Biomedical subjects

Diana Anderson

Publications and source records attributed to Diana Anderson.

At least 19 recordsLinked to original sources

PubMed2006

Genotoxic and antigenotoxic properties of selenium compounds in the in vitro micronucleus assay with human whole blood lymphocytes and TK6 lymphoblastoid cells.

Selenium is known to possess both genotoxic and antigenotoxic properties. In the present study, we have evaluated the genotoxicity and antigenotoxicity of three selenium compounds (sodium selenate, sodium selenite and selenous acid) by measuring in vitro micronucleus induction. Assays were conducted in whole blood lymphocytes and in the TK6 lymphoblastoid cell line, with and without co-treatment with potassium dichromate, a known genotoxic compound. In general, the compounds were more active in TK6 cells than they were in blood lymphocytes. Only 1 microM selenous acid increased the frequency of binucleated cells containing micronuclei (BNMN) in blood lymphocytes, while all three selenium compounds increased BNMN in TK6 cells. In addition, combinations of selenous acid and potassium dichromate resulted in lower frequencies of BNMN than potassium dichromate alone in blood lymphocytes, while combinations of sodium selenate and potassium dichromate produced lower frequencies of BNMN than potassium dichromate alone in TK6 cells. The concentrations of selenium compounds that were used, in combination with the medium components and the biological physiology of the whole blood lymphocytes and TK6 cells, could have affected the redox potential of the compounds, switching the chemicals from a pro-oxidant to antioxidant status and vice-versa. The lower activities of the compounds in blood lymphocytes may be due to the protective effects of blood components. The results indicate that the genotoxic and antigenotoxic properties of selenium compounds are highly dependent upon the conditions under which they are evaluated.

Adult↗

PubMed2006

The responses of lymphocytes from Asian and Caucasian diabetic patients and non-diabetics to hydrogen peroxide and sodium nitrite in the Comet assay.

Numerous factors may influence the incidence of diabetes in the population. The production of reactive oxygen species (ROS) is elevated in diabetes patients. Based on the reported involvement of reactive species and nitrate/nitrite in diabetes, this present study has examined in the alkaline Comet assay, the effect of different levels of NaNO(2) in the presence of the oxygen radical generating agent, hydrogen peroxide (H(2)O(2)). Peripheral lymphocytes from diabetic and non-diabetic Caucasians and Asians of both sexes were studied in vitro. Endogenous factors (e.g., sex, age, body mass index-BMI) and exogenous factors (lifestyle factors e.g., smoking and drinking habits, diet) were taken into account. A preliminary study in two individuals showed that DNA damage remained constant over a wide dose range of NaNO(2) (1-75mM), but when H(2)O(2) was added at a constant concentration of 50microM per dose of NaNO(2), there was an increase in DNA damage corresponding with the varying levels of NaNO(2) investigated. This was also seen with the 44 individuals (non-diabetic, n=24; type 1 diabetic, n=11; type 2 diabetic, n=9) investigated. NaNO(2) was capable of inducing a significant level of DNA damage in lymphocytes (p<0.001), but only with the addition of H(2)O(2). When levels of DNA damage were analysed in terms of the different variables there were few significant differences in damage between diabetic and non-diabetic subjects, or other sub-population groups, and no statistically significant differences in susceptibility were observed between subject covariates using regression techniques.

Adolescent↗

PubMed2006

An in vitro model to study chemoresistance in non-Hodgkin's lymphoma patients over-expressing mutant p53.

INTRODUCTION: P-Glycoprotein plays a major role in regulating the concentration of chemotherapeutic agents inside the cytoplasm of normal and cancerous cells. The present in vitro study has primarily focused on the evaluation of the chemosensitising drug model, verapamil, as a P-glycoprotein antagonist not only to overcome chemoresistance in non-Hodgkin's lymphoma (NHL) chemoresistant cells (p53(+) i.e. over-expressing p53 mutant protein NHL cells) but also to evaluate and suggest the use of the single cell gel electrophoresis (SCGE) assay in the clinical setting. METHODS: Leucocytes from CHOP-chemoresistant NHL patients (p53(+) cells) were examined in the SCGE assay. Altered levels of DNA damage (tail moments) were determined by the assay not only in the leucocytes from clinical samples, but also in the Raji cell sub-lines (NHL model) which are also over-expressing p53 mutant protein. In addition, as a comparison, P-glycoprotein was examined in normal human leucocytes and Raji cells. RESULTS: Verapamil increased the tail moments induced by doxorubicin in all cell types over-expressing p53 mutant protein. DISCUSSION: The assay was successful for evaluating P-glycoprotein regulation. This suggests the applicability at the cellular level of the method as suitable for use in the clinical setting since it is reliable and could be used pre-clinically or perhaps instead of or alongside clinical trails. Cells from patients with a chemoresistant disease state or whose disease relapses subsequently could be treated with such novel experimental therapies in vitro to determine the necessity for the individual administration of a P-glycoprotein antagonist.

ATP Binding Cassette Transporter, Subfamily B, Mem↗

PubMed2006

Modulation of the cytotoxicity and genotoxicity of the drinking water disinfection byproduct lodoacetic acid by suppressors of oxidative stress.

Drinking water disinfection byproducts (DBPs) are generated by the chemical disinfection of water and may pose a hazard to the public health. Previously we demonstrated that iodoacetic acid was the most cytotoxic and genotoxic DBP analyzed in a mammalian cell system. Little is known of the mechanisms of its genotoxicity. The involvement of oxidative stress in the toxicity of iodoacetic acid was analyzed with the antioxidants catalase and butylated hydroxyanisole (BHA). lodoacetic acid toxicity was quantitatively measured with and without antioxidants in Salmonella typhimurium strain TA100 and with Chinese hamster ovary (CHO) cells. The endpoints included cytotoxicity in S. typhimurium or in CHO cells, mutagenicity in S. typhimurium, and genotoxicity in CHO cells. Neither catalase nor BHA reduced the level of iodoacetic acid induced cytotoxicity in S. typhimurium. In CHO cells neither antioxidant caused a significant reduction in iodoacetic acid induced cytotoxicity. However, in S. typhimurium, BHA or catalase reduced the mutagenicity of iodoacetic acid by 33.5 and 26.8%, respectively. Likewise, BHA or catalase reduced iodoacetic acid induced genomic DNA damage by 86.5 and 42%, respectively. These results support the hypothesis that oxidative stress is involved in the induction of genotoxicity and mutagenicity by iodoacetic acid.

Animals↗

PubMed2006

In vitro studies of DNA damage and its repair in cells from NHL patients with different p53 mutant protein status, resistant (p53(+)) and sensitive (p53(-)) to cancer chemotherapy.

INTRODUCTION: Resistance to an anthracycline-based regimen, such as CHOP, constitutes a problem for curing non-Hodgkin's lymphoma (NHL) patients. Chemoresistance in the clinic manifests itself as a lack of response to treatment or regrowth of a tumour after an initial response. METHODS: In this study, lymphocytes from NHL patients were treated with hydrogen peroxide (H(2)O(2)), a free radical generating model agent, and ethyl methanesulfonate (EMS), a model alkylating agent, to induce DNA damage which was evaluated by SCGE. This study assessed whether or not there were any differences in the patterns of damage and repair between cells from patients with p53 mutant protein abnormalities, i.e. over-expression (p53(+)) not responding to the CHOP regimen and patients responding to the CHOP regimen without p53 protein abnormalities (p53(-)) by comparison with control individuals (wild-type). An NHL cell line model [Raji TK(+) (mex(+)) and TK(-) (mex(-))] with p53 over-expression was also investigated. RESULTS: Results showed that frozen/thawed samples from healthy people were not suitable for use in repair studies, whilst fresh samples or samples incubated for 20 h at room temperature could be used. Tumour cells were more sensitive to damage than control cells. After treatment with H(2)O(2), cells from fresh or incubated blood showed a similar repair capacity. After treatment with EMS, there was a difference between repair in resistant and sensitive cells. DISCUSSION: These results suggest that the repair process is a useful cellular biomarker for investigating chemoresistance. The lack of repair in p53(+) cells may correlate with a low level of MGMT, since there was no repair in the Raji TK(-) cells lacking this enzyme.

Adult↗

PubMed2006

Autophagy promotes tumor cell survival and restricts necrosis, inflammation, and tumorigenesis.

Defective apoptosis renders immortalized epithelial cells highly tumorigenic, but how this is impacted by other common tumor mutations is not known. In apoptosis-defective cells, inhibition of autophagy by AKT activation or by allelic disruption of beclin1 confers sensitivity to metabolic stress by inhibiting an autophagy-dependent survival pathway. While autophagy acts to buffer metabolic stress, the combined impairment of apoptosis and autophagy promotes necrotic cell death in vitro and in vivo. Thus, inhibiting autophagy under conditions of nutrient limitation can restore cell death to apoptosis-refractory tumors, but this necrosis is associated with inflammation and accelerated tumor growth. Thus, autophagy may function in tumor suppression by mitigating metabolic stress and, in concert with apoptosis, by preventing death by necrosis.

Animals↗

PubMed2006

Estrogens can contribute to hydrogen peroxide generation and quinone-mediated DNA damage in peripheral blood lymphocytes from patients with vitiligo.

To date there is ample in vivo and in vitro evidence for increased epidermal and systemic hydrogen peroxide (H(2)O(2)) levels in vitiligo, which can be reduced with a topical application of a pseudocatalase-K.U. Schallreuter (PC-KUS) leading to the recovery of epidermal catalase levels as well as other enzymes in peripheral blood cells. Recently, the generation of H(2)O(2) by oxidative metabolism of estrogens and other aromatic steroids was documented. Therefore, it was tempting to follow estrogen-generated H(2)O(2) and its possible effect on DNA damage in peripheral blood lymphocytes from patients with vitiligo before and after the reduction of epidermal H(2)O(2) with pseudocatalase PC-KUS compared to controls. For this purpose, 20 Caucasian patients were grouped in treated responders (group A, n=11) and untreated active/acute disease (group B, n=9) and compared to Caucasian healthy controls (group C, n=7). Consequently, epidermal catalase protein expression in full skin biopsies was assessed using immunofluorescence labelling together with determination of basal H(2)O(2) levels in peripheral blood lymphocytes. To test the influence of estrogen on H(2)O(2) generation and DNA damage, freshly prepared peripheral blood lymphocytes from all three groups were used for the alkaline comet assay in the presence and absence of catalase. The results of this study demonstrated that reduction of epidermal H(2)O(2) leads to both increased epidermal catalase protein expression as well as decreased H(2)O(2) concentrations in lymphocytes. Moreover, a direct estrogen-mediated DNA damage was identified in both patient groups, which was absent in healthy controls. This effect was not abolished by catalase pointing to direct quinone-mediated DNA damage by estrogens in peripheral blood lymphocytes in vitiligo.

Adult↗

PubMed2005

Male-mediated developmental toxicity.

In recent years, the public has become more aware that exposure of males to certain agents can adversely affect their offspring and cause infertility and cancer. The hazards associated with exposure to ionising radiation have been recognised for nearly a century, but interest was aroused when a cluster of leukaemia cases was identified in young children living in Seascale, close to the nuclear processing plant at Sellafield in West Cumbria. There was a civil court case on behalf of two of the alleged victims of paternal irradiation at Seascale against British Nuclear Fuels. The case foundered on "the balance of probabilities". Nevertheless, there was support for paternal exposure from Japanese experimental X-ray studies in mice. The tumours were clearly heritable as shown by F2 transmission. Also, effects of a relatively non-toxic dose of radiation (1Gy) on cell proliferation transmitted to the embryo were manifested in the germ line of adult male mice even after two generations. In addition in humans, smoking fathers appear to give rise to tumours in the F(1) generation. Using rodent models, developmental abnormalities/congenital malformations and tumours can be studied after exposure of males in an extended dominant lethal assay and congenital malformations can be determined which have similar manifestations in humans. The foetuses can also be investigated for skeletal malformations and litters can be allowed to develop to adulthood when tumours, if present, can be observed. Karyotype analysis can be performed on foetuses and adult offspring to determine if induced genetic damage can be transmitted. Using this study design, cyclophosphamide, 1,3-butadiene and urethane have been examined and each compound produced positive responses: cyclophosphamide in all endpoints examined, 1,3-butadiene in some and urethane only produced liver tumours in F(1) male offspring. This suggests the endpoints are determined by independent genetic events. The results from heritable studies with 1,3-butadiene have been used in the parallelogram approach to determine a risk assessment for the germ cells in man.

Animals↗

PubMed2005

Cadmium chloride-induced DNA and lysosomal damage in a hepatoma cell line.

Cadmium is a toxic metal and no uniform mechanism of toxicity has so far been proposed. The aim of this study was to investigate the biochemical effects of cadmium chloride in a rat hepatoma cell line (HTC cells) and the cellular events mediating DNA damage. HTC cells were exposed to various concentrations of cadmium chloride for 5 and 8 h and lysosomal damage was assessed with the neutral red assay (NR) and fluorescence microscopy. Mitochondrial integrity was assessed from ATP levels and DNA damage determined with the single cell gel electrophoresis/comet assay. The formation of reactive oxygen species (ROS) was also determined under the same experimental conditions with the dichlorofluorescein assay. Cytotoxicity was assessed with the LDH leakage assay and the levels of glutathione were measured and correlated with the other effects. The results indicate that lysosomal damage occurs at a lower concentration of cadmium chloride (20 microM) than DNA damage (500 microM) in HTC cells. The latter effect was accompanied by an increase of reactive oxygen species without any significant LDH leakage whereas lysosomal damage was significant as determined by the neutral red assay and confirmed with fluorescence microscopy. The effect of CdCl2 on mitochondria and glutathione levels were observed at concentrations or incubation times higher than the ones required to induce lysosomal damage. The data suggest that DNA damage may be due to the formation of reactive oxygen species. It is possible that cadmium induced lysosomal damage is an earlier event than DNA damage and can mediate other cellular events that lead to cell death.

Adenosine Triphosphate↗

PubMed2004

Hypoxia and defective apoptosis drive genomic instability and tumorigenesis.

Genomic instability is a hallmark of cancer development and progression, and characterizing the stresses that create and the mechanisms by which cells respond to genomic perturbations is essential. Here we demonstrate that antiapoptotic BCL-2 family proteins promoted tumor formation of transformed baby mouse kidney (BMK) epithelial cells by antagonizing BAX- and BAK-dependent apoptosis. Cell death in vivo correlated with hypoxia and induction of PUMA (p53 up-regulated modulator of apoptosis). Strikingly, carcinomas formed by transformed BMK cells in which apoptosis was blocked by aberrant BCL-2 family protein function displayed prevalent, highly polyploid, tumor giant cells. Examination of the transformed BMK cells in vivo revealed aberrant metaphases and ploidy changes in tumors as early as 9 d after implantation, which progressed in magnitude during the tumorigenic process. An in vitro ischemia system mimicked the tumor microenvironment, and gain of BCL-2 or loss of BAX and BAK was sufficient to confer resistance to apoptosis and to allow for accumulation of polyploid cells in vitro. These data suggest that in vivo, even in cells in which p53 function is compromised, apoptosis is an essential response to hypoxia and ischemia in the tumor microenvironment and that abrogation of this response allows the survival of cells with abnormal genomes and promotes tumorigenesis.

Animals↗

PubMed2004

Modulation by flavonoids of DNA damage induced by estrogen-like compounds.

Reactive oxygen species (ROS) are produced by a wide variety of exogenous chemicals and metabolic processes and cause a broad spectrum of damage to biological systems. As a consequence, ROS react with DNA, among many other biological targets, disrupting its structure and functionality. Estrogen-like compounds mediate DNA damage by ROS generation, implying that their effects can be modulated by antioxidants such as catalase, superoxide dismutase, and vitamin C. We examined DNA damage in human lymphocytes and sperm after treatment with four estrogen-like compounds (beta-estradiol, diethylstilbestrol, daidzein, and genistein) and its modulation by flavonoids (quercetin and kaempferol) using the Comet assay. The results indicated that quercetin and kaempferol reduced the DNA damage produced in sperm and lymphocytes by the four estrogenic compounds. The flavonoids also reduced the DNA damage induced by hydrogen peroxide, which was used as a positive control. Our results demonstrate that the antioxidant properties of flavonoids can protect the integrity of human sperm and lymphocyte DNA from ROS induced by estrogenic compounds.

Antineoplastic Agents↗

PubMed2004

Antioxidants modulate thyroid hormone- and noradrenaline-induced DNA damage in human sperm.

The genotoxic effects of steroidal oestrogens are probably brought about by metabolic changes in their phenolic groups accompanied by the generation of quinones and reactive oxygen species. Although non-steroidal oestrogens and related compounds have not been thoroughly investigated for genotoxicity, some of them also contain phenolic groups that could be involved in redox cycling. Therefore, the aim of the present study was to evaluate the possible DNA-damaging effects of the thyroid hormones triiodothyronine (T3) and L-thyroxine sodium salt (T4) and the neurotransmitter noradrenaline (NA) in human sperm using the Comet assay. They were compared with diethylstilboestrol (DES), a steroidal oestrogen, as a positive control. After dose-response studies, doses of 80 microM T3, 80 microM T4, 300 microM NA and 175 microM DES, which produced DNA damage but retained good cell viability, were chosen for further experiments with the antioxidant catalase and the flavonoids kaempferol and quercetin. Since the scavenging enzyme catalase reduced the DNA-damaging effects of T3, T4 and NA, it can be surmised that these compounds under these conditions induced DNA damage mainly via the production of reactive oxygen species. This was further confirmed by the inhibitory responses produced by the flavonoids, which are known to have antioxidant effects. Therefore, the mechanism of mutagenic action of both steroidal and non-steroidal compounds imply the creation of oxidative stress and subsequent DNA damage due to reactive oxygen species and possibly due to reactive hormone derivatives created during their redox cycling.

Adult↗

PubMed2003

Sensitivity of different thalassaemia genotypes to food mutagens in the Comet assay.

Thalassaemia is a heterogeneous group of inherited anaemias, characterised by a reduction or total absence of one or more of the globin chains of haemoglobin. Individuals with thalassaemia major require regular blood transfusions in order to maintain their haemoglobin concentration at an appropriate level. An essential treatment in parallel with transfusions is iron chelation therapy to remove excess iron deposited in tissues from the transfused blood. The high iron levels in these patients make free oxygen radicals accessible, for example, through Fenton-type chemistry, and generate superoxide and hydroxyl radicals. Increased oxygen radical capacity is known to be associated with cancer and ageing. In a previous study, it has been shown that peripheral blood lymphocytes isolated from a sickle/beta thal double heterozygote-sickle phenotype thalassaemia patient, who was not undergoing chelation therapy, showed increased sensitivity to the effects of oxygen radicals and iron salts by comparison with lymphocytes from normal controls. Furthermore, in a later study, this patient also showed increased sensitivity to the dietary food mutagen 3-amino-1-methyl-5H-pyridol(4,3-b)indole (Trp-P-2) when compared to the control. The present study, therefore, investigated whether the above observation could be duplicated using different food mutagens in different thalassaemia genotypes. The effect of the food mutagens 2-amino-2-methylimidazolo(4,5-f)quinolone (IQ) and 2-amino-1-methyl-6-phenylimidazol(4,5-b)pyridine (PhIP) on lymphocytes of three different thalassaemia patients, a beta-thalassaemia major, a beta-thalassaemia/Hb E, and an alpha-thalassaemia trait with a 3.7-kb deletion, who were not undergoing chelation therapy were investigated using the Comet assay. All three thalassaemia genotypes showed increased sensitivity to both IQ and PhIP in comparison to the control, although with PhIP at the highest two concentrations (50 and 75 microM) the differences monitored with the alpha-thalassaemia trait were found not to be statistically significant (P > 0.05).

Child, Preschool↗

PubMed2003

Oxygen-induced DNA damage in freshly isolated brain cells compared with cultured astrocytes in the Comet assay.

Brain cells are continuously exposed to reactive oxygen species generated by oxidative metabolism, and in certain pathological conditions defence mechanisms against oxygen radicals may be weakened and/or overwhelmed. DNA is a potential target for oxidative damage, and genomic damage can contribute to neuropathogenesis. It is important, therefore, to identify tools for the quantitative analysis of DNA damage in models of neurological disorders. The aim of this study was to compare the susceptibility of DNA to oxidative stress in cells freshly dissociated from the mouse brain, to that in cultured brain cells. Both primary cultures and a continuous cell line of astrocytes were considered. All cells were treated by xanthine/xanthine oxidase, a superoxide generator or hydrogen peroxide, applied alone or in the presence of the oxygen radical scavengers, superoxide dismutase, catalase, or ascorbic acid. DNA damage, quantified with the Comet assay, was consistent in all the different cell preparations exposed to oxidative stress, and was attenuated in similar ways by superoxide dismutase and catalase, scavengers of superoxide anion and hydrogen peroxide, respectively. The results with ascorbic acid were more variable, presumably because this compound may switch from anti- to pro-oxidant status depending on its concentration and other experimental conditions. Overall, similar responses were found in freshly dissociated and cultured brain cells. These results suggest that the Comet assay can be directly applied to cells freshly dissociated from the brain of rodents, including models of neurological disorders, such as stroke models and animals with targeted mutations that mimic human diseases.

Animals↗

PubMed2003

Antigenotoxic properties of selenium compounds on potassium dichromate and hydrogen peroxide.

Selenium is an environmental metal that occurs ubiquitously and is produced throughout the world for various industrial activities. Selenium has been reported to have anticarcinogenic and preventive effects in clinical and epidemiological studies. Selenium supplements can inhibit chemically-induced tumours. From the viewpoint of genotoxicity, selenium has not been adequately studied and an IARC review concluded that there were not sufficient data to consider it a carcinogen for man. In contrast, hexavalent chromium is classified as a known respiratory carcinogen producing DNA damage through free oxygen radicals. In the present study, a collaborative study has been carried out to evaluate the genotoxicity of selenium compounds and their possible interactions with potassium dichromate and hydrogen peroxide. Thus, in laboratory 1 (U.K.), the genotoxic effects of three selenium compounds were examined. Sodium selenate, sodium selenite, and selenous acid were investigated in the Ames test using strain TA102 and in the Comet assay using human lymphocytes, and also investigated for their interaction with potassium dichromate. In the Ames test, it was shown that potassium dichromate produced a highly mutagenic response, whilst the three selenium compounds did not. In combination, sodium selenate reduced the genotoxicity of potassium dichromate, but sodium selenite and selenous acid had no effect. In the Comet assay, potassium dichromate induced DNA damage, but so did the selenium compounds. In combination with potassium dichromate, however, only sodium selenate reduced its effect, whereas sodium selenite and selenous acid exacerbated DNA damage. In laboratory 2 (Spain), in the TK6 lymphoblastoid cell line, the Comet assay showed that sodium selenite was non-genotoxic, while potassium dichromate and hydrogen peroxide induced DNA damage. It was also shown that sodium selenite did not decrease the genotoxicity of potassium dichromate or hydrogen peroxide when administered as a pre-treatment or at the same time, or when potassium dichromate and sodium selenite treatments were for different time periods. Thus, only sodium selenate has shown antigenotoxic properties against potassium dichromate in the Ames test and in human lymphocytes in the Comet assay.

Comet Assay↗

PubMed2003

Evaluation of EMS-induced DNA damage in the single cell gel electrophoresis (Comet) assay and with flow cytometric analysis of micronuclei.

Toxic agents in the environment pose serious threats to ecosystems and to the public health. The single cell gel electrophoresis (SCGE) or Comet assay quantitatively measures genomic damage as DNA strand breaks. The micronucleus (MCN) test is an established assay that measures chromosomal damage. Micronuclei are formed from chromosome fragments or from whole chromosomes that have not undergone mitosis properly. This test is usually conducted microscopically. However, micronuclei can also be analyzed using flow cytometry. Chinese hamster ovary (CHO) cells were exposed to ethylmethanesulfonate (EMS), for 4 h in a total volume of 25 microl. These cells were immediately analyzed for genomic DNA damage by SCGE. In concurrent parallel experiments, CHO cells were treated with EMS in 6-well plates for 4 h, the cells were washed and fresh medium was added. The cells were allowed to grow for 45 to 48 h to express micronuclei. The data demonstrated that both DNA strand breaks and micronuclei were induced in a significant and concentration-dependent manner. There was a significant and high correlation (r = 0.91; P < or = 0.001) between the acute induction of DNA strand breaks and the subsequent generation of micronuclei. These data indicate that using molecular and computer technologies, the genotoxic impact of toxic and environmental agents can be rapidly and comprehensively evaluated in mammalian cell systems.

Animals↗

PubMed2003

Genotoxicity studies on DNA-interactive telomerase inhibitors with application as anti-cancer agents.

Telomerase-targeted strategies have aroused recent interest in anti-cancer chemotherapy, because DNA-binding drugs can interact with high-order tetraplex rather than double-stranded (duplex) DNA targets in tumour cells. However, the protracted cell-drug exposure times necessary for clinical application require that telomerase inhibitory efficacy must be accompanied by both low inherent cytotoxicity and the absence of mutagenicity/genotoxicity. For the first time, the genotoxicity of a number of structurally diverse DNA-interactive telomerase inhibitors is examined in the Ames test using six Salmonella typhimurium bacterial strains (TA1535, TA1537, TA1538, TA98, TA100, and TA102). DNA damage induced by each agent was also assessed using the Comet assay with human lymphocytes. The two assay procedures revealed markedly different genotoxicity profiles that are likely to reflect differences in metabolism and/or DNA repair between bacterial and mammalian cells. The mutational spectrum for a biologically active fluorenone derivative, shown to be mutagenic in the TA100 strain, was characterised using a novel and rapid assay method based upon PCR amplification of a fragment of the hisG46 allele, followed by RFLP analysis. Preliminary analysis indicates that the majority (84%) of mutations induced by this compound are C --> A transversions at position 2 of the missense proline codon of the hisG46 allele. However, despite its genotoxic bacterial profile, this fluorenone agent gave a negative response in the Comet assay, and demonstrates how unwanted systemic effects (e.g., cytotoxicity and genotoxicity) can be prevented or ameliorated through suitable molecular fine-tuning of a candidate drug in targeted human tumour cells.

Antineoplastic Agents↗

PubMed2003

Modulation by flavonoids of the effects of a food mutagen in different thalassaemia genotypes in the Comet assay.

Thalassaemia is an inherited group of disorders caused by a reduction or total absence of one or more of the globin chains of the haemoglobin molecule. It has been shown that lymphocytes isolated from a sickle/beta thal double heterozygote-sickle phenotype patient showed increased sensitivity to the dietary food mutagen 3-amino-1-methyl-5H-pyridol(4,3-b)indole (Trp-P-2) when compared to the control. Furthermore, when a combination of Trp-P-2 with either quercitin or kaempferol was compared, the responses to Trp-P-2 were reduced to untreated control levels at the highest doses of quercitin and kaempferol. It has now been shown that using the food mutagens 2-amino-2-methylimidazolo(4,5-f)quinolone (IQ) and 2-amino-1-methyl-6-phenylimidazol(4,5-b)pyridine (PhIP) on lymphocytes of three different thalassaemia patients, a beta-thalassaemia major, a beta-thalassaemia/Hb E, and an alpha-thalassaemia trait with a 3.7-kb deletion, similar increased sensitivity could also be demonstrated. The present study investigated whether the modulatory effects of the flavonoids could be demonstrated in lymphocytes isolated from a beta-thalassaemia major and a beta-thalassaemia/Hb E patient. Lymphocytes from both a beta-thalassaemia major and beta-thalassaemia/Hb E patient showed increased sensitivity to PhIP when compared to the normal control. When a combination of PhIP and either quercitin or kaempferol was used, a reduction in the responses was seen, and at the highest doses of quercitin and kaempferol the responses were reduced to near untreated control levels and were significantly different when compared to PhIP alone (P < 0.05). It was concluded that lymphocytes from different thalassaemia genotypes showed increased sensitivity to different dietary food mutagens compared to normal lymphocytes and that flavonoids such as quercitin and kaempferol modulated the effects of these food mutagens in an antigenotoxic/antioxidant manner.

Child, Preschool↗