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In many research settings, the intervention implied by the average causal effect of a time-varying exposure is impractical or unrealistic, and we might instead prefer a more realistic target estimand. Instead of requiring all individuals to be always exposed versus unexposed, incremental effects quantify the impact of merely shifting each individual's probability of being exposed.We demonstrate the estimation of incremental effects in the time-varying setting, using data from the Effects of Aspirin in Gestation and Reproduction trial, which assessed the effect of preconception low-dose aspirin on pregnancy outcomes. Compliance to aspirin or placebo was summarized weekly and was affected by time-varying confounders such as bleeding or nausea. We sought to estimate what the incidence of pregnancy by 26 weeks postrandomization would have been if we shifted each participant's probability of taking aspirin or placebo each week by odds ratios (OR) between 0.30 and 3.00.Under no intervention (OR = 1), the incidence of pregnancy was 77% (95% CI: 74%, 80%). Decreasing women's probability of complying with aspirin had little estimated effect on pregnancy incidence. When we increased women's probability of taking aspirin, estimated incidence of pregnancy increased, from 83% (95% confidence interval [CI] = 79%, 87%) for OR = 2 to 89% (95% CI = 84%, 93%) for OR=3. We observed similar results when we shifted women's probability of complying with a placebo.These results estimated that realistic interventions to increase women's probability of taking aspirin would have yielded little to no impact on the incidence of pregnancy, relative to similar interventions on placebo.

Colorectal cancer (CRC) has a high mortality rate among cancers worldwide. To reduce this mortality rate, chemotherapy (5-fluorouracil, oxaliplatin, and irinotecan) or targeted therapy (bevacizumab, cetuximab, and panitumumab) has been used to treat CRC. However, due to various side effects and poor responses to CRC treatment, novel therapeutic targets for drug development are needed. In this study, we identified the overexpression of EHMT1 in CRC using RNA sequencing (RNA-seq) data derived from TCGA, and we observed that knocking down EHMT1 expression suppressed cell growth by inducing cell apoptosis in CRC cell lines. In Gene Ontology (GO) term analysis using RNA-seq data, apoptosis-related terms were enriched after EHMT1 knockdown. Moreover, we identified the CHOP gene as a direct target of EHMT1 using a ChIP (chromatin immunoprecipitation) assay with an anti-histone 3 lysine 9 dimethylation (H3K9me2) antibody. Finally, after cotransfection with siEHMT1 and siCHOP, we again confirmed that CHOP-mediated cell apoptosis was induced by EHMT1 knockdown. Our findings reveal that EHMT1 plays a key role in regulating CRC cell apoptosis, suggesting that EHMT1 may be a therapeutic target for the development of cancer inhibitors.

A high-throughput cell-based screen identified redox-active small molecules that produce a period lengthening of the circadian rhythm. The strongest period lengthening phenotype was induced by a phenazine carboxamide (VU661). Comparison to two isomeric benzquinoline carboxamides (VU673 and VU164) shows the activity is associated with the redox modulating phenazine functionality. Furthermore, ex vivo cell analysis using optical redox ratio measurements shows the period lengthening phenotype to be associated with a shift to the NAD/FAD oxidation state of nicotinamide and flavine coenzymes.

Introduction: This study reports the effect of herbal medicine (Samulanshin-tang-gamibang) and acupuncture on complete atrioventricular (AV) block.Case presentation: A 63-year-old female with complete AV block was experiencing dyspnea, palpitation, dizziness, headache, bradycardia, and insomnia, and she was treated with Samulanshin-tang-gamibang and acupuncture for 12 days. To evaluate the treatment, a numeric rating scale (NRS) and the New York Heart Association (NYHA) functional classification was used. The patient’s NRS scores decreased from 6 to 2 for dyspnea and palpitation and from 5 to 1 for dizziness and headache. Her NYHA Class improved from Class II to Class I. No side effects were observed during treatment.Conclusion: This study suggests that herbal medicine and acupuncture may be effective in relieving symptoms caused by complete AV block. However, the long-term effects of the treatment were not observed, and so further studies are still needed.

The purpose of this report is to describe the effects of Qingfei Paidu decoction (QPD) and QPD without Ephedra Herba on COVID-19 patients. One COVID-19 patient was treated with QPD and two others who had been sensitive to ephedra were treated with QPD without it. The fever, cough, and sore throat of all three were monitored three times a day through non-face-to-face assessment for a total of five to seven days, and cough and throat pain changes were evaluated with a numeric rating scale (NRS). After treatment, the fever and NRS scores were improved for all three patients. These results indicate that both QPD and QPD without Ephedra Herba is an effective treatment for COVID-19 symptoms.

The human microbiome plays an essential role in the human immune system, food digestion, and protection from harmful bacteria by colonizing the human intestine. Recently, although the human microbiome affects colorectal cancer (CRC) treatment, the mode of action between the microbiome and CRC remains unclear. This study showed that propionate suppressed CRC growth by promoting the proteasomal degradation of euchromatic histone-lysine N-methyltransferase 2 (EHMT2) through HECT domain E3 ubiquitin protein ligase 2 (HECTD2) upregulation. In addition, EHMT2 downregulation reduced the H3K9me2 level on the promoter region of tumor necrosis factor α-induced protein 1 (TNFAIP1) as a novel direct target of EHMT2. Subsequently, TNFAIP1 upregulation induced the apoptosis of CRC cells. Furthermore, using Bacteroides thetaiotaomicron culture medium, we confirmed EHMT2 downregulation via upregulation of HECTD2 and TNFAIP1 upregulation. Finally, we observed the synergistic effect of propionate and an EHMT2 inhibitor (BIX01294) in 3D spheroid culture models. Thus, we suggest the anticancer effects of propionate and EHMT2 as therapeutic targets for colon cancer treatment and may provide the possibility for the synergistic effects of an EHMT2 inhibitor and microbiome in CRC treatment.

N-acyl-phosphatidylethanolamine hydrolyzing phospholipase D (NAPE-PLD) is a zinc metallohydrolase that hydrolyzesN-acyl-phosphatidylethanolamine (NAPEs) to formN-acyl-ethanolamides (NAEs) and phosphatidic acid. Several lines of evidence suggest that reduced NAPE-PLD activity could contribute to cardiometabolic diseases. For instance,NAPEPLDexpression is reduced in human coronary arteries with unstable atherosclerotic lesions, defective efferocytosis is implicated in the enlargement of necrotic cores of these lesions, and NAPE-PLD products such as palmitoylethanolamide and oleoylethanolamide have been shown to enhance efferocytosis. Thus, enzyme activation mediated by a small molecule may serve as a therapeutic treatment for cardiometabolic diseases. As a proof-of-concept study, we sought to identify small molecule activators of NAPE-PLD. High-throughput screening followed by hit validation and primary lead optimization studies identified a series of benzothiazole phenylsulfonyl-piperidine carboxamides that variably increased activity of both mouse and human NAPE-PLD. From this set of small molecules, two NAPE-PLD activators (VU534 and VU533) were shown to increase efferocytosis by bone-marrow derived macrophages isolated from wild-type mice, while efferocytosis was significantly reduced inNapepld-/-BMDM or after Nape-pld inhibition. Together these studies demonstrate an essential role for NAPE-PLD in the regulation of efferocytosis and the potential value of NAPE-PLD activators as a strategy to treat cardiometabolic diseases.

Objectives: The aim of this study was to evaluate the effectiveness of additional administration of Banha-sasim-tang formulation to patients with gastrointestinal symptoms that did not improve with Western medicines.Methods: The patients who met the inclusion and exclusion criteria were analyzed retrospectively. A paired t-test was performed on the Nepean Dyspepsia Index-Korean version (NDI-K) scores before and after administering the Banha-sasim-tang formulation.Results: A total of 27 subjects were included. The mean duration of taking the Banha-sasim-tang formulation was 15.6±3.1 days. The most commonly used Western medicine before taking the Banha-sasim-tang formulation was rebamipide. The means of the NDI-K score were 57.4±21.8 before administering Banha-sasim-tang formulation and 34.9±16.5 after administration, and the difference was statistically significant (p

The lipid peroxidation product malondialdehyde and the DNA peroxidation product base-propenal react with dG to generate the exocyclic adduct, M1dG. This mutagenic lesion has been found in human genomic and mitochondrial DNA. M1dG in genomic DNA is enzymatically oxidized to 6-oxo-M1dG, a lesion of currently unknown mutagenic potential. Here, we report the synthesis of an oligonucleotide containing 6-oxo-M1dG and the results of extension experiments aimed at determining the effect of the 6-oxo-M1dG lesion on the activity of human polymerase iota (hPol ι). For this purpose, a liquid chromatography-tandem mass spectrometry (LC-MS/MS) assay was developed to obtain reliable quantitative data on the utilization of poorly incorporated nucleotides. Results demonstrate that hPol ι primarily incorporates deoxycytidine triphosphate (dCTP) and thymidine triphosphate (dTTP) across from 6-oxo-M1dG with approximately equal efficiency, whereas deoxyadenosine triphosphate (dATP) and deoxyguanosine triphosphate (dGTP) are poor substrates. Following the incorporation of a single nucleotide opposite the lesion, 6-oxo-M1dG blocks further replication by the enzyme.

The Receptor for Advanced Glycation End products (RAGE) is a pattern recognition receptor that signals for inflammation via the NF-κB pathway. RAGE has been pursued as a potential target to suppress symptoms of diabetes and is of interest in a number of other diseases associated with chronic inflammation, such as inflammatory bowel disease and bronchopulmonary dysplasia. Screening and optimization have previously produced small molecules that inhibit the activity of RAGE in cell-based assays, but efforts to develop a therapeutically viable direct-binding RAGE inhibitor have yet to be successful. Here, we show that a fragment-based approach can be applied to discover fundamentally new types of RAGE inhibitors that specifically target the ligand-binding surface. A series of systematic assays of structural stability, solubility, and crystallization were performed to select constructs of the RAGE ligand-binding domain and optimize conditions for NMR-based screening and co-crystallization of RAGE with hit fragments. An NMR-based screen of a highly curated ~14 000-member fragment library produced 21 fragment leads. Of these, 3 were selected for elaboration based on structure-activity relationships generated through cycles of structural analysis by X-ray crystallography, structure-guided design principles, and synthetic chemistry. These results, combined with crystal structures of the first linked fragment compounds, demonstrate the applicability of the fragment-based approach to the discovery of RAGE inhibitors. This article is protected by copyright. All rights reserved.

Electroless composite plating enables uniform and thin surface treatment along with composite deposition using nanoparticles. Among such particles, polytetrafluoroethylene (PTFE) is capable of dry lubrication because of its self-lubricating properties. Specifically, the PTFE content in a plating layer increases with the concentration of PTFE in the plating bath. However, a high concentration of PTFE interferes with the co-deposition of Ni and P, thereby reducing the plating speed. Additionally, PTFE is unevenly deposited on the surface of the plating layer. Consequently, a method for increasing the PTFE content at low PTFE concentrations is required. Therefore, in this study, a stirring process in a low-PTFE-concentration plating bath and a process wherein PTFE precipitates on a specimen without stirring were combined. The PTFE content of the plated layer deposited on high carbon steel, plated layer deposition rate, average friction co-efficient, static contact angle, and surface energy were evaluated as 25.96%, 3.44 µm/40 min, 0.195, 141.9°, and 2.74 mN/m, respectively. This technique prevented the decrease in the deposition rate of the plating layer and led to high PTFE content in the plating layer. Notably, even a thin plating layer (5 μm or less in thickness) showed excellent surface characteristics.

Chemical synthesis has been described as a central science. Its practice provides access to the chemical structures of known and/or designed function. In particular, human health is greatly impacted by synthesis that enables advancements in both basic science discoveries in chemical biology as well as translational research that can lead to new therapeutics. To support the chemical synthesis needs of investigators across campus, the Vanderbilt Institute of Chemical Biology established a chemical synthesis core as part of its foundation in 2008. Provided in this Review are examples of synthetic products, known and designed, produced in the core over the past 10 years.

We have previously identified that in the absence of MET amplification or mutation, RTK hyperphosphorylation, indicative of its active state, may contribute to resistance to the EGFR-directed therapeutic, cetuximab in colorectal cancer (CRC). In our in vitro 3D cultures and in vivo xenografts, we showed that inhibition of MET (and RON) by crizotinib can overcome cetuximab resistance in CRC lines and xenografts. We now show that MET may get hyperphosphorylated by the autocrine expression and maturation of its ligand, HGF. This non-genetic mode of cetuximab resistance contrasts with the usual genetic modes of CRC cetuximab resistance, with majority of them being RAS/RAF mutations. Exogenous addition as well as autocrine overexpression of HGF led to cetuximab resistance, which could be overcome by MET inhibition by crizotinib. Interestingly, HGF/HGFL are synthesized as inactive precursors and require cleavage by proteases (HGFA, Matriptase, and Hepsin) to be biologically active. A survey of TCGA datasets indicated that HGF/HGFL were overexpressed in several CRC CMS subtypes, while the endogenous inhibitor of HGF proteases, HAI-I, was downregulated across all CRC CMS subtypes. We next expressed human full-length HGF in cetuximab-sensitive CC cells and observed that HGF overexpression imparts cetuximab resistance. Moreover, cetuximab resistance induced by upstream HGF overexpression could be overcome by the downstream MET inhibition. Next, in a subset of KRAS WT CRC patient-derived xenografts (PDXs) we tested and found that at majority of PDXs exhibit elevated pMET/pRON levels. RNA-seq-based principal component analysis showed that the maximal differences in expression among PDXs could be explained by their pMET/pRON phosphorylation status. Next, we generated nude mice xenografts for one PDX with high pMET/pRON levels and showed that cetuximab inhibition alone was not sufficient to inhibit regrowth of tumors after cessation of drug treatment, but a combined cetuximab/crizotinib treatment was, indicating endogenous MET/RON activation contributes to cetuximab resistance. In a parallel tumor stratification strategy, we have optimized conditions to generate [18F]crizotinib and are testing its efficacy as a new non-invasive PET-based imaging probe to identify crizotinib-avid tumors with high MET/RON activity. Together, these studies may provide novel mechanistic insights into RTK cross-talk and provide a positive predictive biomarker for precision therapy in colorectal cancer. Citation Format: Vivian T. Jones, Ramona Graves-Deal, KyuOK Jeon, Adam J. Rosenberg, Zheng Cao, Galina T. Bogatcheva, Sarah J. Harmych, James N. Higginbotham, Kwangho Kim, James W. Janetka, Gary A. Sulikowski, Bhuminder Singh. Role of MET signaling in cetuximab resistance and generation of an imaging biomarker to identify MET-driven cetuximab resistance in colorectal cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2562.

Several phase 1/2 clinical trials showed that low-dose interleukin-2 (IL-2) treatment is a safe and effective strategy for the treatment of chronic graft-versus-host disease, hepatitis C virus-induced vasculitis, and type 1 diabetes. Ulcerative colitis (UC) is a chronic inflammatory condition of the colon that lacks satisfactory treatment. In this study, we aimed to determine the effects of low-dose IL-2 as a therapeutic for UC on dextran sulfate sodium (DSS)-induced colitis. Methods: Mice with DSS-induced colitis were intraperitoneally injected with low-dose IL-2. Survival, body weight, disease activity index, colon length, histopathological score, myeloperoxidase activity and inflammatory cytokine levels as well as intestinal barrier integrity were examined. Differential gene expression after low-dose IL-2 treatment was analyzed by RNA-sequencing. Results: Low-dose IL-2 significantly improved the symptoms of DSS-induced colitis in mice and attenuated pro-inflammatory cytokine production and immune cell infiltration. The most effective dose range of IL-2 was 16K-32K IU/day. Importantly, low-dose IL-2 was effective in ameliorating the disruption of epithelial barrier integrity in DSS-induced colitis tissues by restoring tight junction proteins and mucin production and suppressing apoptosis. The colon tissue of DSS-induced mice exposed to low-dose IL-2 mimic gene expression patterns in the colons of control mice. Furthermore, we identified the crucial role of the PI3K-AKT pathway in exerting the therapeutic effect of low-dose IL-2. Conclusions: The results of our study suggest that low-dose IL-2 has therapeutic effects on DSS-induced colitis and potential clinical value in treating UC.

Imaging chromatin organization at the molecular-scale resolution remains an important endeavor in basic and translational research. Stochastic optical reconstruction microscopy (STORM) is a powerful superresolution imaging technique to visualize nanoscale molecular organization down to the resolution of ~20 to 30 nm. Despite the substantial progress in imaging chromatin organization in cells and model systems, its routine application on assessing pathological tissue remains limited. It is, in part, hampered by the lack of simple labels that consistently generates high-quality STORM images on the highly processed clinical tissue. We developed a fast, simple, and robust small-molecule fluorescent probe—cyanine 5–conjugated Hoechst—for routine superresolution imaging of nanoscale nuclear architecture on clinical tissue. We demonstrated the biological and clinical significance of imaging superresolved chromatin structure in cancer development and its potential clinical utility for cancer risk stratification.