-up period, 68.1% of the PDTC/ATC cohort were dead, 51.6% of which were attributed to thyroid malignancies and 16.5% to non-thyroid causes. The top three common non-thyroid causes of death were miscellaneous cancers, lower respiratory system disease, and heart disease. The histological feature of papillary thyroid cancer (PTC) was the leading pathological category for PDTC patients (51.7%), whereas 76.7% of ATC patients' pathological feature was characterized as unidentifiable. Sarcoma histological characteristics found in ATC cases suffer the highest overall mortality (vs. PTC, HR = 2.61, 95% CI 1.68-4.06, P < 0.001). Older age unidentifiable histology feature, more advanced AJCC N1b, AJCC M1, and SEER stage, tumor size larger than 5 cm, and more invasive tumor extension were independent bad outcome predictors. Conclusion: The populational analysis of the PDTC/ATC cohort has provided reliable support for better understanding of the difference between PDTC and ATC cases and the guidance of clinical practice and further studies.

Cytotoxicity of bismuth(III) dithiocarbamate derivatives by promoting a mitochondrial-dependent apoptotic pathway and suppressing MCF-7 breast adenocarcinoma cell invasion.

We previously reported that the bismuth(III) dithiocarbamate derivative, bismuth diethyldithiocarbamate (1) exhibited greater cytotoxicity while inducing apoptosis via the intrinsic pathway in MCF-7 cells. We further evaluated the other bismuth(III) dithiocarbamate derivatives, Bi[S2CNR]3, with R = (CH2CH2OH)(iPr), (CH2)4, and (CH2CH2OH)(CH3), denoted as 2, 3, and 4, respectively, in the same MCF-7 cell line. 2-4 were found to exhibit IC50 values of 10.33 ± 0.06 µM, 1.07 ± 0.01 µM and 25.37 ± 0.12 µM, respectively, compared to that of cisplatin at 30.53 ± 0.23 µM. Apoptotic promotion via the mitochondrial-dependent pathway was due to the elevation of intracellular reactive oxygen species (ROS), promotion of caspases, release of cytochrome c, fragmentation of DNA, and results of staining assay observed in all compound-treated cells. 2-4 are also capable of suppressing MCF-7 cell invasion and modulate Lys-48 also Lys-63 linked polyubiquitination, leading to proteasomal degradation. Analysis of gene expression via qRT-PCR revealed their modulation, which supported all activities conducted upon treatment with 2-4. Altogether, bismuth dithiocarbamate derivatives, with bismuth(III) as the metal center bound to ligands, isopropyl ethanol, pyrrolidine, and methyl ethanol dithiocarbamate, are potential anti-breast cancer agents that induce apoptosis and suppress metastasis. Further studies using other breast cancer cell lines and in vivo studies are recommended to clarify the anticancer effects of these compounds.

Deep response to a combination of mTOR inhibitor temsirolimus and dual immunotherapy of nivolumab/ipilimumab in poorly differentiated thyroid carcinoma with PTEN mutation: a case report and literature review.

Treating advanced thyroid cancer presents challenges due to its resistance to various treatment modalities, thereby limiting therapeutic options. To our knowledge, this study is the first to report the efficacy of temsirolimus in conjunction with dual immunotherapy of nivolumab/ipilimumab to treat heavily treated advanced PDTC. A 50-year-old female initially presented with a rapidly enlarging mass on her right neck. Subsequent diagnosis revealed poorly differentiated thyroid carcinoma, leading to a total thyroidectomy followed by post-operative radioablation therapy. After four years, an examination for persistent cough revealed a recurrence of the disease within multiple mediastinal nodes. Genetic analysis of blood samples uncovered somatic mutations in the tumor, specifically involving PTEN and TP53. The disease progressed despite palliative radiation, lenvatinib, and nivolumab/ipilimumab therapy. Consequently, temsirolimus, functioning as an mTOR inhibitor, was introduced as an adjunct to the nivolumab/ipilimumab regimen. This combination approach yielded remarkable clinical improvement and disease control for a duration of approximately six months. Temsirolimus likely suppressed the aberrantly activated PI3K/AKT/mTOR signaling pathway, facilitated by the PTEN genetic alteration, thus engendering an effective treatment response. This synergy between targeted agents and immunotherapy presents a promising therapeutic strategy for advanced PDTC patients with limited treatment alternatives. In previous clinical trials, mTOR inhibitors have demonstrated the ability to maintain stable disease (SD) in 65% to 74% for advanced thyroid cancer patients, including those with PDTC. When combined with other targeted therapies, the observed SD or partial response rates range from 80% to 97%. Many of these trials primarily involved differentiated thyroid carcinoma, with diverse genetic mutations. Thyroid cancer patients with alterations in the PI3K/mTOR/Akt appeared to benefit most from mTOR inhibitors. However, no clear association between the efficacy of mTOR inhibitors and specific histologies or genetic mutations has been established. Future studies are warranted to elucidate these associations.

Dissipation and evaluation of different treatments on the residues of different insecticides in/on cauliflower curd.

A field study to understand dissipation rates and effect of various washing treatments on the residues of seven different insecticides, i.e. tetraniliprole 200 SC, emamectin benzoate 5 SG, lufenuron 5.4 EC, indoxacarb 14.5 SC, thiodicarb 75 WP, profenofos 50 EC and cypermethrin 25 EC in/on cauliflower curd has been conducted. The results showed that initial deposits (just after the last insecticide application, i.e. 0 d) of tetraniliprole, emamectin benzoate, lufenuron, indoxacarb, thiodicarb, profenofos and cypermethrin were 0.43, 0.03, 0.25, 0.28, 0.38, 6.70 and 0.68 mg kg-1, respectively. The dissipation pattern of all the tested insecticides followed monophasic, first order kinetics with the half-lives of 6.25, 8.85, 3.27, 7.71, 4.36, 2.98 and 3.76 d, respectively. Proposed pre-harvest intervals for these insecticides are 6, 9, 3, 8, 4, 3 and 4 d, respectively. All the decontamination techniques showed reductions in residue levels. However, treatment by soaking in 5% sodium bicarbonate aqueous solution showed 54%, 42%, 53%, 48%, 22%, 54% and 77% maximum reductions in residues of tetraniliprole, emamectin benzoate, lufenuron, indoxacarb, profenofos and cypermethrin, respectively, in cauliflower curds. The next best treatment was soaking cauliflower curds in water at 45-50 °C for 10 min, which reduced the residues of cypermethrin, profenofos, tetraniliprole, thiodicarb, emamectin benzoate and lufenuron.

M2 macrophage­derived exosomes alleviate KCa3.1 channel expression in rapidly paced HL­1 myocytes via the NF­κB (p65)/STAT3 signaling pathway.

The present study was designed to explore the role of M2 macrophage­derived exosomes (M2­exos) on the KCa3.1 channel in a cellular atrial fibrillation (AF) model using rapidly paced HL­1 myocytes. M2 macrophages and M2­exos were isolated and identified. MicroRNA (miR)­146a­5p levels in M2 macrophages and M2­exos were quantified using reverse transcription­quantitative PCR (RT­qPCR). HL­1 myocytes were randomly divided into six groups: Control group, pacing group, pacing + coculture group (pacing HL­1 cells cocultured with M2­exos), pacing + mimic­miR­146a­5p group, pacing + NC­miR­146a­5p group and pacing + pyrrolidine dithiocarbamate (PDTC; a special blocker of the NF­κB signaling pathway) group. Transmission electron microscopy, nanoparticle tracking analysis, western blotting, RT­qPCR and immunohistochemistry were performed in the present study. A whole­cell clamp was also applied to record the current density of KCa3.1 and action potential duration (APD) in each group. The results revealed that miR­146a­5p was highly expressed in both M2 macrophages and M2­exos. Pacing HL­1 cells led to a shorter APD, an increased KCa3.1 current density and higher protein levels of KCa3.1, phosphorylated (p­)NF­κB p65, p­STAT3 and IL­1ß compared with the control group. M2­exos, miR­146a­5p­mimic and PDTC both reduced the protein expression of KCa3.1, p­NF­κB p65, p­STAT3 and IL­1ß and the current density of KCa3.1, resulting in a longer APD in the pacing HL­1 cells. In conclusion, M2­exos and their cargo, which comprised miR­146a­5p, decreased KCa3.1 expression and IL­1ß secretion in pacing HL­1 cells via the NF­κB/STAT3 signaling pathway, limiting the shorter APD caused by rapid pacing.

The frequency of mutations in advanced thyroid cancer in Japan: a single-center study.

We analyzed the outcomes of genetic testing to study the frequency of mutations in advanced thyroid cancer in Japan. Patients (n = 96) with unresectable or metastatic thyroid carcinoma were included for retrospective chart review. Results of gene panel testing, which was performed between May 2020 and April 2023, were analyzed. The median age of the patients was 73.5 years (range, 17-88); 59 were women, and 39 were men. Overall, 17 patients had anaplastic thyroid carcinoma (ATC), 68 had papillary thyroid carcinoma (PTC), 7 had follicular thyroid carcinoma, and 6 had poorly differentiated thyroid carcinoma (PDTC). Of the 81 patients with differentiated thyroid carcinoma (DTC) and PDTC, 88.9% were radioactive iodine-refractory, and 32.7% of all cases had previously been treated with multiple kinase inhibitors. Of ATC cases, 52.9% had BRAF mutations, and 5.9% had RET fusion. Of PTC cases, 83.1% had BRAF mutations, 9.2% had RET fusion, and 1.5% had NTRK fusion. One case each of ATC and PTC had a tumor mutation burden of ≥10. ATC cases had a significantly higher prevalence of TP53 alterations than the other cases (82.3% vs. 11.8%), whereas the frequencies of TERT promoter mutations were 88.2% in ATC cases and 64.7% in the other cases, albeit without a significant difference. In conclusion, 58.8% of ATC, 93.8% of PTC, and 42.9% of PDTC had genetic alterations linked to therapeutic agents. Active gene panel testing is required to increase treatment options.

Characterization of the Thyroid Cancer Genomic Landscape by Plasma-Based Circulating Tumor DNA Next-Generation Sequencing.

Background: The limited availability of targeted therapies in thyroid cancer (TC) has challenged conventional treatment algorithms and has established urgency for the identification of targetable genomic abnormalities. In addition to widely adopted tissue-based next-generation sequencing (NGS), plasma-based circulating tumor DNA (ctDNA) NGS is rapidly emerging as a genomic biomarker detection method and is steadily gaining utility across solid tumors. To date, plasma-based genomic alterations in TC have not been determined. Herein, we profile potential actionable mutations detected through ctDNA in patients with TC subtypes. Methods: A retrospective data analysis of the Guardant Health, Inc. database was performed using the commercially available Guardant360® plasma-NGS test on TC samples from adult patients collected between 2016 and 2021. The landscape of genomic alterations and blood tumor mutation burden (bTMB) were analyzed in patients with different types of TC: anaplastic TC (ATC), papillary TC (PTC), follicular TC (FTC), oncolytic carcinoma of the thyroid (OCA), poorly differentiated TC (PDTC), medullary TC (MTC), and TC not otherwise specified (TC NOS). Results: Of the 1094 patients included most of the patients n = 876 had TC NOS, and 20% had a specific diagnosis (92 ATC, 62 PTC, 14 FTC, 16 OCA, 2 PDTC, and 32 MTC patients). The median age was 65 (range 10-98) and 47.3% were male. 78.3% of patients had one or more genomic alteration detected by ctDNA NGS. TP53 (46.9%) was the most common mutation detected among all TC. BRAFV600E was detected in 27.2% of ATC, 35.7% of PTC, and in none of FTC. RAS was detected in 18.5% of ATC, 11.9% of PTC, and 62.5% of FTC. RET, ALK, and NTRK fusions were seen in 1.1%, 0.5%, and 0.2% of all TC, respectively. RET mutations were detected in 66.7% of MTC. bTMB analysis was performed on 159 patients. The mean bTMB was higher in ATC compared with other types of TC (p = 0.0011, 0.0557, and <0.0001, respectively). Conclusions: Plasma-based comprehensive NGS is a promising NGS method in TC; however, future validation of the clinical utility by analysis of paired tumor and plasma samples is needed.

Pelargonidin alleviates acrolein-induced inflammation in human umbilical vein endothelial cells by reducing COX-2 expression through the NF-κB pathway.

Acrolein, a common environmental pollutant, is linked to the development of cardiovascular inflammatory diseases. Pelargonidin is a natural compound with anti-inflammation activity. In this study, we aimed to explore the effects of pelargonidin on inflammation induced by acrolein in human umbilical vein endothelial cells (HUVECs). MTT assay was utilized for assessing cell viability in HUVECs. LDH release in HUVECs was measured using the LDH kit. Western blot was used to detect the protein expression of p-p65, p65 and COX-2. Inflammation was evaluated through determining the levels of PGE2, IL-1ß, IL-6, IL-8 and TNF-α in HUVECs after treatment. COX-2 mRNA expression and COX-2 content were examined using RT-qPCR and a human COX-2 ELISA kit, respectively. Acrolein treatment at 50 µM resulted in a 45% decrease in the viability and an increase in LDH release (2.2-fold) in HUVECs. Pelargonidin at 5, 10, 20, and 40 µM alleviated acrolein-caused inhibitory effect on cell viability (increased to 1.3-, 1.5-, 1.8-, and 1.9-fold, respectively, compared to acrolein treatment group) and promoting effect on LDH release (decreased to 82%, 75%, 62%, and 58%, respectively, compared to acrolein treatment group) in HUVECs. Moreover, pelargonidin or pyrrolidine dithiocarbamate (PDTC; an NF-κB pathway inhibitor) inhibited acrolein-induced activation of the NF-κB pathway. Acrolein elevated the levels of PGE2, IL-1ß, IL-6, IL-8 and TNF-α (from 40.2, 27.3, 67.2, 29.0, 24.8 pg/mL in control group to 224.0, 167.3, 618.3, 104.6, and 275.1 pg/mL in acrolein treatment group, respectively), which were retarded after pelargonidin (decreased to 134.8, 82.3, 246.2, 70.2, and 120.8 pg/mL in acrolein + pelargonidin treatment group) or PDTC (decreased to 107.9, 80.1, 214.6, 64.0, and 96.6 pg/mL in acrolein + PDTC treatment group) treatment in HUVECs. Pelargonidin inactivated the NF-κB pathway to reduce acrolein-induced COX-2 expression. Furthermore, pelargonidin relieved acrolein-triggered inflammation through decreasing COX-2 expression by inactivating the NF-κB pathway in HUVECs. In conclusion, pelargonidin could protect against acrolein-triggered inflammation in HUVECs through attenuating COX-2 expression by inactivating the NF-κB pathway.

The Frequency of Differentiated Thyroid Cancer Recurrence in 2302 Patients With Excellent Response to Primary Therapy.

CONTEXT: Discrepant data on the recurrence rate of differentiated thyroid cancer (DTC) are reported. OBJECTIVE: To evaluate the frequency and risk factors of true recurrence in DTC patients with excellent responses (ExR) to initial therapy. METHODS: A retrospective analysis of the 2302 consecutive DTC patients with ExR to primary therapy, treated during 24 years at single center. The percentage of recurrence and cumulative recurrence rate (CRR) were analyzed. Risk factors for recurrence for patients with papillary thyroid cancer (PTC) were investigated and methods for establishing a diagnosis of recurrence were evaluated. RESULTS: Of DTC patients, 32 (1.4%) experienced recurrence. PTC patients with recurrence were more likely to have younger age (P = .0182), larger tumor size (P = .0013), lymph node metastases (P = .0013), incomplete resection (P = .0446), higher ATA risk (P = .0002), and had more frequently been treated with 131I (P = .0203). CRRs at 5, 10, 15, 20, and 24 years after surgery were 1.2%, 1.9%, 2.5%, 2.9%, and 2.9%, respectively. The CRRs according to histological type were highest for poorly differentiated thyroid cancer (PDTC), lower for oncocytic (OTC) and follicular thyroid cancer (FTC), and lowest for PTC. Most recurrences occurred within the first 5 years of observation. The most effective method for detecting local recurrence was ultrasonography with fine needle aspiration cytology, and for distant metastases, 18F-FDG PET. CONCLUSION: True recurrence is rare in DTC patients. PTC patients with ExR to primary therapy and N0/Nx can be dismissed from oncological follow-up. Despite ExR to primary therapy, DTC patients with N1, and PDTC, OTC, FTC should remain under oncological follow-up.

TNF-α-induced down-regulation of type I interferon receptor contributes to acquired resistance of cervical squamous cancer to Cisplatin.

We aimed to investigate the effects of tumor necrosis factor (TNF)-α on the expression of interferon α/ß receptor subunit 1 (IFNAR1) and cervical squamous cancer (CSCC) resistance to Cisplatin, as well as the underlying mechanisms. Kaplan-Meier analysis was used to plot the overall survival curves. SiHa cells were treated with 20 ng/ml TNF-α to determine cell proliferation in human CSCC cells and the expression of IFNAR1. The effects of TNF-α on the downstream signaling pathway, including casein kinase 1α (CK1α), were investigated using the caspase protease inhibitor FK009, the c-Jun kinase inhibitor SP600125, and the nuclear factor kappa-B inhibitor ammonium pyrrolidinedithiocarbamate (PDTC). TNF-α induced down-regulation of IFNAR1 in human CSCC cells and promoted proliferation of SiHa cells. SiHa cells were transfected with the catalytic inactive mutant CK1α K49A, and the ability of TNF-α to induce down-regulation of IFNAR1 expression was found to be significantly diminished in this context. FK009 and PDTC had no obvious effect on the expression of CK1α, however, SP600125 significantly reduced the expression of CK1α in the presence of TNF-α. SiHa cells treated with TNF-α showed reduced sensitivity to Cisplatin and exhibited higher cell viability, while the sensitivity of SiHa cells to Cisplatin was restored after treatment with CK1α inhibitor D4476. Additionally, we constructed a TNF-α overexpressing SiHa cell line and a transplanted tumor model. The results were similar to those of in vitro efficacy. We demonstrate that TNF-α-induced down-regulation of type I interferon receptor contributes to acquired resistance of cervical squamous cancer to Cisplatin.

CDK4 phosphorylation status and rational use for combining CDK4/6 and BRAF/MEK inhibition in advanced thyroid carcinomas.

Background: CDK4/6 inhibitors (CDK4/6i) have been established as standard treatment against advanced Estrogen Receptor-positive breast cancers. These drugs are being tested against several cancers, including in combinations with other therapies. We identified the T172-phosphorylation of CDK4 as the step determining its activity, retinoblastoma protein (RB) inactivation, cell cycle commitment and sensitivity to CDK4/6i. Poorly differentiated (PDTC) and anaplastic (ATC) thyroid carcinomas, the latter considered one of the most lethal human malignancies, represent major clinical challenges. Several molecular evidence suggest that CDK4/6i could be considered for treating these advanced thyroid cancers. Methods: We analyzed by two-dimensional gel electrophoresis the CDK4 modification profile and the presence of T172-phosphorylated CDK4 in a collection of 98 fresh-frozen tissues and in 21 cell lines. A sub-cohort of samples was characterized by RNA sequencing and immunohistochemistry. Sensitivity to CDK4/6i (palbociclib and abemaciclib) was assessed by BrdU incorporation/viability assays. Treatment of cell lines with CDK4/6i and combination with BRAF/MEK inhibitors (dabrafenib/trametinib) was comprehensively evaluated by western blot, characterization of immunoprecipitated CDK4 and CDK2 complexes and clonogenic assays. Results: CDK4 phosphorylation was detected in all well-differentiated thyroid carcinomas (n=29), 19/20 PDTC, 16/23 ATC and 18/21 thyroid cancer cell lines, including 11 ATC-derived ones. Tumors and cell lines without phosphorylated CDK4 presented very high p16CDKN2A levels, which were associated with proliferative activity. Absence of CDK4 phosphorylation in cell lines was associated with CDK4/6i insensitivity. RB1 defects (the primary cause of intrinsic CDK4/6i resistance) were not found in 5/7 tumors without detectable phosphorylated CDK4. A previously developed 11-gene expression signature identified the likely unresponsive tumors, lacking CDK4 phosphorylation. In cell lines, palbociclib synergized with dabrafenib/trametinib by completely and permanently arresting proliferation. These combinations prevented resistance mechanisms induced by palbociclib, most notably Cyclin E1-CDK2 activation and a paradoxical stabilization of phosphorylated CDK4 complexes. Conclusion: Our study supports further clinical evaluation of CDK4/6i and their combination with anti-BRAF/MEK therapies as a novel effective treatment against advanced thyroid tumors. Moreover, the complementary use of our 11 genes predictor with p16/KI67 evaluation could represent a prompt tool for recognizing the intrinsically CDK4/6i insensitive patients, who are potentially better candidates to immediate chemotherapy.

Photobiomodulation reduces neuropathic pain after spinal cord injury by downregulating CXCL10 expression.

BACKGROUND: Many studies have recently highlighted the role of photobiomodulation (PBM) in neuropathic pain (NP) relief after spinal cord injury (SCI), suggesting that it may be an effective way to relieve NP after SCI. However, the underlying mechanisms remain unclear. This study aimed to determine the potential mechanisms of PBM in NP relief after SCI. METHODS: We performed systematic observations and investigated the mechanism of PBM intervention in NP in rats after SCI. Using transcriptome sequencing, we screened CXCL10 as a possible target molecule for PBM intervention and validated the results in rat tissues using reverse transcription-polymerase chain reaction and western blotting. Using immunofluorescence co-labeling, astrocytes and microglia were identified as the cells responsible for CXCL10 expression. The involvement of the NF-κB pathway in CXCL10 expression was verified using inhibitor pyrrolidine dithiocarbamate (PDTC) and agonist phorbol-12-myristate-13-acetate (PMA), which were further validated by an in vivo injection experiment. RESULTS: Here, we demonstrated that PBM therapy led to an improvement in NP relative behaviors post-SCI, inhibited the activation of microglia and astrocytes, and decreased the expression level of CXCL10 in glial cells, which was accompanied by mediation of the NF-κB signaling pathway. Photobiomodulation inhibit the activation of the NF-κB pathway and reduce downstream CXCL10 expression. The NF-κB pathway inhibitor PDTC had the same effect as PBM on improving pain in animals with SCI, and the NF-κB pathway promoter PMA could reverse the beneficial effect of PBM. CONCLUSIONS: Our results provide new insights into the mechanisms by which PBM alleviates NP after SCI. We demonstrated that PBM significantly inhibited the activation of microglia and astrocytes and decreased the expression level of CXCL10. These effects appear to be related to the NF-κB signaling pathway. Taken together, our study provides evidence that PBM could be a potentially effective therapy for NP after SCI, CXCL10 and NF-kB signaling pathways might be critical factors in pain relief mediated by PBM after SCI.

Molinate induces organ defects by promoting apoptosis, inflammation, and endoplasmic reticulum stress during the developmental stage of zebrafish.

Molinate is classified as a thiocarbamate herbicide and is mainly used in paddy fields to culture rice. However, the toxic effects of molinate and the associated mechanisms in the process of development have not been completely elucidated. Therefore, in the present study, we demonstrated that molinate reduced the viability of zebrafish larvae and the probability of successful hatching using zebrafish (Danio rerio), one of the remarkable in vivo models for testing the toxicity of chemicals. In addition, molinate treatment triggered the occurrence of apoptosis, inflammation, and endoplasmic reticulum (ER) stress response in zebrafish larvae. Furthermore, we identified that an abnormal cardiovascular phenotype through wild type zebrafish, neuronal defects through transgenic olig2:dsRed zebrafish, and developmental toxicity in the liver through transgenic lfabp:dsRed zebrafish. Collectively, these results provide evidence of the hazardous effects of molinate on the developmental stage of non-target organisms by elucidating the toxic mechanisms of molinate in developing zebrafish.

Reactivity of a Dithiocarbamate-Ligated [WVI≡S] Complex with Hydride Donors: Toward a Synthetic Mimic of Formate Dehydrogenase.

Formate dehydrogenase (FDH) enzymes catalyze redox interconversion of CO2 and HCO2-, with a key mechanistic step being the transfer of H- from HCO2- to an oxidized active site featuring a [MVI≡S] group in a sulfur-rich environment (M = Mo or W). Here, we report reactivity studies with HCO2- and other reducing agents of a synthetic [WVI≡S] model complex ligated by dithiocarbamate (dtc) ligands. Reactions of [WVIS(dtc)3][BF4] (1) conducted in MeOH solvent generated [WVIS(S2)(dtc)2] (2) and [WVS(µ-S)(dtc)]2 (3) products by a solvolysis pathway that was accelerated by the presence of [Me4N][HCO2] but did not require it. Under MeOH-free conditions, the reaction of 1 with [Et4N][HCO2] produced some [WIV(µ-S)(µ-dtc)(dtc)]2 (4), but predominantly [WV(dtc)4]+ (5), along with stoichiometric CO2 detected by headspace gas chromatography (GC) analysis. Stronger hydride sources such as K-selectride generated the more reduced analogue, 4, exclusively. The reaction of 1 with the electron donor, CoCp2, also produced 4 and 5 in varying amounts depending on reaction conditions. These results indicate that formates and borohydrides act as electron donors rather than hydride donors toward 1, an outcome that diverges from the behavior of FDHs. The difference is ascribed to the more oxidizing potential of [WVI≡S] complex 1 when supported by monoanionic dtc ligands that allows electron transfer to outcompete hydride transfer, as compared to the more reduced [MVI≡S] active sites supported by dianionic pyranopterindithiolate ligands in FDHs.

Masking thiol reactivity with thioamide, thiourea, and thiocarbamate-based MBPs.

Thioamides, thioureas, and thiocarbamates are introduced as stable, sulfur-based metal-binding pharmacophores (MBPs) for use in metalloenzyme fragment-based drug discovery (mFBDD). MBP reactivity, bioactivity, and structural studies show that these molecules can act as ligands for Zn(II)-dependent metalloenzymes including human carbonic anhydrase II (hCAII) and matrix metalloproteinase-2 (MMP-2).