Major Carcinogen Research Articles

Highly sensitive fluorescent bioassay of 2,3,7,8-tetrachloro-dibenzo-p-dioxin based on abnormal expression of cytochrome P450 1A2 in human cells

Current in vitro bioassays of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD, a major threat carcinogen) are relied on murine cells and fluorescent probe 7-ethoxyresorufin (7-ER), in which TCDD mostly causes abnormal expression of cytochrome P450 1A1 (CYP1A1). However, for human cells, TCDD mainly leads to a distinct abnormal expression of cytochrome P450 1A2 (CYP1A2). The poor response of 7-ER to CYP1A2 limits the traditional bioassay for human cells. Herein, we report a fluorescent probe N-(3-hydroxybutyl)-4-methoxy-1,8-naphthalimide (HBMN) for in vitro bioassay of TCDD with human cells. HBMN had ca. 60 times higher affinity to CYP1A2 than 7-ER. As such, the sensing sensitivity increased by 10 times, and different expression of CYP1A2 by TCDD induction in different human cells was found. Besides, HBMN was also feasible in rapid screening of TCDD concentration by naked eye. It would open a new way to highly sensitive detect TCDD and understand the pathogenesis of TCDD in different human organs.

Hexavalent Chromium-Induced Chromosome Instability Drives Permanent and Heritable Numerical and Structural Changes and a DNA Repair-Deficient Phenotype.

A key hypothesis for how hexavalent chromium [Cr(VI)] causes cancer is that it drives chromosome instability (CIN), which leads to neoplastic transformation. Studies show chronic Cr(VI) can affect DNA repair and induce centrosome amplification, which can lead to structural and numerical CIN. However, no studies have considered whether these outcomes are transient or permanent. In this study, we exposed human lung cells to particulate Cr(VI) for three sequential 24-hour periods, each separated by about a month. After each treatment, cells were seeded at colony-forming density, cloned, expanded, and retreated, creating three generations of clonal cell lines. Each generation of clones was tested for chromium sensitivity, chromosome complement, DNA repair capacity, centrosome amplification, and the ability to grow in soft agar. After the first treatment, Cr(VI)-treated clones exhibited a normal chromosome complement, but some clones showed a repair-deficient phenotype and amplified centrosomes. After the second exposure, more than half of the treated clones acquired an abnormal karyotype including numerical and structural alterations, with many exhibiting deficient DNA double-strand break repair and amplified centrosomes. The third treatment produced new abnormal clones, with previously abnormal clones acquiring additional abnormalities and most clones exhibiting repair deficiency. CIN, repair deficiency, and amplified centrosomes were all permanent and heritable phenotypes of repeated Cr(VI) exposure. These outcomes support the hypothesis that CIN is a key mechanism of Cr(VI)-induced carcinogenesis.Significance: Chromium, a major public health concern and human lung carcinogen, causes fundamental changes in chromosomes and DNA repair in human lung cells. Cancer Res; 78(15); 4203-14. ©2018 AACR.

Benzo(a)pyrene in Cigarette Smoke Enhances HIV-1 Replication through NF-\u03baB Activation via CYP-Mediated Oxidative Stress Pathway

Smoking aggravates HIV-1 pathogenesis and leads to decreased responses to antiretroviral therapy. In this study, we aim to find a molecular mechanism that would explain smoking-induced HIV-1 replication. Benzo(a)pyrene (BaP), a major carcinogen in cigarette, requires metabolic activation through cytochrome P450s (CYPs) to exert its toxic effects. We hypothesized that CYP-mediated BaP metabolism generates reactive oxygen species (ROS), and the resultant oxidative stress aggravates HIV-1 replication. As expected, we observed ~3 to 4-fold increase in HIV-1 replication in U1 cells and human primary macrophages after chronic BaP exposure. We also observed ~30-fold increase in the expression of CYP1A1 at mRNA level, ~2.5-fold increase in its enzymatic activity as well as elevated ROS and cytotoxicity in U1 cells. The knock-down of the CYP1A1 gene using siRNA and treatment with selective CYP inhibitors and antioxidants significantly reduced HIV-1 replication. Further, we observed a nuclear translocation of NF-κB subunits (p50 and p65) after chronic BaP exposure, which was reduced by treatment with siRNA and antioxidants/CYP inhibitors. Suppression of NF-κB pathway using specific NF-κB inhibitors also significantly reduced HIV-1 replication. Altogether, our results suggest that BaP enhances HIV-1 replication in macrophages by a CYP-mediated oxidative stress pathway followed by the NF-κB pathway.

Abstract 536: Targeting PRPK and TOPK for skin cancer prevention and therapy

Abstract Solar ultraviolet (sUV) irradiation is a major environmental carcinogen that can cause inflammation and skin cancer. The costs and morbidity associated with skin cancer are increasing, thus identifying molecules that can help prevent skin carcinogenesis is important. In this study, we identified the p53-related protein kinase (PRPK) as a novel oncogenic protein that is phosphorylated by the T-LAK cell-originated protein kinase (TOPK). Knockdown of TOPK inhibited PRPK phosphorylation and conferred resistance to solar simulated light (SSL)-induced skin carcinogenesis in mouse models. In the clinic, acute SSL irradiation significantly increased epidermal thickness as well as total protein and phosphorylation levels of TOPK and PRPK in human skin tissues. We identified two PRPK inhibitors, FDA-approved rocuronium bromide (Zemuron®) or betamethasone 17-valerate (Betaderm®) that could attenuate TOPK-dependent PRPK signaling. Importantly, topical application of either rocuronium or betamethasone attenuated SSL-induced epidermal hyperplasia, neovascularization and cutaneous squamous cell carcinoma (cSCC) development in SKH1 (Crl: SKH1-Hrhr) hairless mice by inhibiting PRPK activation, and also reduced expression of the proliferation and oncogenesis markers COX-2, cyclin D1 and MMP-9. This study is the first to demonstrate that PRPK- targeted therapy could be useful against sUV-induced cSCC. Citation Format: Eunmiri Roh, Mee-Hyun Lee, Tatyana A. Zykova, Feng Zhu, Hong-Gyum Kim, Ki Beom Bae, Yong Yeon Cho, Ann M. Bode, Zigang Dong. Targeting PRPK and TOPK for skin cancer prevention and therapy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 536.

Abstract 5061: ALDH2 restricts esophageal stem/progenitor cell expansion via autophagy under alcohol-induced mitochondrial stress

Abstract Introduction: Esophageal cancer is linked to heavy alcohol (EtOH) drinking and polymorphic aldehyde dehydrogenase 2 (ALDH2) mutation. Mutant ALDH2E487K protein delays mitochondrial clearance of acetaldehyde, the chief metabolite of EtOH and a major human carcinogen. Autophagy maintains mitochondrial redox homeostasis and may act as a tumor suppressor by decreasing oxidative stress. The role of ALDH2 and autophagy in esophageal epithelial homeostasis remains elusive. Methods: We analyzed EtOH-exposed esophageal epithelial cells and EtOH-fed Aldh2E487 or Aldh2WT mice by immunoblotting, immunofluorescence, flow cytometry and respirometry to determine mTORC1 signaling, autophagy, mitochondrial dysfunction and superoxide production. Mitochondrial superoxide was documented via mitochondria-targeted antioxidant mito-CP. Autophagy flux was blocked pharmacologically by chloroquine (CQ) or RNA interference (RNAi) against Beclin, a mediator of autophagy. Mitochondria-targeted autophagy (mitophagy) was further evaluated by RNAi against Parkin. To evaluate esophageal stem/progenitor cell function, we assessed the formation of single cell-derived three-dimensional (3D) organoids from mice with Aldh2E487K or Atg7loxP/loxP (autophagy mediator). We determined organoid formation following EtOH exposure, autophagy inhibition by CQ or adenoviral Cre-mediated Atg7 deletion. Rapamycin was used to inhibit mTORC1 and activate autophagy. Results: EtOH induced mitochondria damage in esophageal epithelial cells in vitro and in vivo with enhanced mitochondrial dysfunction by Aldh2E487K. EtOH induced mitochondrial dysfunction and superoxide production was antagonized by mito-CP. EtOH robustly suppressed mTORC1 signaling with concurrent activation of autophagy flux. Pharmacological or genetic inhibition of autophagy or mitophagy aggravated EtOH-induced mitochondrial dysfunction, superoxide production and cell death. Recapitulating esophageal epithelial proliferation-differentiation gradient ex vivo, Aldh2E487K murine esophageal cells displayed increased 3D organoid formation capability where resulting organoids contained more basaloid (undifferentiated) cells with mitochondrial dysfunction as compared to Aldh2WT cells. Moreover, EtOH stimulated 3D organoid formation by Aldh2E487K cells in a dose-dependent manner. Aldh2 dysfunction was phenocopied by CQ or Atg7 deletion, which increased 3D organoid formation. Interestingly, rapamycin suppressed organoid formation. Conclusions: Our innovative approach reveals that autophagy and ALDH2 protect against EtOH-induced mitochondrial dysfunction to maintain redox homeostasis and prevent abnormal propagation of esophageal stem/progenitor cells, providing novel mechanistic insights into the role of dysfunctional alcohol metabolism and autophagy in esophageal carcinogenesis. Citation Format: Prasanna Modayur Chandramouleeswaran, Kelly A. Whelan, Noah Engel, Jason Correnti, Koji Tanaka, Satish Srinivasan, Manti Guha, Rotonya Carr, Andres J. Klein-Szanto, Balaraman Kalyanaraman, Che-hong Chen, Daria Mochly-Rosen, Narayan G. Avadhani, Hiroshi Nakagawa. ALDH2 restricts esophageal stem/progenitor cell expansion via autophagy under alcohol-induced mitochondrial stress [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 5061.

A molecular mechanism of nickel (II): reduction of nucleotide excision repair activity by structural and functional disruption of p53.

Nickel is a major carcinogen that is implicated in tumor development through occupational and environmental exposure. Although the exact molecular mechanisms of carcinogenesis by low-level nickel remain unclear, inhibition of DNA repair is frequently considered to be a critical mechanism of carcinogenesis. Here, we investigated whether low concentrations of nickel would affect p53-mediated DNA repair, especially nucleotide excision repair. Our results showed that nickel inhibited the promoter binding activity of p53 on the downstream gene GADD45A, as a result of the disturbance of p53 oligomerization by nickel. In addition, we demonstrated that nickel exposure trigger the reduction of GADD45A-mediated DNA repair by impairing the physical interactions between GADD45A and proliferating cell nuclear antigen or xeroderma pigmentosum G. Notably, in the GADD45A-knockdown system, the levels of unrepaired DNA photoproducts were higher than wild-type cells, elucidating the importance of GADD45A in the nickel-associated inhibition of DNA repair. These results imply that inhibition of p53-mediated DNA repair can be considered a potential carcinogenic mechanism of nickel at low concentrations.

Aldehydes are the predominant forces inducing DNA damage and inhibiting DNA repair in tobacco smoke carcinogenesis

Tobacco smoke (TS) contains numerous cancer-causing agents, with polycyclic aromatic hydrocarbons (PAHs) and nitrosamines being most frequently cited as the major TS human cancer agents. Many lines of evidence seriously question this conclusion. To resolve this issue, we determined DNA adducts induced by the three major TS carcinogens: benzo(a)pyrene (BP), 4-(methylnitrosamine)-1-(3-pyridyl)-1-butanoe (NNK), and aldehydes in humans and mice. In mice, TS induces abundant aldehyde-induced γ-hydroxy-propano-deoxyguanosine (γ-OH-PdG) and α-methyl-γ-OH-PdG adducts in the lung and bladder, but not in the heart and liver. TS does not induce the BP- and NNK-DNA adducts in lung, heart, liver, and bladder. TS also reduces DNA repair activity and the abundance of repair proteins, XPC and OGG1/2, in lung tissues. These TS effects were greatly reduced by diet with polyphenols. We found that γ-OH-PdG and α-methyl-γ-OH-PdG are the major adducts formed in tobacco smokers' buccal cells as well as the normal lung tissues of tobacco-smoking lung cancer patients, but not in lung tissues of nonsmokers. However, the levels of BP- and NNK-DNA adducts are the same in lung tissues of smokers and nonsmokers. We found that while BP and NNK can induce BPDE-dG and O6-methyl-dG adducts in human lung and bladder epithelial cells, these inductions can be inhibited by acrolein. Acrolein also can reduce DNA repair activity and repair proteins. We propose a TS carcinogenesis paradigm. Aldehydes are major TS carcinogens exerting dominant effect: Aldehydes induce mutagenic PdG adducts, impair DNA repair functions, and inhibit many procarcinogens in TS from becoming DNA-damaging agents.

Targeting PRPK and TOPK for skin cancer prevention and therapy.

Solar ultraviolet (sUV) irradiation is a major environmental carcinogen that can cause inflammation and skin cancer. The costs and morbidity associated with skin cancer are increasing, and therefore identifying molecules that can help prevent skin carcinogenesis is important. In this study, we identified the p53-related protein kinase (PRPK) as a novel oncogenic protein that is phosphorylated by the T-LAK cell-originated protein kinase (TOPK). Knockdown of TOPK inhibited PRPK phosphorylation and conferred resistance to solar simulated light (SSL)-induced skin carcinogenesis in mouse models. In the clinic, acute SSL irradiation significantly increased epidermal thickness as well as total protein and phosphorylation levels of TOPK and PRPK in human skin tissues. We identified two PRPK inhibitors, FDA-approved rocuronium bromide (Zemuron®) or betamethasone 17-valerate (Betaderm®) that could attenuate TOPK-dependent PRPK signaling. Importantly, topical application of either rocuronium bromide or betamethasone decreased SSL-induced epidermal hyperplasia, neovascularization and cutaneous squamous cell carcinoma (cSCC) development in SKH1 (Crl: SKH1-Hrhr) hairless mice by inhibiting PRPK activation, and also reduced expression of the proliferation and oncogenesis markers, COX-2, cyclin D1 and MMP-9. This study is the first to demonstrate that targeting PRPK could be useful against sUV-induced cSCC development.

Toxicological effects of Camellia sinensis (green tea): A review.

Many scientific articles proved that green tea (GT), Camellia sinensis, has a great potential to manage central nervous system, cardiovascular, and metabolic diseases and treat cancer and inflammatory disorders. However, it is important to consider that "natural" is not always "safe." Some relevant articles reported side effects of GT, detrimental effects on health. The aim of this study is to provide a classified report about the toxicity of GT and its main constituents in acute, subacute, subchronic, and chronic states. Furthermore, it discusses on the cytotoxicity, genotoxicity, mutagenicity, carcinogenicity, and developmental toxicity of GT and its main constituents. The most important side effects have been reported hepatotoxicity and gastrointestinal disorders specially while consumed on an empty stomach. GT and its main components are not major teratogen, mutagen, or carcinogen substances. However, there is limited data in using them during pregnancy, and they should be used with caution in pregnancy, breast-feeding, and susceptible people. Because GT and its main components have a wide variety of drug interactions, consideration should be taken in coadministration of them with narrow therapeutic indexed drugs. Furthermore, they evoke selective cytotoxicity on cancerous cells that could engage them as an adjuvant substance in cancer therapy.

Helicobacter pylori infection promotes Aquaporin 3 expression via the ROS-HIF-1α-AQP3-ROS loop in stomach mucosa: a potential novel mechanism for cancer pathogenesis.

Helicobacter pylori (H. pylori) is the major stomach carcinogen, but the molecular mechanism responsible for the pathogenesis of cancer development mediated by H. pylori infection is still unclear. Aquaporin 3 (AQP3), overexpressed in gastric carcinoma, has a crucial role in gastric carcinogenesis and progression. However, the triggers and precise regulations for AQP3 upregulation during gastric carcinogens also remain unknown. Here we report that H. pylori infection-mediated carcinogenesis may be mechanistically depended on the upregulation of AQP3 expression via reactive oxygen species (ROS) pathway activation in the stomach. The retrospective analyses of clinical samples from patients with gastric carcinoma and other different stages of gastric diseases indicated that AQP3 expression was positively associated with gastric mucosal disease progression and H. pylori infection status as well. Furthermore, H. pylori infection significantly upregulated AQP3 and HIF-1α expression and increased ROS amount in human gastric epithelial AGS and GES-1 cells. Blockage of ROS with inhibitors, NAC and DPI, markedly decreased the expression of AQP3 and HIF-1α in both AGS and GES-1 cells simultaneously. Furthermore, the increased AQP3 in cells was mechanistically due to the transcriptional regulation by HIF-1α. In addition, H. pylori infection exerted production of proinflammatory cytokines IL-6, IL-8, and TNF depending on AQP3 level. Importantly, these in vitro novel findings were further investigated in vivo in a mouse H. pylori infectious model. Our current studies identify the mechanistic link between H. pylori infection and AQP3 upregulation in the pathogenesis of gastric carcinoma, which involves the activation of the ROS-HIF-1α axis and the exacerbated ROS-HIF-1α-AQP3-ROS loop.

Suppression of the photocatalytic activity of TiO2 nanoparticles encapsulated by chitosan through a spray-drying method with potential for use in sunblocking applications

Solar exposure, in particular to UVA and UVB radiation, is a major carcinogen through direct DNA damage and the production of reactive oxygen species (ROS). Inorganic UV filters present in sunscreening agents, such as titanium dioxide (TiO2), are commonly employed for protection however, due to their photocatalytic nature, they have been shown to instigate the production of ROS when irradiated with UV radiation, which in turn can lead to the degradation of the sunscreening formulation and subsequent damage to the skin. In this work, chitosan/TiO2 nanocomposite particles were produced via a spray-drying method, in a single step, directly through aqueous solution for the purpose of reducing the photocatalytic activity of commercially available TiO2 nanoparticles. The photocatalytic activity of the nanocomposite materials were assessed using the organic dye, crystal violet, as the degradation target and irradiating in a UV reactor. It was found that the photoactivity of the chitosan encapsulated nanoparticles were greatly reduced compared to that of the pristine TiO2 nanoparticles, from 95% degradation after 120 min for pristine TiO2 to 39.5% for the chitosan/TiO2 spray dried particles, highlighting the potential for this simple coating process and chitosan material for application as an inactive protective coating for sunblocking applications.

Major Chemical Carcinogens in Drinking Water Sources: Health Implications Due to Illegal Gold Mining Activities in Zamfara State-Nigeria

The study aimed at quantifying the levels of three major chemical carcinogens (MCCs): Pb, Cd, and Cr in wells, boreholes, and river water sources using HRCS AAS of Bagega, Sunke, and Dareta villages of Zamfara State, Nigeria; estimating daily MCC intake; and determining the cancer and non-cancer risks associated with MCC exposure. In total, 202 water samples were collected from the three villages and their MCC contents measured. The levels of Pb in the three Wells in Sunke ranged between 21 ± 9.8 and 326 ± 13.1 mg/L. The highest concentrations of Cd and Cr of 15 ± 7.1 and 96 ± 9.7 mg/L, respectively, were obtained in the affected area in Sunke village. A high incremental lifetime cancer risk of 1.59 × 103 for children due to Cd was obtained in Bagega village. The computed average Chronic hazard index (CHI) values for adults and children were 2.81 × 106 and 2.32 × 106, respectively, in drinking water from the river in Bagega village followed by an average values of 1.62 × 106 and 1.35 × 106 in drinking water from boreholes in Sunke village. The target hazard quotients and the CHI values were far greater than 1 indicating a high risk of adverse health outcomes.

Viral E6/E7 oncogene and cellular hexokinase 2 expression in HPV-positive cancer cell lines.

Oncogenic types of human papillomaviruses (HPVs) are major human carcinogens. Cancer cells typically exhibit metabolic alterations which support their malignant growth. These include an enhanced rate of aerobic glycolysis (‘Warburg effect’) which in cancer cells is often linked to an increased expression of the rate-limiting glycolytic enzyme Hexokinase 2 (HK2). Intriguingly, recent studies indicate that the HPV E6/E7 oncogenes cause the metabolic reprogramming in HPV-positive cancer cells by directly upregulating HK2 expression. Notably, however, these results were obtained upon ectopic overexpression of E6/E7. Here, we investigated whether HK2 levels are affected by the endogenous E6/E7 amounts present in HPV-positive cancer cell lines. RNA interference analyses reveal that the sustained E6/E7 expression is critical to maintain HK2 expression levels in HeLa cells. Mechanistically, this effect is linked to the E6/E7-dependent upregulation of HK2-stimulatory MYC expression and the E6/E7-induced downregulation of the HK2-inhibitory micro(mi)RNA miR-143-3p. Importantly, however, a stimulatory effect of E6/E7 on HK2 expression was observed only in HeLa among a panel of 8 different HPV-positive cervical and head and neck cancer cell lines. Thus, whereas these results support the notion that E6/E7 can increase HK2 expression, they argue against the concept that the viral oncogenes, at endogenous expression levels, commonly induce the metabolic switch of HPV-positive cancer cells towards aerobic glycolysis by directly or indirectly stimulating HK2 expression.

Virus/Host Cell Crosstalk in Hypoxic HPV-Positive Cancer Cells.

Oncogenic types of human papillomaviruses (HPVs) are major human carcinogens. The expression of the viral E6/E7 oncogenes plays a key role for HPV-linked oncogenesis. It recently has been found that low oxygen concentrations (“hypoxia”), as present in sub-regions of HPV-positive cancers, strongly affect the interplay between the HPV oncogenes and their transformed host cell. As a result, a state of dormancy is induced in hypoxic HPV-positive cancer cells, which is characterized by a shutdown of viral oncogene expression and a proliferative arrest that can be reversed by reoxygenation. In this review, these findings are put into the context of the current concepts of both HPV-linked carcinogenesis and of the effects of hypoxia on tumor biology. Moreover, we discuss the consequences for the phenotype of HPV-positive cancer cells as well as for their clinical behavior and response towards established and prospective therapeutic strategies.

Benzop(a)pyrene in Cigarette Smoke Enhances HIV‐1 Replication in Monocytic Cells: Potential Role of Cytochrome P450s and Oxidative Stress

Smoking aggravates HIV‐1 pathogenesis and leads to decreased responses to antiretroviral therapy. However, the exact mechanism by which smoking enhances HIV‐1 pathogenesis is unknown. Benzo(a)pyrene (BaP) is a major carcinogen in cigarette smoke. It requires metabolic activation through cytochrome P450s (CYPs) to exert its toxic effects. We hypothesize that BaP is metabolized by CYP‐mediated pathway and produce reactive oxygen species (ROS). Oxidative stress resulting from increased ROS would aggravate HIV‐1 replication. Recently, we have shown that chronic exposure of BaP to U937 monocytic cells results in the significant induction of CYPs (CYP 1A1 and 3A4) and antioxidants (catalase and SOD1), and increase in the generation of ROS and cellular toxicity. We also confirmed these results in human primary macrophages. Next, we examined the chronic effect of BaP in U1 cells (HIV‐1 infected U937 monocytic cells). When U1 cells were exposed to BaP for seven days, we observed ~4 fold increase in HIV‐1 replication. In addition, there was a significant increase in the expression of CYP1A1 at mRNA level as well as there was an increase in CYP1A1 enzymatic activity. Elevated ROS and massive cell death were also observed. In order to confirm the role of CYP1A1 and 1B1 in BaP‐induced oxidative stress pathway and HIV‐1 replication, we knocked down the gene expression of CYP1A1 and 1B1 in HIV‐1 infected human primary macrophages, using SiRNA. Our results showed decrease in the viral replication in CYP1A1 and 1B1 knock‐out cells upon BaP exposure. We will determine oxidative DNA damage in untreated, and CYP1A1 and 1B1 knock‐out cells using TUNNEL assay. We are now in the process of determining the role of oxidative stress mediated through CYP pathway in HIV‐1 replication. For this, we will expose HIV‐infected primary macrophages with general and specific antioxidants, followed by exposure to BaP. Finally, we will quantify BaP metabolites in U1 cells and/or HIV‐1 infected human primary macrophages using Liquid Chromatography‐Mass Spectrometry. The results from the present work are clinically relevant as they would help to find a novel therapeutic target to improve drug therapy outcomes in HIV positive smokers.Support or Funding InformationDepartment of Pharmaceutical Sciences, University of Tennessee Health Science Center

Smokeless tobacco (paan and gutkha) consumption, prevalence, and contribution to oral cancer.

Smokeless tobacco consumption, which is widespread throughout the world, leads to oral submucous fibrosis (OSMF), which is a long-lasting and devastating condition of the oral cavity with the potential for malignancy. In this review, we mainly focus on the consumption of smokeless tobacco, such as paan and gutkha, and the role of these substances in the induction of OSMF and ultimately oral cancer. The list of articles to be examined was established using citation discovery tools provided by PubMed, Scopus, and Google Scholar. The continuous chewing of paan and swallowing of gutkha trigger progressive fibrosis in submucosal tissue. Generally, OSMF occurs due to multiple risk factors, especially smokeless tobacco and its components, such as betel quid, areca nuts, and slaked lime, which are used in paan and gutkha. The incidence of oral cancer is higher in women than in men in South Asian countries. Human oral epithelium cells experience carcinogenic and genotoxic effects from the slaked lime present in the betel quid, with or without areca nut. Products such as 3-(methylnitrosamino)-proprionitrile, nitrosamines, and nicotine initiate the production of reactive oxygen species in smokeless tobacco, eventually leading to fibroblast, DNA, and RNA damage with carcinogenic effects in the mouth of tobacco consumers. The metabolic activation of nitrosamine in tobacco by cytochrome P450 enzymes may lead to the formation of N-nitrosonornicotine, a major carcinogen, and micronuclei, which are an indicator of genotoxicity. These effects lead to further DNA damage and, eventually, oral cancer.

Induction of dormancy in hypoxic human papillomavirus-positive cancer cells

Oncogenic human papillomaviruses (HPVs) are closely linked to major human malignancies, including cervical and head and neck cancers. It is widely assumed that HPV-positive cancer cells are under selection pressure to continuously express the viral E6/E7 oncogenes, that their intracellular p53 levels are reconstituted on E6/E7 repression, and that E6/E7 inhibition phenotypically results in cellular senescence. Here we show that hypoxic conditions, as are often found in subregions of cervical and head and neck cancers, enable HPV-positive cancer cells to escape from these regulatory principles: E6/E7 is efficiently repressed, yet, p53 levels do not increase. Moreover, E6/E7 repression under hypoxia does not result in cellular senescence, owing to hypoxia-associated impaired mechanistic target of rapamycin (mTOR) signaling via the inhibitory REDD1/TSC2 axis. Instead, a reversible growth arrest is induced that can be overcome by reoxygenation. Impairment of mTOR signaling also interfered with the senescence response of hypoxic HPV-positive cancer cells toward prosenescent chemotherapy in vitro. Collectively, these findings indicate that hypoxic HPV-positive cancer cells can induce a reversible state of dormancy, with decreased viral antigen synthesis and increased therapeutic resistance, and may serve as reservoirs for tumor recurrence on reoxygenation.

Association between UGT1A1*28*28 genotype and lung cancer in the Japanese population.

Lung cancer is the leading cause of cancer death and is closely linked to tobacco smoking. Genetic polymorphisms in genes that encode enzymes involved in metabolizing tobacco carcinogens could affect an individual's risk for lung cancer. While polymorphism of UDP-glucuronosyltransferase1A1 (UGT1A1) is involved in detoxification of benzo(a)pyrene-7,8-dihydrodiol(-), a major tobacco carcinogen, the association between UGT1A1 genotype and lung cancer has not been examined. We retrieved the clinical data of 5,285 patients who underwent systemic chemotherapy at Kyoto University Hospital. A total of 765 patients (194 lung cancer patients and 671 patients with other malignancies) with UGT1A1 genotyping data were included in this analysis. We used logistic regression with recessive, dominant, and additive models to identify differences in genotype frequencies between lung cancer and other malignancies. In the recessive model, UGT1A1*28*28 genotype was significantly associated with lung cancer compared to other malignancies (odds ratio5.3, P=0.0083). Among lung cancer patients with a smoking history, squamous cell carcinoma was significantly predominant in patients with UGT1A1*28*28 compared to those with other UGT1A1 genotypes (P=0.024). This is the first study to demonstrate a significant association between the homozygous UGT1A1*28 genotype and lung cancer.

Exposure to Nicotine and Selected Toxicants in Cigarette Smokers Who Switched to Electronic Cigarettes: A Longitudinal Within-Subjects Observational Study.

Electronic cigarettes (e-cigarettes) are purported to deliver nicotine aerosol without any toxic combustion products present in tobacco smoke. In this longitudinal within-subjects observational study, we evaluated the effects of e-cigarettes on nicotine delivery and exposure to selected carcinogens and toxicants. We measured seven nicotine metabolites and 17 tobacco smoke exposure biomarkers in the urine samples of 20 smokers collected before and after switching to pen-style M201 e-cigarettes for 2 weeks. Biomarkers were metabolites of 13 major carcinogens and toxicants in cigarette smoke: one tobacco-specific nitrosamine (NNK), eight volatile organic compounds (1,3-butadiene, crotonaldehyde, acrolein, benzene, acrylamide, acrylonitrile, ethylene oxide, and propylene oxide), and four polycyclic aromatic hydrocarbons (naphthalene, fluorene, phenanthrene, and pyrene). Changes in urine biomarkers concentration were tested using repeated measures analysis of variance. In total, 45% of participants reported complete abstinence from cigarette smoking at 2 weeks, while 55% reported continued smoking. Levels of total nicotine and some polycyclic aromatic hydrocarbon metabolites did not change after switching from tobacco to e-cigarettes. All other biomarkers significantly decreased after 1 week of using e-cigarettes (p < .05). After 1 week, the greatest percentage reductions in biomarkers levels were observed for metabolites of 1,3-butadiene, benzene, and acrylonitrile. Total NNAL, a metabolite of NNK, declined by 57% and 64% after 1 and 2 weeks, respectively, while 3-hydroxyfluorene levels declined by 46% at week 1, and 34% at week 2. After switching from tobacco to e-cigarettes, nicotine exposure remains unchanged, while exposure to selected carcinogens and toxicants is substantially reduced. To our knowledge, this is the first study that demonstrates that substituting tobacco cigarettes with an e-cigarette may reduce user exposure to numerous toxicants and carcinogens otherwise present in tobacco cigarettes. Data on reduced exposure to harmful constituents that are present in tobacco cigarettes and e-cigarettes can aid in evaluating e-cigarettes as a potential harm reduction device.

Single-molecule force spectroscopy study of the effect of cigarette carcinogens on thrombomodulin–thrombin interaction

Exposure to cigarette smoke is a major risk factor for cancer and cardiovascular disease. Thrombosis is regarded as the main reason for smoking-related cardiovascular disease. However, the detail mechanism of how smoking promotes thrombosis is not fully understood. In this work, we investigated the impacts of one major cigarette carcinogens 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) as well as its metabolite 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol (NNAL) on a key process in thrombosis regulation: thrombin–thrombomodulin (TM) binding. Atomic force microscopy based single-molecule force spectroscopy was applied to measure both in vitro and in vivo binding force of thrombin to TM in the absence and presence of NNK and NNAL respectively. The results revealed that NNK and NNAL can reduce the binding probability of TM and thrombin. The inhibition effect and underlying mechanism was further studied by molecular simulation. As indicated by our results, the cigarette carcinogens could cause a higher risk of thrombosis through the disruption of TM–thrombin interaction.