From cultivation to cancer: formation of N-nitrosamines and other carcinogens in smokeless tobacco and their mutagenic implications.

Tobacco use is a major cause of preventable morbidity and mortality globally. Tobacco products, including smokeless tobacco (ST), generally contain tobacco-specific N-nitrosamines (TSNAs), such as N'-nitrosonornicotine (NNN) and 4-(methylnitrosamino)-1-(3-pyridyl)-butanone (NNK), which are potent carcinogens that cause mutations in critical genes in human DNA. This review covers the series of biochemical and chemical transformations, related to TSNAs, leading from tobacco cultivation to cancer initiation. A key aim of this review is to provide a greater understanding of TSNAs: their precursors, the microbial and chemical mechanisms that contribute to their formation in ST, their mutagenicity leading to cancer due to ST use, and potential means of lowering TSNA levels in tobacco products. TSNAs are not present in harvested tobacco but can form due to nitrosating agents reacting with tobacco alkaloids present in tobacco during certain types of curing. TSNAs can also form during or following ST production when certain microorganisms perform nitrate metabolism, with dissimilatory nitrate reductases converting nitrate to nitrite that is then released into tobacco and reacts chemically with tobacco alkaloids. When ST usage occurs, TSNAs are absorbed and metabolized to reactive compounds that form DNA adducts leading to mutations in critical target genes, including the RAS oncogenes and the p53 tumor suppressor gene. DNA repair mechanisms remove most adducts induced by carcinogens, thus preventing many but not all mutations. Lastly, because TSNAs and other agents cause cancer, previously documented strategies for lowering their levels in ST products are discussed, including using tobacco with lower nornicotine levels, pasteurization and other means of eliminating microorganisms, omitting fermentation and fire-curing, refrigerating ST products, and including nitrite scavenging chemicals as ST ingredients.

Impact of the WHO FCTC on non-cigarette tobacco products.

INTRODUCTION: This paper investigates to what extent Framework Convention on Tobacco Control (FCTC) parties have successfully implemented regulatory measures against non-cigarette tobacco product (NCTP) use, considers the challenges and peculiarities in applying such regulations and proposes effective means. DATA AND METHODS: This review was based on many sources mainly: International Legal Consortium, International Tobacco Control, Campaign for Tobacco-Free Kids, FCTC, expert group visits and published literature. FINDINGS AND CONCLUSION: The FCTC provided a framework that applies to all forms of tobacco and this encouraged some parties to adopt control measures against NCTP and to incorporate them into their national tobacco control plans. Although a number of countries have adopted measures specifically targeted towards smokeless and waterpipe tobacco, greater global progress is needed. The strongest achievements have been in protection from exposure to tobacco smoke; controlling advertising, promotion and sponsorship; controlling sales to and by minors; education, communication and public awareness; and packaging and labelling of NCTP. Countries which adopted broad definitions of tobacco products have demonstrated encouraging trends in curbing their use. Future work should address the deep-rooted social acceptance of NCTP, the laxity in their control, their exclusion from regulations in some countries and the failure to subject them to increased taxation. Control measures should also specifically target the initiation risk to youth and adolescents and all factors that contribute to that such as banning flavourings and promotions through social media. Stronger global surveillance of NCTP use, tracking of policy implementation and evaluation of policy impact will provide important evidence to assist parties in fully implementing the FCTC to control their use.

ST1926, an orally active synthetic retinoid, induces apoptosis in chronic myeloid leukemia cells and prolongs survival in a murine model.

The tyrosine kinase inhibitor, imatinib, is the first line of treatment for chronic myeloid leukemia (CML) patients. Unfortunately, patients develop resistance and relapse due to bcr-abl point mutations and the persistence of leukemia initiating cells (LIC). Retinoids regulate vital biological processes such as cellular proliferation, apoptosis, and differentiation, in particular of hematopoietic progenitor cells. The clinical usage of natural retinoids is hindered by acquired resistance and undesirable side effects. However, bioavailable and less toxic synthetic retinoids, such as the atypical adamantyl retinoid ST1926, have been developed and tested in cancer clinical trials. We investigated the preclinical efficacy of the synthetic retinoid ST1926 using human CML cell lines and the murine bone marrow transduction/transplantation CML model. In vitro, ST1926 induced irreversible growth inhibition, cell cycle arrest and apoptosis through the dissipation of the mitochondrial membrane potential and caspase activation. Furthermore, ST1926 induced DNA damage and downregulated BCR-ABL. Most importantly, oral treatment with ST1926 significantly prolonged the longevity of primary CML mice, and reduced tumor burden. However, ST1926 did not eradicate LIC, evident by the ability of splenocytes isolated from treated primary mice to develop CML in untreated secondary recipients. These results support a potential therapeutic use of ST1926 in CML targeted therapy.

Health effects associated with waterpipe smoking.

OBJECTIVE: It is widely held that waterpipe smoking (WPS) is not associated with health hazards. However, several studies have documented the uptake of several toxicants and carcinogens during WPS that is strongly associated with harmful health effects. This paper reviews the literature on the health effects of WPS. DATA SOURCES: Three databases-PubMed, MEDLINE and EMBASE-were searched until August 2014 for the acute and long-term health effects of WPS using the terms 'waterpipe' and its synonyms (hookah, shisha, goza, narghileh, arghileh and hubble-bubble) in various spellings. STUDY SELECTION: We included original clinical studies, case reports and systematic reviews and focused on clinical human studies. ∼10% of the identified studies met the selection criteria. DATA EXTRACTION: Data were abstracted by all three authors and summarised into tables. Abstracted data included study type, results and methodological limitations and were analysed jointly by all three authors. DATA SYNTHESIS: WPS acutely leads to increased heart rate, blood pressure, impaired pulmonary function and carbon monoxide intoxication. Chronic bronchitis, emphysema and coronary artery disease are serious complications of long-term use. Lung, gastric and oesophageal cancer are associated with WPS as well as periodontal disease, obstetrical complications, osteoporosis and mental health problems. CONCLUSIONS: Contrary to the widely held misconception, WPS is associated with a variety of adverse short-term and long-term health effects that should reinforce the need for stronger regulation. In addition, this review highlights the limitations of the published work, which is mostly cross-sectional or retrospective. Prospective studies should be undertaken to assess the full spectrum of health effects of WPS, particularly in view of its growing popularity and attractiveness to youth.

A comparative study of systemic carcinogen exposure in waterpipe smokers, cigarette smokers and non-smokers.

BACKGROUND: In the past decade, waterpipe smoking-also known as hookah, shisha, narghileh-has increased among youth. The scarcity of rigorous studies linking waterpipe smoking to smoking-related diseases has hindered policy and regulatory efforts to confront the waterpipe epidemic. This study compares systemic carcinogen exposure between independent groups of exclusive waterpipe smokers, cigarette smokers and non-smokers. METHODS: This study was conducted at the Syrian Center for Tobacco Studies (SCTS) in Aleppo, Syria, between 2010 and 2011. First morning urinary samples were collected from three groups of subjects; exclusive daily waterpipe smokers (n=24), exclusive daily cigarette smokers (n=23), and non-smokers (n=28). These samples were analysed for carcinogenic tobacco-specific nitrosamines 4-(methylnitrosamino)-1-(3-pyridyl)-1- butanol (NNAL) using liquid chromatography-tandem mass spectrometry (LC-MS/MS). RESULTS: Our results show that waterpipe smokers are exposed to about 5-10 times greater NNAL than non-smokers. Mean (95% CI) free and total NNAL was 0.7 (0.3 to 1. 4) and 3.9 (1.6 to 9.5) pg/mL urine for non-smokers, 8.4 (4.8 to 14.8) and 33.0 (21.6 to 50.6) pg/mL urine for waterpipe smokers, and 10.7 (5.0 to 22.6) and 46.8 (27.6 to 79.3) pg/mL urine for cigarette smokers (p<0.001 for all comparisons). Daily waterpipe smokers were less exposed to NNAL than daily cigarette smokers, although the difference did not reach statistical significance for all measurements. CONCLUSIONS: These results provide the clearest indication to date about systemic exposure to harmful carcinogens associated with long-term waterpipe smoking. Such evidence can support policy and regulatory efforts designed to confront the emerging global waterpipe epidemic, as well as drive interventions aimed at increasing the public awareness about the cancer risk associated with waterpipe smoking.

Effective targeting of chronic myeloid leukemia initiating activity with the combination of arsenic trioxide and interferon alpha.

Imatinib is the standard of care in chronic meloid leukemia (CML) therapy. However, imatinib is not curative since most patients who discontinue therapy relapse indicating that leukemia initiating cells (LIC) are resistant. Interferon alpha (IFN) induces hematologic and cytogenetic remissions and interestingly, improved outcome was reported with the combination of interferon and imatinib. Arsenic trioxide was suggested to decrease CML LIC. We investigated the effects of arsenic and IFN on human CML cell lines or primary cells and the bone marrow retroviral transduction/transplantation murine CML model. In vitro, the combination of arsenic and IFN inhibited proliferation and activated apoptosis. Importantly, arsenic and IFN synergistically reduced the clonogenic activity of primary bone marrow cells derived from CML patients. Finally, in vivo, combined interferon and arsenic treatment, but not single agents, prolonged the survival of primary CML mice. Importantly, the combination severely impaired engraftment into untreated secondary recipients, with some recipients never developing the disease, demonstrating a dramatic decrease in CML LIC activity. Arsenic/IFN effect on CML LIC activity was significantly superior to that of imatinib. These results support further exploration of this combination, alone or with imatinib aiming at achieving CML eradication rather than long-term disease control.

ASPL-TFE3 translocation in vulvovaginal alveolar soft part sarcoma.

Alveolar soft part sarcoma of the vulvovaginal region is limited to only 8 reported vaginal cases and 1 vulvar case in the English literature. The histogenesis of the tumor remains intriguing with postulates favoring a myogenic versus nonmyogenic origin. A reciprocal translocation for ASPL-TFE3 gene fusion, frequently detected in ~90% of cases, combined with TFE3 protein immunoexpression are highly sensitive and specific methods for diagnostic confirmation. The current report describes a unique case of vulvovaginal alveolar soft part sarcoma showing the classic morphologic features with documentation of TFE3 protein expression and the ASPL-TFE3 gene rearrangement. Furthermore, a brief review of the literature of vulvar and vaginal alveolar soft part sarcoma cases with the various treatment modalities is outlined.

Chronic lymphocytic leukemia associated with myelodysplastic syndrome with ring sideroblasts.

Diagnosis of myelodysplastic syndrome/acute myeloid leukemia (MDS/AML) and chronic lymphocytic leukemia (CLL) in the same patient is extremely rare. We describe a 69-year-old CLL patient who developed MDS with ring sideroblasts 1 year after diagnosis of CLL and without any previous treatment. Diagnosis was based on flow cytometry, bone marrow aspirate morphology, and iron stain. Our findings indicate that the 2 disorders belong to 2 different hematopoietic clones in this patient. In cases of worsening anemia in CLL patients, it is recommended that an iron stain be performed on bone marrow aspirates to exclude a coexistent malignancy such as refractory anemia with ring sideroblasts.

Prevalence of factor V R2 (H1299R) polymorphism in the Lebanese population.

AIMS: A recently identified polymorphism in factor V gene (His1299Arg; also named HR2) has been reported to be a possible risk factor for the development of venous thromboembolism (VTE), with a high prevalence of 9.5-15.2% in patients of different ethnic groups in different parts of the world. The aim of this study is to assess the prevalence of HR2 haplotype in Lebanon. METHODS: We randomly selected 125 samples from unrelated donors logged into our HLA registry; these represent healthy Lebanese individuals originating from different provinces and religious communities of the country. Their DNA was extracted using the Pel-Freez extraction kit and stored at -80 degrees C for later use. The CVD StripAssay was used for PCR and reverse hybridisation. It screens for several gene mutations including factor V H1299R. RESULTS: A total of 125 controls were studied: 72 males and 53 females with a median age 42 years. Thirteen (10.4%) had the HR2 haplotype; 11 (8.8%) were heterozygous (R1/R2), and two (1.6%) were homozygous (R2/R2), with an allelic frequency of 0.06. CONCLUSIONS: Our study is the first report from Lebanon that describes the prevalence of HR2 haplotype and the frequency of its alleles. We are reporting a high prevalence of the HR2 in our population (10.4%). The hypothesis that A4070G polymorphism might contribute to the expression of a thrombotic phenotype deserves to be tested in our population through larger studies.

Cervical ganglioneuroma in collision with a metastatic undifferentiated carcinoma.

Cervical ganglioneuromas are extremely rare with approximately six case reports. The current report highlights a unique collision tumor between a cervical ganglioneuroma and a metastatic undifferentiated carcinoma arising from a primary gingival mass. A 53-year-old male presented with a 2 cm left gingival mass that was excised and treated with systemic chemotherapy. Consequently, 9 months later, he developed a 3.2 cm left submandibular mass followed by recurrence of the left gingival mass. From the clinicopathologic perspective, this had to be separated from the differentials: ganglioneuroblastoma or metastatic involvement of a lymph node from primary gingival undifferentiated carcinoma.

Nicotine exposure in daily waterpipe smokers and its relation to puff topography.

Waterpipe tobacco smoking is increasing in popularity worldwide and available evidence point to its addictive and harmful potential. This study is conducted to assess nicotine exposure in daily waterpipe smokers, and its correlation with puff topography parameters. Sixty-one waterpipe tobacco smokers (56 males; mean age±SD, 30.9±9.5years; mean number of weekly waterpipe smoking episodes 7.8±5.7) abstained from smoking for at least 24h, and then smoked tobacco from a waterpipe ad libitum in a laboratory setting. During the session puff topography parameters were monitored continuously, and pre- and post-smoking expired-air CO was measured. Before and after smoking, venous blood was sampled for the assessment of plasma nicotine using Gas Chromatography-Mass Spectrometry. The average pre- and post-smoking expired-air CO was 4±1.7 and 35.5±32.7ppm, respectively (i.e., a CO boost of 31.5ppm, p<.001). Mean plasma nicotine concentration increased from 3.07±3.05ng/ml pre-smoking to 15.7±8.7ng/ml post-smoking (p<.001). Plasma nicotine boost was correlated with total session time (Pearson correlation coefficient r=.31, p=.04), cumulative puff duration (r=.37, p=.01), mean puff duration (r=.34, p=.02), and total smoke inhaled in the session (r=.34, p=.02. These data show considerable nicotine exposure in daily waterpipe smokers, and that nicotine exposure is a function of waterpipe smoking patterns.

Essential thrombocythemia with myelofibrosis transformed into acute myeloid leukemia with der(1;15)(q10;q10): case report and literature review.

Translocations involving chromosomes 1 and 15 are uncommon in hematologic malignancies. So far, only 42 cases have been reported with t(1;15) as a reciprocal or complex chromosomal abnormalities. We herein report the first case in the literature, to our knowledge, of a 44-year-old female with essential thrombocythemia and severe myelofibrosis who developed acute myeloid leukemia (AML-M4) with der(1;15)(q10;q10) after 13 years of treatment. In addition, we reviewed the literature for all up-to-date published cases with t(1;15).

Killer cell immunoglobulin-like receptor genotypes in Behçet's disease patients: any role for the 3DP1*001/002 pseudogene?

AIMS: Genotypic profiles of the natural killer cell immunoglobulin-like receptors (KIR) have been reported to vary among different ethnic groups and variable clinical entities. This study represents the second report on its distribution among patients with Behçet's disease (BD). We studied 43 unrelated Lebanese Behçet's patients, had their DNA typed using sequence-specific primer technique for the presence of 16 KIR genes and pseudogenes loci, and compared them to the general Lebanese population. RESULTS: In addition to sharing common features with the general population, the AA genotype was still the most frequent--however, with five new KIR profiles identified. There was no statistically significant distribution of the different KIR genes between the cases (BD patients) and controls (Lebanese population); however, KIR3DP1*001/002 was found to be significantly different between the BD patients and the Lebanese population, but this significance was lost after correction for all KIR loci. CONCLUSION: The results lead to an interesting future research question of whether or not KIR genotype is involved in the predisposition to or pathogenesis of BD especially that a pseudogene is controversially in question. This is the second report that describes the KIR genotypic profile in such an important clinical disease but the first to shed a light on the possible role of a pseudogene.

Distribution of killer cell immunoglobulin-like receptor (KIR) genotypes in patients with familial Mediterranean fever.

Genotypic profiles of the natural killer cell immunoglobulin-like receptors (KIR) have been reported to vary among different ethnic groups and variable clinical entities. This study represents the first report on its distribution among patients with familial Mediterranean fever (FMF). We studied 56 unrelated Lebanese FMF patients, had their DNA typed using sequence-specific primer (SSP) technique for the presence of 16 KIR gene and pseudogene loci, and compared them to the general Lebanese population. The AA1 genotype was the most frequent in both the FMF and control groups. Six new KIR profiles were identified. The FMF group showed a higher prevalence of KIR 3DP1*003 (p<0.05) and an increase in the BB genotype compared with controls. The results lead to an interesting future research question of whether or not KIR genotype is involved in the predisposition to or pathogenesis of FMF. This is the first report that describes the KIR genotypic profile in this important clinical disease.

Plasminogen Activator Inhibitor-1 (PAI-1) gene 4G/5G alleles frequency distribution in the Lebanese population.

Plasminogen activator inhibitor-1 (PAI-1) is an inhibitor of fibrinolysis. Increased plasma PAI-1 levels play an essential role in the pathogenesis of cardiovascular risk and other diseases associated with thrombosis. The 4G/5G polymorphism of the PAI-1 promoter region has been extensively studied in different populations. We studied 160 healthy unrelated Lebanese individuals using a reverse hybridization PCR assay to detect the 5G/5G, 4G/5G and, 4G/4G genotypes of the PAI-1 gene and the frequencies of the 4G and 5G alleles. We found that 4G/5G genotype was the most prevalent (45.6%) followed by 5G/5G (36.9%) and 4G/4G (17.5%). The frequencies of the 4G and 5G alleles were calculated to be 0.403 and 0.597, respectively. Compared to other ethnic communities, the Lebanese population was found to harbour a relatively high prevalence of the rare 4G allele. This, in turn, may predispose this population to develop cardiovascular diseases and other thrombotic clinical conditions. This study aids to enhance our understanding of the genetic features of the Lebanese population.

Frequency distribution of the G/A alleles of the beta-fibrinogen gene in the Lebanese population.

Fibrinogen is a plasma protein that has been reported to be associated with an increased risk of atherothrombotic diseases and venous thrombosis. The most common polymorphism that has been studied so far in different populations is the G-455-->A polymorphism in the promoter region of the beta-fibrinogen gene. We studied 160 healthy unrelated Lebanese individuals for the prevalence of -455G/G, -455G/A and -455A/A genotypes of the beta-fibrinogen gene and the frequency of G and A alleles using a reverse hybridization PCR assay. The prevalence of the G/G, G/A, and A/A genotypes were found to be 60.6, 31.9 and 7.5%, respectively. The frequency of the G and A alleles were found to be 0.77 and 0.23, respectively. As compared to other ethnic groups, the Lebanese individuals were found to have a relatively high prevalence of the A allele which may predispose them to develop cardiovascular diseases as well as thrombotic events. This study provides additional unique genetic information pertaining to the Lebanese population.