Genomic modelling of the ESR1 Y537S mutation for evaluating function and new therapeutic approaches for metastatic breast cancer

Drugs that inhibit estrogen receptor-α (ER) activity have been highly successful in treating and reducing breast cancer progression in ER-positive disease. However, resistance to these therapies presents a major clinical problem. Recent genetic studies have shown that mutations in the ER gene are found in >20% of tumours that progress on endocrine therapies. Remarkably, the great majority of these mutations localize to just a few amino acids within or near the critical helix 12 region of the ER hormone binding domain, where they are likely to be single allele mutations. Understanding how these mutations impact on ER function is a prerequisite for identifying methods to treat breast cancer patients featuring such mutations. Towards this end, we used CRISPR-Cas9 genome editing to make a single allele knock-in of the most commonly mutated amino acid residue, tyrosine 537, in the estrogen-responsive MCF7 breast cancer cell line. Genomic analyses using RNA-seq and ER ChIP-seq demonstrated that the Y537S mutation promotes constitutive ER activity globally, resulting in estrogen-independent growth. MCF7-Y537S cells were resistant to the anti-estrogen tamoxifen and fulvestrant. Further, we show that the basal transcription factor TFIIH is constitutively recruited by ER-Y537S, resulting in ligand-independent phosphorylation of Serine 118 (Ser118) by the TFIIH kinase, cyclin-dependent kinase (CDK)7. The CDK7 inhibitor, THZ1 prevented Ser118 phosphorylation and inhibited growth of MCF7-Y537S cells. These studies confirm the functional importance of ER mutations in endocrine resistance, demonstrate the utility of knock-in mutational models for investigating alternative therapeutic approaches and highlight CDK7 inhibition as a potential therapy for endocrine-resistant breast cancer mediated by ER mutations

Maternal hyperleptinemia is associated with male offspring’s altered vascular function and structure in mice

Children of mothers with gestational diabetes have greater risk of developing hypertension but little is known about the mechanisms by which this occurs. The objective of this study was to test the hypothesis that high maternal concentrations of leptin during pregnancy, which are present in mothers with gestational diabetes and/or obesity, alter blood pressure, vascular structure and vascular function in offspring. Wildtype (WT) offspring of hyperleptinemic, normoglycemic, Lepr db/+ dams were compared to genotype matched offspring of WT-control dams. Vascular function was assessed in male offspring at 6, and at 31 weeks of age after half the offspring had been fed a high fat, high sucrose diet (HFD) for 6 weeks. Blood pressure was increased by HFD but not affected by maternal hyperleptinemia. On a standard diet, offspring of hyperleptinemic dams had outwardly remodeled mesenteric arteries and an enhanced vasodilatory response to insulin. In offspring of WT but not Leprdb/+ dams, HFD induced vessel hypertrophy and enhanced vasodilatory responses to acetylcholine, while HFD reduced insulin responsiveness in offspring of hyperleptinemic dams. Offspring of hyperleptinemic dams had stiffer arteries regardless of diet. Therefore, while maternal hyperleptinemia was largely beneficial to offspring vascular health under astandard diet, it had detrimental effects in offspring fed HFD. These results suggest that circulating maternal leptin concentrations may interact with other factors in the pre- and post-natal environments to contribute to altered vascular function in offspring of diabetic pregnancie

NF-κB Hyper-Activation by HTLV-1 Tax Induces Cellular Senescence, but Can Be Alleviated by the Viral Anti-Sense Protein HBZ

Activation of I-κB kinases (IKKs) and NF-κB by the human T lymphotropic virus type 1 (HTLV-1) trans-activator/oncoprotein, Tax, is thought to promote cell proliferation and transformation. Paradoxically, expression of Tax in most cells leads to drastic up-regulation of cyclin-dependent kinase inhibitors, p21CIP1/WAF1 and p27KIP1, which cause p53-/pRb-independent cellular senescence. Here we demonstrate that p21CIP1/WAF1-/p27KIP1-mediated senescence constitutes a checkpoint against IKK/NF-κB hyper-activation. Senescence induced by Tax in HeLa cells is attenuated by mutations in Tax that reduce IKK/NF-κB activation and prevented by blocking NF-κB using a degradation-resistant mutant of I-κBα despite constitutive IKK activation. Small hairpin RNA-mediated knockdown indicates that RelA induces this senescence program by acting upstream of the anaphase promoting complex and RelB to stabilize p27KIP1 protein and p21CIP1/WAF1 mRNA respectively. Finally, we show that down-regulation of NF-κB by the HTLV-1 anti-sense protein, HBZ, delay or prevent the onset of Tax-induced senescence. We propose that the balance between Tax and HBZ expression determines the outcome of HTLV-1 infection. Robust HTLV-1 replication and elevated Tax expression drive IKK/NF-κB hyper-activation and trigger senescence. HBZ, however, modulates Tax-mediated viral replication and NF-κB activation, thus allowing HTLV-1-infected cells to proliferate, persist, and evolve. Finally, inactivation of the senescence checkpoint can facilitate persistent NF-κB activation and leukemogenesis

Exhaled Aerosol Transmission of Pandemic and Seasonal H1N1 Influenza Viruses in the Ferret

Person-to-person transmission of influenza viruses occurs by contact (direct and fomites) and non-contact (droplet and small particle aerosol) routes, but the quantitative dynamics and relative contributions of these routes are incompletely understood. The transmissibility of influenza strains estimated from secondary attack rates in closed human populations is confounded by large variations in population susceptibilities. An experimental method to phenotype strains for transmissibility in an animal model could provide relative efficiencies of transmission. We developed an experimental method to detect exhaled viral aerosol transmission between unanesthetized infected and susceptible ferrets, measured aerosol particle size and number, and quantified the viral genomic RNA in the exhaled aerosol. During brief 3-hour exposures to exhaled viral aerosols in airflow-controlled chambers, three strains of pandemic 2009 H1N1 strains were frequently transmitted to susceptible ferrets. In contrast one seasonal H1N1 strain was not transmitted in spite of higher levels of viral RNA in the exhaled aerosol. Among three pandemic strains, the two strains causing weight loss and illness in the intranasally infected ‘donor’ ferrets were transmitted less efficiently from the donor than the strain causing no detectable illness, suggesting that the mucosal inflammatory response may attenuate viable exhaled virus. Although exhaled viral RNA remained constant, transmission efficiency diminished from day 1 to day 5 after donor infection. Thus, aerosol transmission between ferrets may be dependent on at least four characteristics of virus-host relationships including the level of exhaled virus, infectious particle size, mucosal inflammation, and viral replication efficiency in susceptible mucosa

Somatic growth in the first six months of life of infants exposed to maternal smoking in pregnancy

Background: Some studies suggest a relationship between maternal smoking during pregnancy and not only intrauterine fetal growth restriction or low birth weight, but also with changes in the postnatal growth and development. The objective of the present study was to investigate the effects of smoking during pregnancy on infants growth in the first 6 months of life compared with a control group and a group with idiopathic intrauterine growth restriction. Methods: Longitudinal observational study using a convenience sample of newborns divided into three groups: infants of smoking mothers (tobacco), with idiopathic intrauterine growth restriction (IUGR) and a control group. The sample was selected from two hospitals in Porto Alegre, located in southern Brazil, between 2011 and 2015. Newborns were evaluated at birth, 7 and 15 days, and in the first, third, and sixth month. Anthropometric measures were weight, length and head circumference. The growth indicators used were expressed as z-scores. The analyses were performed using the generalized estimating equation method. Results: The sample included 273 mother/newborn pairs: 86 tobacco group, 34 IUGR group, and 153 control group. In terms of weight at birth, all groups differed significantly (p < 0.001). The birth length of tobacco and control groups were similar, but the IUGR group was lower than both (p < 0.001). We found no differences in growth trajectory between tobacco and control group, but there were differences in the growth of the IUGR group when compared with the other groups. At 6 months of age, all groups had similar anthropometric measurements. Conclusion: Intrauterine growth restriction had major impact on the growth trajectory of the infants studied, regardless of other factors, such as smoking and diet

Changes in leisure-time physical activity among Brazilian pregnant women: comparison between two birth cohort studies (2004 – 2015)

Abstract Background Low levels of leisure-time physical activity (LTPA) during pregnancy have been shown in studies conducted worldwide. Surveillance is extremely important to monitor the progress of physical activity patterns over time and set goals for effective interventions to decrease inactivity among pregnant women. The aim of this study was to evaluate time changes in LTPA among Brazilian pregnant women in an 11-year period (2004–2015) by comparing data from two birth cohort studies. Methods Two population-based birth cohort studies were carried out in the city of Pelotas, southern Brazil, in 2004 and 2015. A total of 4244 and 4271 mothers were interviewed after delivery. Weekly frequency and duration of each session of LTPA in a typical week were reported for the pre-pregnancy period and for each trimester of pregnancy. Trends in both recommended LTPA (≥150 min/week) and any LTPA (regardless of weekly amount) were analysed overtime. Changes were also calculated separately for subgroups of maternal age, schooling, family income, parity, pre-pregnancy body mass index and pre-pregnancy LTPA. Results The proportion of women engaged in recommended levels of LTPA pre-pregnancy increased from 11.2% (95%CI 10.0–12.2) in 2004 to 15.8% (95%CI 14.6–16.9) in 2015. During pregnancy, no changes were observed over the period for the first (10.6 to 10.9%) and second (8.7 to 7.9%) trimesters, whereas there was a decrease from 3.4% (95%CI 2.9–4.0) to 2.4% (95%CI 1.9–2.8) in the last trimester. Major decreases in LTPA in the last trimester were observed among women who were younger, with intermediate to high income, high schooling, primiparous, pre-pregnancy obese and, engaged in LTPA before pregnancy. Changes in any LTPA practice followed the same patterns described for recommended LTPA. Conclusions Despite the increase in the proportion of women engaged in LTPA before pregnancy between 2004–2015, LTPA levels remained stable during the first and second trimesters of pregnancy and declined during the third gestational trimester over the period. Interventions to encourage the maintenance of LTPA practice throughout pregnancy are urgently needed

Fetal Exposure to Maternal Active and Secondhand Smoking with Offspring Early-life Growth in the Healthy Start Study

Background Previous studies have modeled the association between fetal exposure to tobacco smoke and body mass index (BMI) growth trajectories, but not the timing of catch-up growth. Research on fetal exposure to maternal secondhand smoking is limited. Objectives To explore the associations between fetal exposure to maternal active and secondhand smoking with body composition at birth and BMI growth trajectories through age 3 years. Methods We followed 630 mother-child pairs enrolled in the Healthy Start cohort through age 3 years. Maternal urinary cotinine was measured at ~ 27 weeks gestation. Neonatal body composition was measured using air displacement plethysmography. Child weight and length/height were abstracted from medical records. Linear regression models examined the association between cotinine categories (no exposure, secondhand smoke, active smoking) with weight, fat mass, fat-free mass, and percent fat mass at birth. A mixed-effects regression model estimated the association between cotinine categories and BMI. Results Compared to unexposed offspring, birth weight was significantly lower among offspring born to active smokers (−343-g; 95% CI: −473, −213), but not among offspring of women exposed to secondhand smoke (−47-g; 95% CI: −130, 36). There was no significant difference in the rate of BMI growth over time between offspring of active and secondhand smokers (p = 0.58). Therefore, our final model included a single growth rate parameter for the combined exposure groups of active and secondhand smokers. The rate of BMI growth for the combined exposed group was significantly more rapid (0.27 kg/m2 per year; 95% CI: 0.05, 0.69; p \u3c 0.01) than the unexposed. Conclusions Offspring prenatally exposed to maternal active or secondhand smoking experience rapid and similar BMI growth in the first three years of life. Given the long-term consequences of rapid weight gain in early childhood, it is important to encourage pregnant women to quit smoking and limit their exposure to secondhand smoke

A decision support tool to inform local suicide prevention activity in Greater Western Sydney (Australia)

Objectives: This study describes the development of a decision support tool to identify the combination of suicide prevention activities and service priorities likely to deliver the greatest reductions in suicidal behaviour in Western Sydney (Australia) over the period 2018–2028. Methods: A dynamic simulation model for the WentWest – Western Sydney Primary Health Network population-catchment was developed in partnership with primary health network stakeholders based on defined pathways to mental health care and suicidal behaviour, and which represented the current incidence of suicide and attempted suicide in Western Sydney. A series of scenarios relating to potential suicide prevention activities and service priorities identified by primary health network stakeholders were investigated to identify the combination of interventions associated with the largest reductions in the forecast number of attempted suicide and suicide cases for a 10-year follow-up period. Results: The largest number of cases averted for both suicide and attempted suicide was associated with (1) post-suicide attempt assertive aftercare (6.1% for both attempted suicide and suicide), (2) improved community support and reductions in psychological distress in the community (5.1% for attempted suicide and 14.8% for suicide), and (3) reductions in the proportion of those lost to services following a mental health service contact (10.5% for both attempted suicide and suicide). In combination, these interventions were forecast to avert approximately 29.7% of attempted suicides and 37.1% of suicides in the primary health network catchment over the 10-year period. Conclusion: This study demonstrates the utility of dynamic simulation models, co-designed with multi-disciplinary stakeholder groups, to capture and analyse complex mental health and suicide prevention regional planning problems. The model can be used by WentWest – Western Sydney Primary Health Network as a decision support tool to guide the commissioning of future service activity, and more efficiently frame the monitoring and evaluation of interventions as they are implemented in Western Sydney