Molecular classification reveals the sensitivity of lung adenocarcinoma to radiotherapy and immunotherapy: multi-omics clustering based on similarity network fusion.

BACKGROUND: Due to individual differences in tumors and immune systems, the response rate to immunotherapy is low in lung adenocarcinoma (LUAD) patients. Combinations with other therapeutic strategies improve the efficacy of immunotherapy in LUAD patients. Although radioimmunotherapy has been demonstrated to effectively suppress tumors, the underlying mechanisms still need to be investigated. METHODS: Total RNA from LUAD cells was sequenced before and after radiotherapy to identify differentially expressed radiation-associated genes. The similarity network fusion (SNF) algorithm was applied for molecular classification based on radiation-related genes, immune-related genes, methylation data, and somatic mutation data. The changes in gene expression, prognosis, immune cell infiltration, radiosensitivity, chemosensitivity, and sensitivity to immunotherapy were assessed for each subtype. RESULTS: We used the SNF algorithm and multi-omics data to divide TCGA-LUAD patients into three subtypes. Patients with the CS3 subtype had the best prognosis, while those with the CS1 and CS2 subtypes had poorer prognoses. Among the strains tested, CS2 exhibited the most elevated immune cell infiltration and expression of immune checkpoint genes, while CS1 exhibited the least. Patients in the CS2 subgroup were more likely to respond to PD-1 immunotherapy. The CS2 patients were most sensitive to docetaxel and cisplatin, while the CS1 patients were most sensitive to paclitaxel. Experimental validation of signature genes in the CS2 subtype showed that inhibiting the expression of RHCG and TRPA1 could enhance the sensitivity of lung cancer cells to radiation. CONCLUSIONS: In summary, this study identified a risk classifier based on multi-omics data that can guide treatment selection for LUAD patients.

Exposure to polycyclic aromatic hydrocarbons, volatile organic compounds, and tobacco-specific nitrosamines and incidence of esophageal cancer.

BACKGROUND: Studying carcinogens in tobacco and nontobacco sources may be key to understanding the pathogenesis and geographic distribution of esophageal cancer. METHODS: The Golestan Cohort Study has been conducted since 2004 in a region with high rates of esophageal squamous cell carcinoma. For this nested study, the cases comprised of all incident cases by January 1, 2018; controls were matched to the case by age, sex, residence, time in cohort, and tobacco use. We measured urinary concentrations of 33 exposure biomarkers of nicotine, polycyclic aromatic hydrocarbons, volatile organic compounds, and tobacco-specific nitrosamines. We used conditional logistic regression to calculate odds ratios (ORs) and 95% confidence intervals for associations between the 90th vs the 10th percentiles of the biomarker concentrations and incident esophageal squamous cell carcinoma. RESULTS: Among individuals who did not currently use tobacco (148 cases and 163 controls), 2 acrolein metabolites, 2 acrylonitrile metabolites, 1 propylene oxide metabolite, and one 1,3-butadiene metabolite were significantly associated with incident esophageal squamous cell carcinoma (adjusted odds ratios between 1.8 and 4.3). Among tobacco users (57 cases and 63 controls), metabolites of 2 other volatile organic compounds (styrene and xylene) were associated with esophageal squamous cell carcinoma (OR = 6.2 and 9.0, respectively). In tobacco users, 2 tobacco-specific nitrosamines (NNN and N'-Nitrosoanatabine) were also associated with esophageal squamous cell carcinoma. Suggestive associations were seen with some polycyclic aromatic hydrocarbons (especially 2-hydroxynaphthalene) in nonusers of tobacco products and other tobacco-specific nitrosamines in tobacco users. CONCLUSION: These novel associations based on individual-level data and samples collected many years before cancer diagnosis, from a population without occupational exposure, have important public health implications.

High-performance liquid chromatography-tandem mass spectrometry method for measuring cotinine and trans-3'-hydroxycotinine in bronchoalveolar lavage fluid of patients with e-cigarette, or vaping, product use-associated lung injury.

In 2019, nearly 3000 U.S. residents developed severe lung injury associated with recent use of e-cigarette or vaping products. The Centers for Disease Control and Prevention responded to the outbreak, which was formally defined as e-cigarette, or vaping, product use-associated lung injury (EVALI). Centers for Disease Control and Prevention Laboratory rapidly developed assays to analyze potentially harmful and addictive substances in bronchoalveolar lavage (BAL) fluid collected from EVALI case patients. This report describes the development and validation of a high-throughput isotope-dilution high performance liquid chromatography-tandem mass spectrometry method for measuring two nicotine biomarkers, cotinine (COT) and trans-3'-hydroxycotinine (HCT), in bronchoalveolar lavage fluid samples. COT and HCT are the major metabolites of nicotine, the addictive alkaloid presents in tobacco products. This method had good specificity and sensitivity. The limit of detection is 0.033 and 0.0165 ng/mL for COT and HCT, respectively, using only 200 µL of sample volume. The within-run and between-run precision were 2-10%. The overall accuracy, calculated from recovery in three different sample matrices spiked at three concentrations, was 94.8% and 93.6% for COT and HCT, respectively. This novel HPLC-MS-MS method was utilized to characterize recent tobacco exposure in EVALI case patients. This method is useful for characterizing tobacco exposure that may be related to acute and chronic lung injury.

Influence of Half-life and Smoking/Nonsmoking Ratio on Biomarker Consistency between Waves 1 and 2 of the Population Assessment of Tobacco and Health Study.

BACKGROUND: Biomarkers of exposure are tools for understanding the impact of tobacco use on health outcomes if confounders like demographics, use behavior, biological half-life, and other sources of exposure are accounted for in the analysis. METHODS: We performed multiple regression analysis of longitudinal measures of urinary biomarkers of alkaloids, tobacco-specific nitrosamines, polycyclic aromatic hydrocarbons, volatile organic compounds (VOC), and metals to examine the sample-to-sample consistency in Waves 1 and 2 of the Population Assessment of Tobacco and Health (PATH) Study including demographic characteristics and use behavior variables of persons who smoked exclusively. Regression coefficients, within- and between-person variance, and intra-class correlation coefficients (ICC) were compared with biomarker smoking/nonsmoking population mean ratios and biological half-lives. RESULTS: Most biomarkers were similarly associated with sex, age, race/ethnicity, and product use behavior. The biomarkers with larger smoking/nonsmoking population mean ratios had greater regression coefficients related to recency of exposure. For VOC and alkaloid metabolites, longer biological half-life was associated with lower within-person variance. For each chemical class studied, there were biomarkers that demonstrated good ICCs. CONCLUSIONS: For most of the biomarkers of exposure reported in the PATH Study, for people who smoke cigarettes exclusively, associations are similar between urinary biomarkers of exposure and demographic and use behavior covariates. Biomarkers of exposure within-subject consistency is likely associated with nontobacco sources of exposure and biological half-life. IMPACT: Biomarkers measured in the PATH Study provide consistent sample-to-sample measures from which to investigate the association of adverse health outcomes with the characteristics of cigarettes and their use.

Development of stimuli responsive polymeric nanomedicines modulating tumor microenvironment for improved cancer therapy.

The complexity of the tumor microenvironment (TME) severely hinders the therapeutic effects of various cancer treatment modalities. The TME differs from normal tissues owing to the presence of hypoxia, low pH, and immune-suppressive characteristics. Modulation of the TME to reverse tumor growth equilibrium is considered an effective way to treat tumors. Recently, polymeric nanomedicines have been widely used in cancer therapy, because their synthesis can be controlled and they are highly modifiable, and have demonstrated great potential to remodel the TME. In this review, we outline the application of various stimuli responsive polymeric nanomedicines to modulate the TME, aiming to provide insights for the design of the next generation of polymeric nanomedicines and promote the development of polymeric nanomedicines for cancer therapy.

Validating Wave 1 (2014) Urinary Cotinine and TNE-2 Cut-points for Differentiating Wave 4 (2017) Cigarette Use from Non-use in the United States Using Data from the PATH Study.

BACKGROUND: Sex and racial/ethnic identity-specific cut-points for validating tobacco use using Wave 1 (W1) of the Population Assessment of Tobacco and Health (PATH) Study were published in 2020. The current study establishes predictive validity of the W1 (2014) urinary cotinine and total nicotine equivalents-2 (TNE-2) cut-points on estimating Wave 4 (W4; 2017) tobacco use. METHODS: For exclusive and polytobacco cigarette use, weighted prevalence estimates based on W4 self-report alone and with exceeding the W1 cut-point were calculated to identify the percentage missed without biochemical verification. Sensitivity and specificity of W1 cut-points on W4 self-reported tobacco use status were examined. ROC curves were used to determine the optimal W4 cut-points to distinguish past 30-day users from non-users, and evaluate whether the cut-points significantly differed from W1. RESULTS: Agreement between W4 self-reported use and exceeding the W1 cut-points was high overall and when stratified by demographic subgroups (0.7%-4.4% of use was missed if relying on self-report alone). The predictive validity of using the W1 cut-points to classify exclusive cigarette and polytobacco cigarette use at W4 was high (>90% sensitivity and specificity, except among polytobacco Hispanic smokers). Cut-points derived using W4 data did not significantly differ from the W1-derived cut-points [e.g., W1 exclusive = 40.5 ng/mL cotinine (95% confidence interval, CI: 26.1-62.8), W4 exclusive = 29.9 ng/mL cotinine (95% CI: 13.5-66.4)], among most demographic subgroups. CONCLUSIONS: The W1 cut-points remain valid for biochemical verification of self-reported tobacco use in W4. IMPACT: Findings from can be used in clinical and epidemiologic studies to reduce misclassification of cigarette smoking status.

Tobacco Smoke Is a Major Source of Aromatic Amine Exposure in U.S. Adults: 2013-2014 National Health and Nutrition Examination Survey (NHANES).

BACKGROUND: Cigarette smoking increases the risk of cancer, cardiovascular diseases, and premature death. Aromatic amines (AA) are found in cigarette smoke and are well-established human bladder carcinogens. METHODS: We measured and compared total urinary levels of 1-aminonaphthalene (1AMN), 2-aminonaphthalene (2AMN), and 4-aminobiphenyl (4ABP) in adults who smoked cigarettes exclusively and in adult nonusers of tobacco products from a nationally representative sample of non-institutionalized U.S. population in the 2013-2014 National Health and Nutrition Examination Survey. RESULTS: Sample-weighted geometric mean concentrations of AAs in adults who smoked cigarettes exclusively compared with adult nonusers were 30 times higher for 1AMN and 4 to 6 times higher for 2AMN and 4ABP. We evaluated the association of tobacco-smoke exposure with urinary AAs using sample-weighted multiple linear regression models to control for age, sex, race/ethnicity, diet, and urinary creatinine. Secondhand smoke exposure status was categorized using serum cotinine (SCOT) among adult nonusers (SCOT ≤ 10 ng/mL). The exposure for adults who smoked cigarettes exclusively (SCOT > 10 ng/mL) was categorized on the basis of the average number of self-reported cigarettes smoked per day (CPD) in the five days prior to urine collection. The regression models show AAs concentration increased with increasing CPD (P < 0.001). Dietary-intake variables derived from the 24-hours recall questionnaire were not consistently significant predictors of urinary AAs. CONCLUSIONS: This is the first characterized total urinary AA concentrations of the U.S. adult non-institutionalized population. Our analyses show that smoking status is a major contributor to AA exposures. IMPACT: These data provide a crucial baseline for exposure to three AAs in U.S. non-institutionalized adults.

Short- and Long-Term Stability of Aromatic Amines in Human Urine.

Several aromatic amines (AAs) are established by the International Agency for Research on Cancer as carcinogenic (group 1) or probable/possible carcinogens to humans (group 2A/2B). AAs can be found in mainstream and sidestream smoke from combustible tobacco products, as well as in certain environmental pollution and occupational exposure from several chemical industry sectors. Exposure to AAs can be estimated by measuring their concentrations in urine; however, information about the short-term and long-term stabilities of AAs in urine need to be characterized before conducting large-scale population studies on AA exposure and the potentially harmful effects of AA exposure. In this report, the storage stability of o-toluidine, 2,6-dimethylaniline, o-anisidine, 1-aminonaphthalene, 2-aminonaphthalene, and 4-aminobiphenyl fortified in pooled, filtered, non-smokers' urine is analyzed by isotope dilution gas chromatography-triple quadrupole mass spectrometry (ID GC-MS/MS). The six AAs were measured in urine samples stored at ~20 °C (collection temperature), 4 °C and 10 °C (short-term transit temperatures), and -20 °C and -70 °C (long-term storage temperatures) over a 10-day period. All six analytes were stable for 10 days at transit and long-term storage temperatures but showed reduced recovery at 20 °C. The instability of the target AAs at 20 °C suggests that immediate storage of freshly voided urine at low temperatures is needed to attenuate degradation. A subset of the urine samples was analyzed following a longer storage duration at -70 °C: all AAs were stable for up to 14 months at this temperature. The stability of the six AAs in urine samples can be maintained at the various temperature levels and storage times expected in a typical study set.

CEMIP, acting as a scaffold protein for bridging GRAF1 and MIB1, promotes colorectal cancer metastasis via activating CDC42/MAPK pathway.

Metastasis is the leading cause of treatment failure and tumor-related death in colorectal cancer (CRC). Our previous studies report that CEMIP functionally promotes CRC metastasis and is closely related to poor outcomes. However, the molecular network of CEMIP promoting CRC metastasis is still not fully understood. In the current study, we identify CEMIP interacting with GRAF1, and the combination of high-CEMIP and low-GRAF1 predicts poor survival of patients. Mechanistically, we elucidate that CEMIP interacts with the SH3 domain of GRAF1 through the 295-819aa domain, and negatively regulates the stability of GRAF1. Moreover, we identify MIB1 to be an E3 ubiquitin ligase for GRAF1. Importantly, we uncover that CEMIP acts as a scaffold protein in bridging MIB1 and GRAF1, which is critical to GRAF1 degradation and CEMIP-mediated CRC metastasis. Furthermore, we found that CEMIP activates CDC42/MAPK pathway-regulated EMT by enhancing the degradation of GRAF1, which is indispensable to CEMIP-mediated migration and invasion of CRC cells. Subsequently, we prove that CDC42 inhibitor suppresses CEMIP-mediated CRC metastasis in vitro and in vivo. Collectively, our results reveal that CEMIP promotes CRC metastasis through GRAF1/CDC42/MAPK pathway-regulated EMT, and suggest that CDC42 inhibitor could be a novel therapeutic strategy for CEMIP-mediated CRC metastasis.

Tumor CEMIP drives immune evasion of colorectal cancer via MHC-I internalization and degradation.

BACKGROUND: Loss of major histocompatibility complex class I (MHC-I) in tumor cells limits the use of immune checkpoint blockade (ICB) in colorectal cancer. Nevertheless, the regulatory mechanism of MHC-I downregulation in tumor cells has not been fully elucidated. Overexpression of CEMIP in tumor tissues is associated with a poor prognosis in colorectal cancer. Here, in this research, we aim to address the role of CEMIP in mediating MHC-I expression in tumor cells and investigate the underlying regulatory mechanisms. METHOD: Protein levels were analyzed by western blotting. Flow cytometry analysis was used to examine immune cells. Protein-protein interactions were investigated by co-immunoprecipitation and proximity ligation assays. The intracellular trafficking of MHC-I was revealed by an immunofluorescent technique. In addition, the effect of CEMIP on tumor growth and the antitumor efficacy of targeting CEMIP in combination with ICB therapy were evaluated in murine models of colorectal cancer. RESULTS: We reported that CEMIP specifically downregulated the expression of MHC-I on the surface of murine and human colon cancer cells, hindering the cytotoxicity of CD8+ T cells. We also demonstrated that CEMIP restricted CD8+ T-cell antitumor activities both in vitro and in vivo due to impaired MHC-I-mediated antigen presentation. Correspondingly, the combination of CEMIP inhibition and ICB impeded tumor growth and enhanced therapeutic efficacy. Mechanistically, CEMIP acted as an adaptor for the interaction betweenMHC-I and clathrin, which drove MHC-I internalization via clathrin-dependent endocytosis. Furthermore, CEMIP anchored internalized MHC-I to lysosomes for degradation, disrupting the recycling of MHC-I to the cell surface. CONCLUSION: Overall, our study unveils a novel regulatory mechanism of MHC-I on tumor cell surfaces by CEMIP-mediated internalization and degradation. Furthermore, targeting CEMIP provides an effective strategy for colorectal cancer immunotherapy.

Distinguishing Exposure to Secondhand and Thirdhand Tobacco Smoke among U.S. Children Using Machine Learning: NHANES 2013-2016.

While the thirdhand smoke (THS) residue from tobacco smoke has been recognized as a distinct public health hazard, there are currently no gold standard biomarkers to differentiate THS from secondhand smoke (SHS) exposure. This study used machine learning algorithms to assess which combinations of biomarkers and reported tobacco smoke exposure measures best differentiate children into three groups: no/minimal tobacco smoke exposure (NEG); predominant THS exposure (TEG); and mixed SHS and THS exposure (MEG). Participants were 4485 nonsmoking 3-17-year-olds from the National Health and Nutrition Examination Survey 2013-2016. We fitted and tested random forest models, and the majority (76%) of children were classified in NEG, 16% were classified in TEG, and 8% were classified in MEG. The final classification model based on reported exposure, biomarker, and biomarker ratio variables had a prediction accuracy of 95%. This final model had prediction accuracies of 100% for NEG, 88% for TEG, followed by 71% for MEG. The most important predictors were the reported number of household smokers, serum cotinine, serum hydroxycotinine, and urinary 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol (NNAL). In the absence of validated biomarkers specific to THS, comprehensive biomarker and questionnaire data for tobacco smoke exposure can distinguish children exposed to SHS and THS with high accuracy.

Variability in Urinary Nicotine Exposure Biomarker Levels Between Waves 1 (2013-2014) and 2 (2014-2015) in the Population Assessment of Tobacco and Health Study.

INTRODUCTION: To date, no studies have evaluated the consistency of biomarker levels in people who smoke over a long-time period in real-world conditions with a large number of subjects and included use behavior and measures of nicotine metabolism. We evaluated the variability of biomarkers of nicotine exposure over approximately a 1-year period in people who exclusively smoke cigarettes, including intensity and recency of use and brand switching to assess impact on understanding associations with product characteristics. AIMS AND METHODS: Multivariate regression analysis of longitudinal repeated measures of urinary biomarkers of nicotine exposure from 916 adults in the Population Assessment of Tobacco and Health (PATH) Study with demographic characteristics and use behavior variables. Intraclass correlation coefficients (ICCs) were calculated to examine individual variation of nicotine biomarkers and the uncertainty of repeat measures at two time points (Waves 1 and 2). RESULTS: Age, race, and urinary creatinine were significant covariates of urinary cotinine. When including use behavior, recency, and intensity of use were highly significant and variance decreased to a higher extent between than within subjects. The ICC for urinary cotinine decreased from 0.7530 with no use behavior variables in the model to 0.5763 when included. Similar results were found for total nicotine equivalents. CONCLUSIONS: Urinary nicotine biomarkers in the PATH Study showed good consistency between Waves 1 and 2. Use behavior measures such as time since last smoked a cigarette and number of cigarettes smoked in the past 30 days are important to include when assessing factors that may influence biomarker concentrations. IMPLICATIONS: The results of this study show that the consistency of the nicotine biomarkers cotinine and total nicotine equivalents in spot urine samples from Waves 1 to 2 of the PATH Study is high enough that these data are useful to evaluate the association of cigarette characteristics with biomarkers of exposure under real-world use conditions.

Manipulation of Menthol and Nicotine Content in Cigarettes: Effects on Smoking Behavior and Toxicant Exposure in Women Menthol Smokers.

BACKGROUND: The purpose of this study was to determine the effects of smoking and other outcomes of assigning cigarettes with reduced nicotine and/or no menthol to female menthol smokers. AIMS AND METHODS: Nontreatment-seeking female menthol smokers (N = 263) participated in a randomized controlled trial in which levels of menthol and nicotine in cigarettes were manipulated using experimental cigarettes. After a baseline period, participants were assigned to the following conditions for 6 weeks: (1) their own brand of cigarette (conventional nicotine with menthol), (2) a conventional nicotine cigarette with no menthol, (3) a cigarette with reduced nicotine (RNC) with menthol, or (4) a RNC cigarette and no menthol. Participants then returned to using their own brand and were followed for another 6 weeks. Outcomes included cigarettes smoked, biomarkers of exposure, and dependence measures. RESULTS: Results indicated that, after an initial increase, rates of smoking of all three experimental cigarettes were at or below baseline rates of smoking of one's own brand. Levels of biomarkers also decreased during the experimental phase but rebounded somewhat after participants resumed smoking their own brand. There was evidence that the overall amount of smoking decreased similarly among women who switched to non-menthol and/or RNC cigarettes. CONCLUSIONS: These results suggest that no detrimental effect will occur in nicotine or toxicant exposure levels with a ban on characterizing menthol and/or a product standard on nicotine content in cigarettes. IMPLICATIONS: The implication of this work is that there would be no risk to women menthol smokers associated with regulations restricting nicotine and eliminating menthol in cigarettes.

Anabasine and Anatabine Exposure Attributable to Cigarette Smoking: National Health and Nutrition Examination Survey (NHANES) 2013-2014.

Anabasine and anatabine are minor alkaloids in tobacco products and are precursors for tobacco-specific nitrosamines (TSNAs). The levels of these two compounds have been used to differentiate tobacco product sources, monitor compliance with smoking cessation programs, and for biomonitoring in TSNA-related studies. The concentrations of urinary anabasine and anatabine were measured in a representative sample of U.S. adults who smoked cigarettes (N = 770) during the 2013−2014 National Health and Nutrition Examination Survey (NHANES) study cycle, which was the first cycle where urinary anabasine and anatabine data became available. Weighted geometric means (GM) and geometric least squares means (LSM) with 95% confidence intervals were calculated for urinary anabasine and anatabine categorized by tobacco-use status [cigarettes per day (CPD) and smoking frequency] and demographic characteristics. Smoking ≥20 CPD was associated with 3.6× higher anabasine GM and 4.8× higher anatabine GM compared with smoking <10 CPD. Compared with non-daily smoking, daily smoking was associated with higher GMs for urinary anabasine (1.41 ng/mL vs. 6.28 ng/mL) and anatabine (1.62 ng/mL vs. 9.24 ng/mL). Urinary anabasine and anatabine concentrations exceeded the 2 ng/mL cut point in 86% and 91% of urine samples from people who smoke (PWS) daily, respectively; in comparison, 100% of them had serum cotinine concentrations greater than the established 10 ng/mL cut point. We compared these minor tobacco alkaloid levels to those of serum cotinine to assess their suitability as indicators of recent tobacco use at established cut points and found that their optimal cut point values would be lower than the established values. This is the first time that anabasine and anatabine are reported for urine collected from a U.S. population-representative sample of NHANES study participants, providing a snapshot of exposure levels for adults who smoked during 2013−2014. The results of this study serve as an initial reference point for future analysis of NHANES cycles, where changes in the national level of urinary anabasine and anatabine can be monitored among people who smoke to show the effect of changes in tobacco policy.

Occurrence of methylmercury in aerobic environments: Evidence of mercury bacterial methylation based on simulation experiments.

Methylmercury (MeHg) is mainly produced by anaerobic δ-proteobacteria such as sulfate-reducing bacteria (SRB). However, mercury bio-methylation has also been found to occur in the aerobic soil of the Three Gorges Reservoir (TGR). Using γ-proteobacterial TGR bacteria (TGRB) and δ-proteobacterial Desulfomicrobium escambiense strains, the efficiency of mercury methylation and demethylation was evaluated using an isotope tracer technique. Kinetics simulation showed that the bacterial Hg methylation rate (km) of TGRB3 was 4.36 × 10-9 pg·cell-1·h-1, which was significantly lower than that of D. escambiense (170.74 ×10-9 pg·cell-1·h-1) under anaerobic conditions. Under facultative and/or aerobic conditions, D. escambiense could not survive, while the km of TGRB3 were 0.35 × 10-9 and 0.29 × 10-9 pg·cell-1·h-1, respectively. Furthermore, the bacterial MeHg tolerance threshold of TGRB3 was 3.47 × 10-9 pg·cell-1, which was 98.6-fold lower than that of D. escambiense under anaerobic conditions. However, the MeHg tolerance threshold of TGRB3 remained at 0.50-0.52 × 10-9 pg·cell-1 under facultative and/or aerobic conditions. Notably, bacterial Hg methylation rates (km) were higher than the corresponding bacterial MeHg demethylation rates (kd1). These results establish the contribution of some aerobic and/or facultative anaerobic bacteria to net environmental MeHg production in terrestrial ecosystems and provide a novel understanding of the biogeochemical cycle of MeHg. SYNOPSIS: Hg methylation of facultative and/or aerobic bacteria may contribute to the net production of environmental methylmercury in terrestrial ecosystems.

Geometric Mean Serum Cotinine Concentrations Confirm a Continued Decline in Secondhand Smoke Exposure among U.S. Nonsmokers-NHANES 2003 to 2018.

The objective of this study was to examine long-term trends in serum cotinine (COT) concentrations, as a measure of secondhand smoke (SHS) exposure, in U.S. nonsmokers using data from the National Health and Nutrition Examination Surveys (NHANES) from 2003 to 2018. We analyzed NHANES serum COT results from 8 continuous NHANES 2 year cycles from 2003 to 2018 using a liquid chromatography−tandem mass spectrometry assay that has been maintained continuously at the Centers for Disease Control and Prevention (CDC) since 1992. Serum COT concentrations (based on the geometric means) among nonsmokers in the U.S. decreased by an average of 11.0% (95% confidence interval (CI) [8.8%, 13.1%]; p < 0.0001) every 2 year cycle. From 2003 to 2018, serum COT concentrations in U.S. nonsmokers declined by 55.0%, from 0.065 ng/mL in 2003−2004 to 0.029 ng/mL in 2017−2018 (p < 0.0001). Significant decreases in serum COT concentrations were observed in all demographic groups. While disparities between these groups seems to be shrinking over time, several previously observed disparities in SHS exposure remain in 2017−2018. Serum COT concentrations of the non-Hispanic Black population remained higher than those of non-Hispanic Whites and Mexican Americans (p < 0.0001). Additionally, serum COT concentrations were significantly higher for children aged 3−5 years than other age groups (p ≤ 0.0002), and men continued to have significantly higher serum COT concentrations than women (p = 0.0384). While there is no safe level of exposure to SHS, the decrease in serum COT concentrations in the U.S. population as well as across demographic groupings represents a positive public health outcome and supports the importance of comprehensive smoke-free laws and policies for workplaces, public places, homes, and vehicles to protect nonsmokers from SHS exposure.

Targeting DCLK1 overcomes 5-fluorouracil resistance in colorectal cancer through inhibiting CCAR1/ß-catenin pathway-mediated cancer stemness.

BACKGROUND: To date, 5-fluorouracil-based chemotherapy is very important for locally advanced or metastatic colorectal cancer (CRC). However, chemotherapy resistance results in tumor recurrence and metastasis, which is a major obstacle for treatment of CRC. METHODS: In the current research, we establish 5-fluorouracil resistant cell lines and explore the potential targets associated with 5-fluorouracil resistance in CRC. Moreover, we perform clinical specimen research, in vitro and in vivo experiments and molecular mechanism research, to reveal the biological effects and the mechanism of DCLK1 promoting 5-fluorouracil resistance, and to clarify the potential clinical value of DCLK1 as a target of 5-fluorouracil resistance in CRC. RESULTS: We discover that doublecortin-like kinase 1 (DCLK1), a cancer stem cell maker, is correlated with 5-fluorouracil resistance, and functionally promotes cancer stemness and 5-fluorouracil resistance in CRC. Mechanistically, we elucidate that DCLK1 interacts with cell cycle and apoptosis regulator 1 (CCAR1) through the C-terminal domain, and phosphorylates CCAR1 at the Ser343 site, which is essential for CCAR1 stabilisation. Moreover, we find that DCLK1 positively regulates ß-catenin signalling via CCAR1, which is responsible for maintaining cancer stemness. Subsequently, we prove that blocking ß-catenin inhibits DCLK1-mediated 5-fluorouracil resistance in CRC cells. Importantly, we demonstrate that DCLK1 inhibitor could block CCAR1/ß-catenin pathway-mediated cancer stemness and consequently suppresses 5-fluorouracil resistant CRC cells in vitro and in vivo. CONCLUSIONS: Collectively, our findings reveal that DCLK1 promotes 5-fluorouracil resistance in CRC by CCAR1/ß-catenin pathway-mediated cancer stemness, and suggest that targeting DCLK1 might be a promising method to eliminate cancer stem cells for overcoming 5-fluorouracil resistance in CRC.

Urinary Nicotine Metabolites and Self-Reported Tobacco Use Among Adults in the Population Assessment of Tobacco and Health (PATH) Study, 2013-2014.

INTRODUCTION: The Population Assessment of Tobacco and Health (PATH) Study is a longitudinal cohort study on tobacco use behavior, attitudes and beliefs, and tobacco-related health outcomes, including biomarkers of tobacco exposure in the U.S. population. In this report we provide a summary of urinary nicotine metabolite measurements among adult users and non-users of tobacco from Wave 1 (2013-2014) of the PATH Study. METHODS: Total nicotine and its metabolites including cotinine, trans-3'-hydroxycotinine (HCTT), and other minor metabolites were measured in more than 11 500 adult participants by liquid chromatography tandem mass spectrometry methods. Weighted geometric means (GM) and least square means from statistical modeling were calculated for non-users and users of various tobacco products. RESULTS: Among daily users, the highest GM concentrations of nicotine, cotinine and HCTT were found in exclusive smokeless tobacco users, and the lowest in exclusive e-cigarette users. Exclusive combustible product users had intermediate concentrations, similar to those found in users of multiple products (polyusers). Concentrations increased with age within the categories of tobacco users, and differences associated with gender, race/ethnicity and educational attainment were also noted among user categories. Recent (past 12 months) former users had GM cotinine concentrations that were more than threefold greater than never users. CONCLUSIONS: These urinary nicotine metabolite data provide quantification of nicotine exposure representative of the entire US adult population during 2013-2014 and may serve as a reference for similar analyses in future measurements within this study. IMPLICATIONS: Nicotine and its metabolites in urine provide perhaps the most fundamental biomarkers of recent nicotine exposure. This report, based on Wave 1 of the Population Assessment of Tobacco and Health (PATH) Study, provides the first nationally representative data describing urinary nicotine biomarker concentrations in both non-users, and users of a variety of tobacco products including combustible, e-cigarette and smokeless products. These data provide a urinary biomarker concentration snapshot in time for the entire US population during 2013-2014, and will provide a basis for comparison with future results from continuing, periodic evaluations in the PATH Study.

Nicotine Exposure in the U.S. Population: Total Urinary Nicotine Biomarkers in NHANES 2015-2016.

We characterize nicotine exposure in the U.S. population by measuring urinary nicotine and its major (cotinine, trans-3'-hydroxycotinine) and minor (nicotine 1'-oxide, cotinine N-oxide, and 1-(3-pyridyl)-1-butanol-4-carboxylic acid, nornicotine) metabolites in participants from the 2015−2016 National Health and Nutrition Examination Survey. This is one of the first U.S. population-based urinary nicotine biomarker reports using the derived total nicotine equivalents (i.e., TNEs) to characterize exposure. Serum cotinine data is used to stratify tobacco non-users with no detectable serum cotinine (−sCOT), non-users with detectable serum cotinine (+sCOT), and individuals who use tobacco (users). The molar concentration sum of cotinine and trans-3'-hydroxycotinine was calculated to derive the TNE2 for non-users. Additionally, for users, the molar concentration sum of nicotine and TNE2 was calculated to derive the TNE3, and the molar concentration sum of the minor metabolites and TNE3 was calculated to derive the TNE7. Sample-weighted summary statistics are reported. We also generated multiple linear regression models to analyze the association between biomarker concentrations and tobacco use status, after adjusting for select demographic factors. We found TNE7 is positively correlated with TNE3 and TNE2 (r = 0.99 and 0.98, respectively), and TNE3 is positively correlated with TNE2 (r = 0.98). The mean TNE2 concentration was elevated for the +sCOT compared with the −sCOT group (0.0143 [0.0120, 0.0172] µmol/g creatinine and 0.00188 [0.00172, 0.00205] µmol/g creatinine, respectively), and highest among users (33.5 [29.6, 37.9] µmol/g creatinine). Non-daily tobacco use was associated with 50% lower TNE7 concentrations (p < 0.0001) compared with daily use. In this report, we show tobacco use frequency and passive exposure to nicotine are important sources of nicotine exposure. Furthermore, this report provides more information on non-users than a serum biomarker report, which underscores the value of urinary nicotine biomarkers in extending the range of trace-level exposures that can be characterized.

KIAA1199 drives immune suppression to promote colorectal cancer liver metastasis by modulating neutrophil infiltration.

BACKGROUND AND AIMS: Metastasis is the primary cause of cancer mortality, and colorectal cancer (CRC) frequently metastasizes to the liver. Our previous studies demonstrated the critical role of KIAA1199 in tumor invasion and metastasis in CRC. In the present study, we described an immune regulatory effect of KIAA1199 that creates a permissive environment for metastasis. APPROACH AND RESULTS: Flow cytometry was used to examine the effects of KIAA1199 on the infiltration of tumor immune cells. Neutrophils and T cells were isolated, stimulated, and/or cultured for in vitro function assays. In the patients with CRC, high expression levels of KIAA1199 were associated with an increased neutrophil infiltration into the liver. This result was further validated in mouse metastasis models. The increased influx of neutrophils contributed to the KIAA1199-driven CRC liver metastasis. Mechanistically, KIAA1199 activated the TGFß signaling pathway by interacting with the TGFBR1/2 to stimulate CXCL1 and CXCL3 production, thereby driving the aggregation of immunosuppressive neutrophils. Genetic blockade or pharmacologic inhibition of KIAA1199 restored tumor immune infiltration, impeded tumor progression, and potentiated response to immune checkpoint blockade (ICB). CONCLUSIONS: These findings indicated that KIAA1199 could facilitate the liver infiltration of immunosuppressive neutrophils via the TGFß-chemokine (C-X-C motif) ligand (CXCL)3/1-CXCR2 axis, which might be clinically targeted for the treatment of hepatic metastasis.