Adverse effects of Sudanese toombak vs. Swedish snuff on human oral cells

J Oral Pathol Med (2010) 39 128–140 Background: The high incidence of oral cancer in Sudan has been associated with the use of toombak, the local type of smokeless tobacco. However, its specific effects on human oral cells are not known. We aimed to investigate the effects of toombak on primary normal human oral keratinocytes, fibroblasts, and a dysplastic oral keratinocytic cell line, and to compare them with the effects induced by Swedish snuff. Method: Aqueous extracts were prepared from moist toombak and Swedish snuff and added in serial dilutions on in vitro monolayer cultured cells. Cell viability, morphology and growth, DNA double‐strand breaks (γH2AX staining), expression of phosphatidylserine (Annexin V staining), and cell cycle were assessed after various exposure time periods. Results: Significant decrease in cell number, occurrence of DNA double‐strain breaks, morphological and biochemical signs of programmed cell death were detected in all oral cell types exposed to clinically relevant dilutions of toombak extract, although to a lesser extent in normal oral fibroblasts and dysplastic keratinocytes. G2/M‐block was also detected in normal oral keratinocytes and fibroblasts exposed to clinically relevant dilutions of toombak extract. Swedish snuff extract had less adverse effects on oral cells, mainly at non‐clinically relevant dilutions. Conclusion: This study indicates a potential for toombak, higher than for Swedish snuff, to damage human oral epithelium. Dysplastic oral keratinocytes were less sensitive than their normal counterparts, suggesting that they might have acquired a partially resistant phenotype to toombak‐induced cytotoxic effects while still being prone to DNA damage that could lead to further malignant progression.     

Changes in hepatitis B serologic titers in HIV+ and HIV– children with haemophilia

This longitudinal study examines differences in hepatitis B immune titres in children and adolescents with haemophilia to determine if they are dependent on how immunity was acquired (vaccination or natural infection), and whether they are related to the child's HIV status and/or are influenced by HIV disease progression. Serologic titres (HBcAb, HBsAb) and HBsAg were measured prospectively at baseline, and at years 1, 2 and 3 of follow-up in 126 HIV- and 207 HIV+ children and adolescents with haemophilia. Analyses were performed to assess the impact of HIV status on the measured titres, and for HIV+ subjects to examine the association with CD4+ lymphocyte counts and p24 antigen status. The results show that HIV+ children were more likely than HIV- children to lose vaccine-induced immunity as indicated by the loss of HBsAb. There was an increased risk of losing HBsAb with higher CD4+ counts and younger age. Re-immunization was not successful in seven of eight HIV+ children. Two subjects (one HIV+, one HIV-) entered the study HBsAg- but became HBsAg+ over the course of follow-up. Seven HIV+ subjects lost natural immunity as indicated by the loss of HBcAb. The loss of either HBsAb or HBcAb in HIV--subjects was negligible to absent. In conclusion, because of the loss of immunity in HIV+ children the viral safety of factor replacement concentrates for these children is an important consideration. HIV- children rarely lose immunity, therefore frequent measures of HBsAb are not necessary.     

Influence of Late Side-Effects Upon Daily Life After Radiotherapy for Laryngeal and Pharyngeal Cancer

The influence on daily life from long-term side-effects was studied in patients treated more than 5 years ago with radiotherapy for laryngeal and pharyngeal cancer. Forty-six patients were asked to participate in telephone interviews and 44 participated. Only a minority (10%) stated serious problems related to the treatment. Problems related to the voice and severe xerostomia were especially mentioned. Half of the patients treated for pharyngeal cancer, complained of xerostomia and had occasionally a feeling of being handicapped. Social relations were impaired in 10% of the patients and 10% had retired due to their cancer or therapy related side-effects. It is concluded that radiotherapy seems to be a well tolerated treatment with relatively little impairment of the daily life in patients with laryngeal cancer. In patients with pharyngeal cancer, xerostomia is a major problem, which often persists more than 5 years after treatment.     

Cytotoxicity and characterization of an active metabolite of benzamide riboside,a novel inhibitor of IMP dehydrogenase

Benzamide riboside exhibits significant cytotoxicity against a variety of human tumor cells in culture. On the basis of metabolic studies, the primary target of this drug's action appears to be IMP dehydrogenase (IMPDH). Incubation of human myelogenous leukemia K562 cells with an IC₅₀ concentration of benzamide riboside resulted in an expansion of IMP poofs (5.9-fold), with a parallel reduction in the concentration of GMP (90%), GDP (63%), GTP (55%) and dGTP (40%). On kinetic grounds, it was deduced that benzamide riboside (whose K_i versus IMPDH is 6.4 mM, while that of its 5′-monophosphate is 3.9 mM) or its 5′-monophosphate were unlikely to be responsible for inhibition of this target enzyme, IMPDH, since only micromolar concentrations of benzamide riboside were needed to exert potent inhibition of tumor-cell growth. Studies on the metabolism of this C-nucleoside have revealed the presence of a new peak eluting in the nucleoside diphosphate area on HPLC. Treatment of this peak with venom phosphodiesterase degraded it and concurrently nullified its inhibitory activity versus IMPDH; alkaline phosphatase, on the other hand, totally failed to digest the anabolite. These results suggest that the metabolite in question is the phosphodiester, benzamide adenine dinucleotide (BAD). Evidence that the inhibitor was an analog of NAD, wherein the nicotinamide moiety has been replaced by benzamide, was provided by both NMR and mass spectrometric analysis and confirmed by enzymatic synthesis. Further insight into the nature of the active principle was obtained from kinetic studies, which established that BAD competitively inhibited NAD utilization by partially purified IMPDH from K562 cells with a K_i of O.118 μM. In concert, these studies establish that benzamide riboside exhibits potent antiproliferative activity by inhibiting IMPDH through BAD.     

Mitochondrial Function in Heart Failure

Experimental and clinical studies have detected an impaired respiratory function of cardiomyocyte mitochondria in heart failure. Since the reasons for heart failure are manifold, so is mitochondrial involvement. Characteristics of mitochondrial participation in heart failure are as follows: (1) Inherited or acquired mutations of the mitochondrial or nuclear genome cause defects in different mitochondrial components, eventually resulting in cardiomyopathy. (2a) Oxidative stress depresses mitochondrial function. This occurs slowly and inevitably in the 'physiological' process of ageing, but rapidly in pathophysiologic conditions such as post-ischemic reperfusion. (2b) Free radicals damage mitochondrial DNA, proteins, and membrane lipids. Interactions between altered membrane lipids, respiratory chain components, and carrier proteins further enhance mitochondrial dysfunction. (3) Mitochondrial energy transfer via the adenine nucleotide translocator (ANT) and the mitochondrial creatine kinase is disturbed in heart failure. Especially an altered expression and a functional impairment of the ANT seems to be involved in the disturbed energy metabolism of dilated and inflammatory cardiomyopathy. (4) Mitochondria are mainly involved in the initiation and modulation of the process of programmed cell death (apoptosis). (5) Triggered by a variety of conditions during cellular dysfunction mitochondrial membrane permeability suddenly increases, followed by the collapse of the membrane potential, thus abolishing energy production and further aggravating cellular damage. (6a) Increased levels of cytokines, in particular TNF-, in heart failure and cardiomyopathy modulate mitochondrial function. (6b) Cytokines activate the generation of nitric oxide and heat shock proteins, thus further depressing or preserving mitochondrial activity.These main mechanisms of active and passive participation of mitochondria in heart failure are reviewed in this article. At present, most of them are not completely resolved and some are still hypothetical.     

Glucocorticosteroid receptors in ovarian carcinomas

The use of glucocorticoids (GCs) in oncology, including in the treatment of ovarian carcinomas, is controversial. In vitro experiments suggest that GCs negatively influence the response to chemotherapy, but the few available clinical data show only benefits. Glucocorticoid action is mediated via glucocorticoid receptors (GRs). This study aims to define any clinical implications of GR expression in ovarian cancer to further the debate. Archived tissue samples from patients with histologically confirmed ovarian cancer were analyzed for GR expression and evaluated by immunohistochemistry and immunoreactive score. The results were related to the patients' overall survival. Kaplan-Meier survival and residual survival analyses gave no evidence that GR expression had any prognostic value in the 85 cases studied. No evidence of poorer survival was found in a small subset of GR-positive patients who received GC treatment. Glucocorticoid receptor expression had no prognostic impact in our study. However, GC (cortisol) is being produced continuously by the body, which may have stimulated GR-positive ovarian cancer cells. Our finding does not exclude the possibility that long-term GC treatment has adverse effects, and it should also be emphasized that treatment duration, dosage and dosing regimens, as well as the choice of an appropriate GC and the mode of application, determine the risks and benefits. Our study showed no evidence against using GC for antiemetic prophylaxis in ovarian carcinomas.     

Rational approaches to natural-product-based drug design

Natural-product-based drug discovery has encountered significant challenges during the past decade. In recent years the pharmaceutical industry has placed low emphasis on natural-product-based drug discovery efforts because of an increasing reliance on newer technologies, such as combinatorial synthesis and high-throughput screening, and their associated approaches to drug discovery. However, recent natural-product-based lead-identifying strategies have successfully and rapidly integrated rational approaches that exploit and evolve the structural diversity provided by nature. These rational approaches include the application of structure- and ligand-based design, relationship building between biosynthetic enzymes and targets as well as within the target and natural product scaffold space, and biology-oriented synthesis-guided library design. This review focuses on the recent clinical and preclinical development of natural-product-based compounds derived from these rational approaches, and is organized according to disease areas as well as novel concepts that may provide a rational basis for future developments.     

Stress-induced in vitro apoptosis of native human acute myelogenous leukemia (AML) cells shows a wide variation between patients and is associated with low BCL-2:Bax ratio and low levels of heat shock protein 70 and 90

Spontaneous in vitro apoptosis reflects a true biological heterogeneity between patients which has to be considered when in vitro models are used to study regulation of apoptosis in native human AML cells. Even though the balance between pro- and anti-apoptotic signaling seems to have a prognostic impact in AML, the possible clinical relevance of spontaneous apoptosis remains to be clarified. High apoptosis/low viability was associated with low levels of heat shock proteins 70 and 90 as well as low Bcl-2:Bax ratio for patients heterogeneous with regard to morphology, membrane molecule expression, genetic abnormalities and response to therapy.