PIK3CA mutations and EGFR overexpression predict for lithium sensitivity in human breast epithelial cells

A high frequency of somatic mutations has been found in breast cancers within the gene encoding the catalytic p110α subunit of PI3K, PIK3CA. Using isogenic human breast epithelial cells, we have previously demonstrated that oncogenic PIK3CA "hotspot" mutations predict for response to the toxic effects of lithium. However, other somatic genetic alterations occur within this pathway in breast cancers, and it is possible that these changes may also predict for lithium sensitivity. We overexpressed the epidermal growth factor receptor (EGFR) into the non-tumorigenic human breast epithelial cell line MCF-10A, and compared these cells to isogenic cell lines previously created via somatic cell gene targeting to model Pten loss, PIK3CA mutations, and the invariant AKT1 mutation, E17K. EGFR overexpressing clones were capable of cellular proliferation in the absence of EGF and were sensitive to lithium similar to the results previously seen with cells harboring PIK3CA mutations. In contrast, AKT1 E17K cells and PTEN -/- cells displayed resistance or partial sensitivity to lithium, respectively. Western blot analysis demonstrated that lithium sensitivity correlated with significant decreases in both PI3K and MAPK signaling that were observed only in EGFR overexpressing and mutant PIK3CA cell lines. These studies demonstrate that EGFR overexpression and PIK3CA mutations are predictors of response to lithium, whereas Pten loss and AKT1 E17K mutations do not predict for lithium sensitivity. Our findings may have important implications for the use of these genetic lesions in breast cancer patients as predictive markers of response to emerging PI3K pathway inhibitors.     

Satellite-based global-ocean mass balance estimates of interannual variability and emerging trends in continental freshwater discharge

Freshwater discharge from the continents is a key component of Earth’s water cycle that sustains human life and ecosystem health. Surprisingly, owing to a number of socioeconomic and political obstacles, a comprehensive global river discharge observing system does not yet exist. Here we use 13 years (1994–2006) of satellite precipitation, evaporation, and sea level data in an ocean mass balance to estimate freshwater discharge into the global ocean. Results indicate that global freshwater discharge averaged 36,055 km³/y for the study period while exhibiting significant interannual variability driven primarily by El Niño Southern Oscillation cycles. The method described here can ultimately be used to estimate long-term global discharge trends as the records of sea level rise and ocean temperature lengthen. For the relatively short 13-year period studied here, global discharge increased by 540 km³/y², which was largely attributed to an increase of global-ocean evaporation (768 km³/y²). Sustained growth of these flux rates into long-term trends would provide evidence for increasing intensity of the hydrologic cycle.     

Differential dynamics and activity-dependent regulation of alpha- and beta-neurexins at developing GABAergic synapses

Neurexins (NRXs) and neuroligins are key synaptic adhesion molecules that also recruit synaptic signaling machineries. Neurexins consist of α- and β-isoforms, but how they couple synaptic transmission and adhesion to regulate activity-dependent synapse development remains unclear, in part because of poor understanding of their cell biology and regulation in the relevant neurons. Here, we examined the subaxonal localization, dynamics, and regulation of NRX1α and NRX1β in cortical perisomatic inhibitory synapses. Both isoforms are delivered to presynaptic terminals but show significant and different turnover rate at the membrane. Although NRX1α is highly diffuse along developing axons and filopodia, NRX1β is strictly anchored at terminals through binding to postsynaptic ligands. The turnover rate of NRX1β is attenuated by neural activity and presynaptic GABA(B) receptors. NRXs, thus, are intrinsically dynamic but are stabilized by local transmitter release. Such an activity-adjusted adhesion system seems ideally suited to rapidly explore and validate synaptic partners guided by synaptic transmission.     

Templated Open Flocs of Nanorods for Enhanced Pulmonary Delivery with Pressurized Metered Dose Inhalers

Purpose  A novel concept is presented for the formation of stable suspensions composed of low density flocs of high aspect ratio drug particles in hydrofluoroalkane (HFA) propellants, and for subdividing (templating) the flocs with aerosolized HFA droplets to achieve high fine particle fractions with a pressurized metered dose inhaler. Methods  Bovine serum albumin (BSA) nanorods, produced by thin film freezing (TFF), were added to HFA to form a suspension. Particle properties were analyzed with an Anderson cascade impactor (ACI), static and dynamic light scattering and optical microscopy. Results  The space filling flocs in HFA were stable against settling for one year. The pMDI produced high fine particle fractions (38–47%) with an emitted dose of 0.7 mg/actuation. The atomized HFA droplets break apart, that is template, the highly open flocs. Upon evaporation of HFA, capillary forces shrink the templated flocs to produce porous particles with optimal aerodynamic diameters for deep lung delivery. Conclusions  Open flocs composed of nanorods, stable against settling, may be templated during actuation with a pMDI to produce optimal aerodynamic diameters and high fine particle fractions. This concept is applicable to a wide variety of drugs without the need for surfactants or cosolvents to stabilize the primary particles.     

The growing impact of HIV infection on the epidemiology of tuberculosis in England and Wales: 1999 2003

BACKGROUND: Previous studies have estimated the prevalence of tuberculosis and HIV infection in population subgroups in the UK. This study was undertaken to describe recent trends in the proportion of individuals with HIV infection among reported cases of tuberculosis in England and Wales, and to review the implications for clinical and public health care. METHODS: A population-based matching study using national surveillance databases was used to investigate all persons aged 15 years and over reported with a diagnosis of tuberculosis to the Health Protection Agency in England and Wales in 1999-2003. Record linkage was used to match the national tuberculosis and HIV/AIDS surveillance databases to identify all cases of tuberculosis and determine the proportion of patients with tuberculosis co-infected with HIV. The distribution and characteristics of the cases were determined and the trend examined by year. RESULTS: Of 30,670 cases of tuberculosis reported in England and Wales between 1999 and 2003, an estimated 1743 (5.7%) were co-infected with HIV. There was a year on year increase in the proportion from 3.1% (169/5388) in 1999 to 8.3% (548/6584) in 2003 (p for trend <0.0001). Co-infected patients contributed to almost a third of the increase in the number of cases of tuberculosis during the 5 year period. Patients co-infected with HIV were predominantly those born abroad. 18.5% (n = 323) of co-infected patients had not been reported as active cases of tuberculosis on the national tuberculosis database. CONCLUSION: The proportion of patients with tuberculosis co-infected with HIV in England and Wales is increasing, with the greatest impact on those born abroad regardless of their ethnic origin. With HIV infection contributing substantially to the increase in the number of cases of tuberculosis, close cooperation in the clinical management and accurate notification of patients is vital if appropriate care and public health action is to be achieved.     

Knock-in of mutant K-ras in nontumorigenic human epithelial cells as a new model for studying K-ras mediated transformation

The oncogenic function of mutant ras in mammalian cells has been extensively investigated using multiple human and animal models. These systems include overexpression of exogenous mutant ras transgenes, conditionally expressed knock-in mouse models, and somatic cell knockout of mutant and wild-type ras genes in human cancer cell lines. However, phenotypic discrepancies between knock-in mice and transgenic mutant ras overexpression prompted us to evaluate the consequences of targeted knock-in of an oncogenic K-ras mutation in the nontumorigenic human breast epithelial cell line MCF-10A and hTERT-immortalized human mammary epithelial cells. Our results show several significant differences between mutant K-ras knock-in cells versus their transgene counterparts, including limited phosphorylation of the downstream molecules extracellular signal-regulated kinase and AKT, minor proliferative capacity in the absence of an exogenous growth factor, and the inability to form colonies in semisolid medium. Analysis of 16 cancer cell lines carrying mutant K-ras genes indicated that 50% of cancer cells harbor nonoverexpressed heterozygous K-ras mutations similar to the expression seen in our knock-in cell lines. Thus, this system serves as a new model for elucidating the oncogenic contribution of mutant K-ras as expressed in a large fraction of human cancer cells.