Syrian hamster convalescence from prototype SARS-CoV-2 confers measurable protection against the attenuated disease caused by the Omicron variant.

The mutation profile of the SARS-CoV-2 Omicron (lineage BA.1) variant posed a concern for naturally acquired and vaccine-induced immunity. We investigated the ability of prior infection with an early SARS-CoV-2 ancestral isolate (Australia/VIC01/2020, VIC01) to protect against disease caused by BA.1. We established that BA.1 infection in naïve Syrian hamsters resulted in a less severe disease than a comparable dose of the ancestral virus, with fewer clinical signs including less weight loss. We present data to show that these clinical observations were almost absent in convalescent hamsters challenged with the same dose of BA.1 50 days after an initial infection with ancestral virus. These data provide evidence that convalescent immunity against ancestral SARS-CoV-2 is protective against BA.1 in the Syrian hamster model of infection. Comparison with published pre-clinical and clinical data supports consistency of the model and its predictive value for the outcome in humans. Further, the ability to detect protection against the less severe disease caused by BA.1 demonstrates continued value of the Syrian hamster model for evaluation of BA.1-specific countermeasures.

Natural product-derived antitumor compound phenethyl isothiocyanate inhibits mTORC1 activity via TSC2.

Phenethyl isothiocyanate (1) is a natural dietary phytochemical with cytostatic, cytotoxic, and antitumor activity. The effects of 1 were investigated on the activity of mTOR, a kinase that enhances the translation of many RNAs encoding proteins critical for cancer cell growth, including the angiogenesis regulator HIF1α. Compound 1 effectively blocked HIF1α RNA translation in MCF7 breast cancer cells, and this was associated with reduced phosphorylation of 4E-BP1 and p70 S6K, well-characterized downstream substrates of the mTOR-containing mTORC1 complex. Compound 1 also inhibited mTORC1 activity in mouse embryonic fibroblasts (MEFs). The 1-mediated inhibition of mTORC1 activity appeared to be independent of the upstream regulators PTEN, AKT, ERK1/2, and AMPK. By contrast, 1-mediated inhibition of mTORC1 activity was dependent on the presence of TSC2, part of a complex that regulates mTORC1 activity negatively. TSC2-deficient MEFs were resistant to 1-mediated inhibition of p70 S6K phosphorylation. TSC2-deficient MEFs were also partially resistant to 1-mediated growth inhibition. Overall, the present results confirm that 1 inhibits mTORC1 activity. This is dependent on the presence of TSC2, and inhibition of mTORC1 contributes to optimal 1-induced growth inhibition. Inhibition of RNA translation may be an important component of the antitumor effects of phenethyl isothiocyanate.

Prophylactic intranasal administration of a TLR2/6 agonist reduces upper respiratory tract viral shedding in a SARS-CoV-2 challenge ferret model.

BACKGROUND: The novel human coronavirus SARS-CoV-2 is a major ongoing global threat with huge economic burden. Like all respiratory viruses, SARS-CoV-2 initiates infection in the upper respiratory tract (URT). Infected individuals are often asymptomatic, yet highly infectious and readily transmit virus. A therapy that restricts initial replication in the URT has the potential to prevent progression of severe lower respiratory tract disease as well as limiting person-to-person transmission. METHODS: SARS-CoV-2 Victoria/01/2020 was passaged in Vero/hSLAM cells and virus titre determined by plaque assay. Challenge virus was delivered by intranasal instillation to female ferrets at 5.0 × 106 pfu/ml. Treatment groups received intranasal INNA-051, developed by Ena Respiratory. SARS-CoV-2 RNA was detected using the 2019-nCoV CDC RUO Kit and QuantStudio™ 7 Flex Real-Time PCR System. Histopathological analysis was performed using cut tissues stained with haematoxylin and eosin (H&E). FINDINGS: We show that prophylactic intra-nasal administration of the TLR2/6 agonist INNA-051 in a SARS-CoV-2 ferret infection model effectively reduces levels of viral RNA in the nose and throat. After 5 days post-exposure to SARS-CoV-2, INNA-051 significantly reduced virus in throat swabs (p=<0.0001) by up to a 24 fold (96% reduction) and in nasal wash (p=0.0107) up to a 15 fold (93% reduction) in comparison to untreated animals. INTERPRETATION: The results of our study support clinical development of a therapy based on prophylactic TLR2/6 innate immune activation in the URT, to reduce SARS-CoV-2 transmission and provide protection against COVID-19. FUNDING: This work was funded by Ena Respiratory, Melbourne, Australia.

Dose-dependent response to infection with SARS-CoV-2 in the ferret model and evidence of protective immunity.

There is a vital need for authentic COVID-19 animal models to enable the pre-clinical evaluation of candidate vaccines and therapeutics. Here we report a dose titration study of SARS-CoV-2 in the ferret model. After a high (5 × 106 pfu) and medium (5 × 104 pfu) dose of virus is delivered, intranasally, viral RNA shedding in the upper respiratory tract (URT) is observed in 6/6 animals, however, only 1/6 ferrets show similar signs after low dose (5 × 102 pfu) challenge. Following sequential culls pathological signs of mild multifocal bronchopneumonia in approximately 5-15% of the lung is seen on day 3, in high and medium dosed groups. Ferrets re-challenged, after virus shedding ceased, are fully protected from acute lung pathology. The endpoints of URT viral RNA replication & distinct lung pathology are observed most consistently in the high dose group. This ferret model of SARS-CoV-2 infection presents a mild clinical disease.

ChAdOx1 nCoV-19 protection against SARS-CoV-2 in rhesus macaque and ferret challenge models.

Vaccines against SARS-CoV-2 are urgently required, but early development of vaccines against SARS-CoV-1 resulted in enhanced disease after vaccination. Careful assessment of this phenomena is warranted for vaccine development against SARS CoV-2. Here we report detailed immune profiling after ChAdOx1 nCoV-19 (AZD1222) and subsequent high dose challenge in two animal models of SARS-CoV-2 mediated disease. We demonstrate in rhesus macaques the lung pathology caused by SARS-CoV-2 mediated pneumonia is reduced by prior vaccination with ChAdOx1 nCoV-19 which induced neutralising antibody responses after a single intramuscular administration. In a second animal model, ferrets, ChAdOx1 nCoV-19 reduced both virus shedding and lung pathology. Antibody titre were boosted by a second dose. Data from these challenge models on the absence of enhanced disease and the detailed immune profiling, support the continued clinical evaluation of ChAdOx1 nCoV-19.

Comparison of rhesus and cynomolgus macaques as an infection model for COVID-19.

A novel coronavirus, SARS-CoV-2, has been identified as the causative agent of the current COVID-19 pandemic. Animal models, and in particular non-human primates, are essential to understand the pathogenesis of emerging diseases and to assess the safety and efficacy of novel vaccines and therapeutics. Here, we show that SARS-CoV-2 replicates in the upper and lower respiratory tract and causes pulmonary lesions in both rhesus and cynomolgus macaques. Immune responses against SARS-CoV-2 are also similar in both species and equivalent to those reported in milder infections and convalescent human patients. This finding is reiterated by our transcriptional analysis of respiratory samples revealing the global response to infection. We describe a new method for lung histopathology scoring that will provide a metric to enable clearer decision making for this key endpoint. In contrast to prior publications, in which rhesus are accepted to be the preferred study species, we provide convincing evidence that both macaque species authentically represent mild to moderate forms of COVID-19 observed in the majority of the human population and both species should be used to evaluate the safety and efficacy of interventions against SARS-CoV-2. Importantly, accessing cynomolgus macaques will greatly alleviate the pressures on current rhesus stocks.

In vivo modulation of 4E binding protein 1 (4E-BP1) phosphorylation by watercress: a pilot study.

Dietary intake of isothiocyanates (ITC) has been associated with reduced cancer risk. The dietary phenethyl ITC (PEITC) has previously been shown to decrease the phosphorylation of the translation regulator 4E binding protein 1 (4E-BP1). Decreased 4E-BP1 phosphorylation has been linked to the inhibition of cancer cell survival and decreased activity of the transcription factor hypoxia-inducible factor (HIF), a key positive regulator of angiogenesis, and may therefore contribute to potential anti-cancer effects of PEITC. In the present study, we have investigated the in vitro and in vivo effects of watercress, which is a rich source of PEITC. We first demonstrated that, similar to PEITC, crude watercress extracts inhibited cancer cell growth and HIF activity in vitro. To examine the effects of dietary intake of watercress, we obtained plasma and peripheral blood mononuclear cells following the ingestion of an 80 g portion of watercress from healthy participants who had previously been treated for breast cancer. Analysis of PEITC in plasma samples from nine participants demonstrated a mean maximum plasma concentration of 297 nm following the ingestion of watercress. Flow cytometric analysis of 4E-BP1 phosphorylation in peripheral blood cells from four participants demonstrated significantly reduced 4E-BP1 phosphorylation at 6 and 8 h following the ingestion of watercress. Although further investigations with larger numbers of participants are required to confirm these findings, this pilot study suggests that flow cytometry may be a suitable approach to measure changes in 4E-BP1 phosphorylation following the ingestion of watercress, and that dietary intake of watercress may be sufficient to modulate this potential anti-cancer pathway.

Acellular Pertussis Vaccines Induce Anti-pertactin Bactericidal Antibodies Which Drives the Emergence of Pertactin-Negative Strains.

Despite high vaccination coverage, Bordetella pertussis the causative agent of whooping cough is still a health concern worldwide. A resurgence of pertussis cases has been reported, particularly in countries using acellular vaccines with waning immunity and pathogen adaptation thought to be responsible. A better understanding of protective immune responses is needed for the development of improved vaccines. In our study, B. pertussis strain B1917 variants presenting a single gene deletion were generated to analyze the role of vaccine components or candidate vaccine antigens as targets for bactericidal antibodies generated after acellular vaccination or natural infection. Our results show that acellular vaccination generates bactericidal antibodies that are only directed against pertactin. Serum bactericidal assay performed with convalescent samples show that disease induces bactericidal antibodies against Prn but against other antigen(s) as well. Four candidate vaccine antigens (CyaA, Vag8, BrkA, and TcfA) have been studied but were not targets for complement-mediated bactericidal antibodies after natural infection. We confirm that Vag8 and BrkA are involved in complement resistance and would be targeted by blocking antibodies. Our study suggests that the emergence and the widespread circulation of Prn-deficient strains is driven by acellular vaccination and the generation of bactericidal antibodies targeting Prn.

Transcriptional regulation of plant senescence: from functional genomics to systems biology.

Leaf senescence is an active process that involves the increased expression of many hundreds of genes. Many putative transcription factors show enhanced transcription during leaf senescence in Arabidopsis and functional analysis of these should help to indicate their role in controlling gene expression during leaf senescence. In this paper, we describe the analysis of knockout insertion mutants in two different senescence-enhanced genes, one encodes a heat shock transcription factor and the other a zinc finger protein. Plants mutated in these genes show accelerated leaf senescence and reduced tolerance to drought stress, indicating that expression of these genes during senescence has a protective role to maintain viability during this essential developmental process. Analysis of gene expression changes in both mutants compared to the wild-type plants indicates an increased rate of senescence but does not show clearly the pathway that is dependent on these genes for expression. The complexities of signalling networks in plant stress and the plasticity of plant responses mean that the direct consequences of mutation are very difficult to define. The usefulness of this type of approach to address the burning question of how senescence is regulated is discussed, and an alternative approach aimed at a more global analysis of gene regulation using systems biology methods is described.

PEITC-mediated inhibition of mRNA translation is associated with both inhibition of mTORC1 and increased eIF2α phosphorylation in established cell lines and primary human leukemia cells.

Increased mRNA translation drives carcinogenesis and is an attractive target for the development of new anti-cancer drugs. In this work, we investigated effects of phenethylisothiocyanate (PEITC), a phytochemical with chemopreventive and anti-cancer activity, on mRNA translation. PEITC rapidly inhibited global mRNA translation in human breast cancer-derived MCF7 cells and mouse embryonic fibroblasts (MEFs). In addition to the known inhibitory effects of PEITC on mTORC1 activity, we demonstrate that PEITC increased eIF2α phosphorylation. PEITC also increased formation of stress granules which are typically associated with eIF2α phosphorylation and accumulation of translationally stalled mRNAs. Analysis of genetically modified MEFs demonstrated that optimal inhibition of global mRNA translation by PEITC was dependent on eIF2α phosphorylation, but not mTORC1 inhibition. We extended this study into primary leukemic B cells derived from patients with chronic lymphocytic leukaemia (CLL). CLL cells were stimulated in vitro with anti-IgM to mimic binding of antigen, a major driver of this leukemia. In CLL cells, PEITC increased eIF2α phosphorylation, inhibited anti-IgM-induced mTORC1 activation and decreased both basal and anti-IgM-induced global mRNA translation. PEITC also inhibited transcription and translation of MYC mRNA and accumulation of the MYC oncoprotein, in anti-IgM-stimulated cells. Moreover, treatment of CLL cells with PEITC and the BTK kinase inhibitor ibrutinib decreased anti-IgM-induced translation and induced cell death to a greater extent than either agent alone. Therefore, PEITC can inhibit both global and mRNA specific translation (including MYC) via effects on multiple regulatory pathways. Inhibition of mRNA translation may contribute to the chemopreventive and anti-cancer effects of PEITC.

Area deprivation, social class, and quality of life among people aged 75 years and over in Britain.

BACKGROUND: There is a shortage of research studies that assess how selected characteristics of neighbourhood and personal social circumstances contribute towards health-related quality of life (QoL) among older people. METHODS: Analysis of baseline data for 5581 people aged > or =75 years and over from the Trial of Assessment and Management of Older People in the Community. The scores for four dimensions from the UK version of the Sickness Impact Profile and for the Philadelphia Geriatric Morale Scale were analysed in relation to individual social class and the Carstairs score of socioeconomic deprivation for the enumeration district of residence. RESULTS: In age and sex adjusted analyses, the proportion of participants of social class IV/V living in the most deprived areas who were in the quintile with worst QoL scores was more than double that among those from social class I/II living in the least deprived areas. Individual social class and area deprivation score contributed roughly equally to this doubling for home management, self-care and social interaction, whereas social class appeared a stronger determinant for mobility. Adjustment for living circumstances, health symptoms, and health behaviours substantially reduced the excess risk associated with social class and area deprivation. Being in a rural area was associated with lower risk of poor morale. CONCLUSION: Poor socioeconomic characteristics of both the area and the individual are associated with worse functioning (QoL) of older people in the community. This is not fully explained by health status. Policy should consider community-level interventions as well as those directed at individuals.

Socioeconomic differences in the prevalence of common chronic diseases: an overview of eight European countries.

BACKGROUND: Few studies have compared socioeconomic inequalities in the prevalence of both fatal and non-fatal diseases. This paper aims to give the first international overview for several common chronic diseases. METHODS: Micro-level data were pooled from non-standardized national health surveys conducted in eight European countries in the 1990s. Surveys ranged in size from 3700 to 41 200 participants. The prevalence of 17 chronic disease groups were analysed in relation to education. Standardized prevalence rates and age-adjusted odds ratios (ORs) were calculated. RESULTS: Most diseases showed higher prevalence among the lower education group. Stroke, diseases of the nervous system, diabetes, and arthritis displayed relatively large inequalities (OR > 1.50). No socioeconomic differences were evident for cancer, kidney diseases, and skin diseases. Allergy was more common in the higher education group. Relative socioeconomic differences were often smaller among the 60-79 age group as compared with the 25-59 age group. Cancer was more prevalent among the lower educated in the 25-59 age group, but among the higher educated in the 60-79 age group. For diabetes, hypertension, and heart disease, socioeconomic differences were larger among women as compared with men. Inequalities in heart disease were larger in northern European countries as compared with southern European countries. CONCLUSION: There are large variations between chronic diseases in the size and pattern of socioeconomic differences in their prevalence. The large inequalities that are found for some specific fatal diseases (e.g. stroke) and non-fatal diseases (e.g. arthritis) require special attention in equity-oriented research and policies.

Socioeconomic and demographic predictors of mortality and institutional residence among middle aged and older people: results from the Longitudinal Study.

STUDY OBJECTIVES: To identify socioeconomic and demographic predictors of long term mortality and institutional residence in old age, taking into account changes in socioeconomic and demographic circumstances between the 1971 and 1981 censuses. DESIGN: Multivariate logistic regression modelling of outcomes for 10 year age cohorts of each gender. The outcomes were death by 31 December 1992; being in an institution in 1991. SETTING: Members of the Longitudinal Study (a 1% sample of the British Census): 43,092 men and 50,839 women aged 55-74 in 1971. MAIN RESULTS: Being in rented accommodation and in a household without access to a car carried 35-45% higher mortality rate over 21 years and similar excess risk of being in an institution in 1991. Marital status and living arrangements were weaker predictors of death but being single was a major predictor of moving to an institution for men. Losing household access to a car was a strong factor for mortality for men and for institutionalisation for men aged 55-64 in 1971. The effects were weaker for women. Moving into rented accommodation was a predictor of both outcomes for women and of death for the younger cohort of men. People who started to live alone in the inter-census period were at reduced risk of dying. CONCLUSIONS: These results demonstrate persistence of inequalities in health related outcomes throughout old age, both in those with unfavourable circumstances in mid-life and in those who, in later life, have lost earlier advantages.

Design, objectives, and lessons from a pilot 25 year follow up re-survey of survivors in the Whitehall study of London Civil Servants.

DESIGN: To assess the feasibility of conducting a re-survey of men who are resident in the United Kingdom 25 years after enrollment in the Whitehall study of London Civil Servants. METHODS: A random sample of 401 study survivors resident in three health authority areas was selected for this pilot study. They were mailed a request to complete a self administered questionnaire, and then asked to attend their general practice to have their blood pressure, weight, and height measured and a blood sample collected into a supplied vacutainer, and mailed to a central laboratory. Using a 2 x 2 factorial design, the impact of including additional questions on income and of an informant questionnaire on cognitive function was assessed. RESULTS: Accurate addresses were obtained from the health authorities for 96% of the sample. Questionnaires were received from 73% and blood samples from 61% of the sample. Questions on income had no adverse effect on the response rate, but inclusion of the informant questionnaire did. Between 1970 and 1995 there were substantial changes within men in the mean blood pressure and blood total cholesterol recorded, as reflected by correlation coefficients between 1970 and 1995 values of 0.26, and 0.30 for systolic and diastolic blood pressure and 0.38 for total cholesterol. CONCLUSION: This pilot study demonstrated the feasibility of conducting a re-survey using postal questionnaires and mailed whole blood samples. The magnitude of change in blood pressure and blood total cholesterol concentrations within individuals was greater than anticipated, suggesting that such remeasurements may be required at different intervals in prospective studies to help interpret risks associations properly. These issues will be considered in a re-survey of the remaining survivors of the Whitehall study.

Quality of life and cough on antihypertensive treatment: a randomised trial of eprosartan, enalapril and placebo.

The objective of this study was to compare the quality of life and incidence of dry cough with the angiotensin II antagonist eprosartan, the ACE-inhibitor enalapril, and placebo, in hypertensive patients with a history of ACE-inhibitor cough. The study was a multicentre, randomised, double-blind, parallel group controlled trial. A total of 136 patients judged to have ACE-inhibitor cough during single-blind enalapril treatment which was lost during a subsequent placebo washout phase, were randomised to receive either eprosartan 300 mg twice daily, or enalapril 20 mg once daily, or placebo for 6 weeks. Self-completion questionnaires assessing quality of life and cough were examined at baseline and end of study. At study end point 23% of patients in the enalapril group and 5% in the eprosartan and placebo groups reported cough (which included definite, probable and possible coughs) (P = 0.02). After adjusting for multiple comparisons, the eprosartan group was not significantly different from either placebo or enalapril. There were no significant differences in the Psychological General Wellbeing Index (PGWB). In conclusion the incidence of self-reported cough on eprosartan was similar to that on placebo, and lower than on enalapril but this difference was not significant when adjustments were made for multiple comparisons. There were no differences in quality of life.

Comparison of quality of life and cough on eprosartan and enalapril in people with moderate hypertension.

The objective of this study was to compare quality of life and incidence of dry persistent cough among patients treated with eprosartan and enalapril for mild-moderate hypertension. This was a randomised 26-week double-blind controlled trial carried out in clinics in nine countries of North America, Europe and South Africa. A total of 529 patients aged 18 and over with diastolic blood pressure between 95 mm Hg and 114 mm Hg were studied. Treatment comprised of eprosartan or enalapril monotherapy for 12 weeks with the option of hydrochlorothiazide addition for the remaining 14 weeks. The primary outcome measures were cough and the Psychological General Wellbeing Index (PGWB) total and subscales (anxiety, self-control, depression, general health, positive wellbeing and vitality). The results were that 17.8% of enalapril patients and 13.2% of eprosartan patients withdrew from randomised treatment. Those on enalapril were twice as likely to have gained a definite or possible cough by study end point as those on eprosartan (7.6% vs 3.2%) P = 0.099. At monotherapy end point the differences were greater (9.9% vs 2.1%) and of statistical significance, P = 0.001. Patients treated with enalapril, however, had small but significant improvements in measures of self-control and total PGWB compared with those on eprosartan. The effect sizes of 0.2 or less indicated that there were small differences. In conclusion eprosartan was associated with fewer coughs than enalapril but it performed less well on some aspects of quality of life.

Prevalence of visual impairment in people aged 75 years and older in Britain: results from the MRC trial of assessment and management of older people in the community.

AIMS: To measure the prevalence of visual impairment in a large representative sample of people aged 75 years and over participating in the MRC trial of assessment and management of older people in the community. METHODS: 53 practices in the MRC general practice research framework. Data were obtained from 14 600 participants aged 75 years and older. Prevalence of visual impairment overall (binocular visual acuity <6/18) which was categorised separately into low vision (binocular visual acuity <6/18-3/60) or blindness (binocular visual acuity of <3/60). The prevalence of binocular acuity <6/12 was presented for comparison with other studies. Visual acuity was measured using Glasgow acuity charts; glasses, if worn, were not removed. RESULTS: Visual acuity was available for 14 600 people out of 21 241 invited (69%). Among people with visual acuity data, 12.4% overall (1803) were visually impaired (95% confidence intervals 10.8% to 13.9%); 1501 (10.3%) were categorised as having low vision (8.7% to 11.8%), and 302 (2.1%) were blind (1.8% to 2.4%). At ages 75-79, 6.2% of the cohort were visually impaired (5.1% to 7.3%) with 36.9% at age 90+ (32.5% to 41.3%). At ages 75-79, 0.6% (0.4% to 0.8%) of the study population were blind, with 6.9% (4.8% to 9.0%) at age 90+. In multivariate regression, controlling for age, women had significant excess risk of visual impairment (odds ratio 1.43, 95% confidence interval 1.29 to 1.58). Overall, 19.9% of study participants had a binocular acuity of less than 6/12 (17.8% to 22.0%). CONCLUSION: The results from this large study show that visual impairment is common in the older population and that this risk increases rapidly with advancing age, especially for women. A relatively conservative measure of visual impairment was used. If visual impairment had been defined as visual acuity of <6/12 (American definition of visual impairment), the age specific prevalence estimates would have increased by 60%.

Randomised comparison of three methods of administering a screening questionnaire to elderly people: findings from the MRC trial of the assessment and management of older people in the community.

OBJECTIVE: To compare three different methods of administering a brief screening questionnaire to elderly people: post, interview by lay interviewer, and interview by nurse. DESIGN: Randomised comparison of methods within a cluster randomised trial. SETTING: 106 general practices in the United Kingdom. PARTICIPANTS: 32 990 people aged 75 years or over registered with participating practices. MAIN OUTCOME MEASURES: Response rates, proportion of missing values, prevalence of self reported morbidity, and sensitivity and specificity of self reported measures by method of administration of questionnaire for four domains. RESULTS: The response rate was higher for the postal questionnaire than for the two interview methods combined (83.5% v 74.9%; difference 8.5%, 95% confidence interval 4.4% to 12.7%, P<0.001). The proportion of missing or invalid responses was low overall (mean 2.1%) but was greater for the postal method than for the interview methods combined (4.1% v 0.9%; difference 3.2%, 2.7% to 3.6%, P<0.001). With a few exceptions, levels of self reported morbidity were lower in the interview groups, particularly for interviews by nurses. The sensitivity of the self reported measures was lower in the nurse interview group for three out of four domains, but 95% confidence intervals for the estimates overlapped. Specificity of the self reported measures varied little by method of administration. CONCLUSIONS: Postal questionnaires were associated with higher response rates but also higher proportions of missing values than were interview methods. Lower estimates of self reported morbidity were obtained with the nurse interview method and to a lesser extent with the lay interview method than with postal questionnaires.

Differential induction of apoptosis in human breast cancer cell lines by phenethyl isothiocyanate, a glutathione depleting agent.

Phenethyl isothiocyanate (PEITC) is a naturally occurring electrophile which depletes intracellular glutathione (GSH) levels and triggers accumulation of reactive oxygen species (ROS). PEITC is of considerable interest as a potential chemopreventive/chemotherapeutic agent, and in this work, we have investigated the effects of PEITC on human breast cancer cell lines. Whereas PEITC readily induced apoptosis in MDA-MB-231 cells (associated with rapid activation of caspases 9 and 3, and decreased expression of BAX), MCF7 cells were relatively resistant to the apoptosis promoting effects of PEITC. The relative resistance of MCF7 cells was associated with high basal expression of NRF2, a transcription factor that coordinates cellular protective responses to oxidants and electrophiles and raised intracellular levels of GSH. This raised basal expression of NRF2 appeared to be a response to on-going production of ROS, since treatment with the antioxidant and GSH precursor N-acetylcysteine (NAC) reduced NRF2 expression. Moreover, pre-treatment of MDA-MB-231 cells with NAC rendered these cells relatively resistant to PEITC-induced apoptosis. In summary, our data confirm that PEITC may be an effective chemopreventive/therapeutic agents for breast cancer. However, differences in the basal expression of NRF2 and resultant changes in GSH levels may be an important determinant of sensitivity to PEITC-induced apoptosis.