Targeting AKT1-E17K and the PI3K/AKT Pathway with an Allosteric AKT Inhibitor,ARQ 092

As a critical component in the PI3K/AKT/mTOR pathway, AKT has become an attractive target for therapeutic intervention. ARQ 092 and a next generation AKT inhibitor, ARQ 751 are selective, allosteric, pan-AKT and AKT1-E17K mutant inhibitors that potently inhibit phosphorylation of AKT. Biochemical and cellular analysis showed that ARQ 092 and ARQ 751 inhibited AKT activation not only by dephosphorylating the membrane-associated active form, but also by preventing the inactive form from localizing into plasma membrane. In endometrial PDX models harboring mutant AKT1-E17K and other tumor models with an activated AKT pathway, both compounds exhibited strong anti-tumor activity. Combination studies conducted in in vivo breast tumor models demonstrated that ARQ 092 enhanced tumor inhibition of a common chemotherapeutic agent (paclitaxel). In a large panel of diverse cancer cell lines, ARQ 092 and ARQ 751 inhibited proliferation across multiple tumor types but were most potent in leukemia, breast, endometrial, and colorectal cancer cell lines. Moreover, inhibition by ARQ 092 and ARQ 751 was more prevalent in cancer cell lines containing PIK3CA/PIK3R1 mutations compared to those with wt-PIK3CA/PIK3R1 or PTEN mutations. For both ARQ 092 and ARQ 751, PIK3CA/PIK3R1 and AKT1-E17K mutations can potentially be used as predictive biomarkers for patient selection in clinical studies.     

Selection, trans-species polymorphism, and locus identification of major histocompatibility complex class IIβ alleles of New World ranid frogs

Genes encoded by the major histocompatibility complex (MHC) play key roles in the vertebrate immune system. However, our understanding of the evolutionary processes and underlying genetic mechanisms shaping these genes is limited in many taxa, including amphibians, a group currently impacted by emerging infectious diseases. To further elucidate the evolution of the MHC in frogs (anurans) and develop tools for population genetics, we surveyed allelic diversity of the MHC class II β1 domain in both genomic and complementary DNA of seven New World species in the genus Rana (Lithobates). To assign locus affiliation to our alleles, we used a “gene walking” technique to obtain intron 2 sequences that flanked MHC class IIβ exon 2. Two distinct intron sequences were recovered, suggesting the presence of at least two class IIβ loci in Rana. We designed a primer pair that successfully amplified an orthologous locus from all seven Rana species. In total, we recovered 13 alleles and documented trans-species polymorphism for four of the alleles. We also found quantitative evidence of selection acting on amino acid residues that are putatively involved in peptide binding and structural stability of the β1 domain of anurans. Our results indicated that primer mismatch can result in polymerase chain reaction (PCR) bias, which influences the number of alleles that are recovered. Using a single locus may minimize PCR bias caused by primer mismatch, and the gene walking technique was an effective approach for generating single-copy orthologous markers necessary for future studies of MHC allelic variation in natural amphibian populations.     

iNKT cells require CCR4 to localize to the airways and to induce airway hyperreactivity

iNKT cells are required for the induction of airway hyperreactivity (AHR), a cardinal feature of asthma, but how iNKT cells traffic to the lungs to induce AHR has not been previously studied. Using several models of asthma, we demonstrated that iNKT cells required the chemokine receptor CCR4 for pulmonary localization and for the induction of AHR. In both allergen-induced and glycolipid-induced models of AHR, wild-type but not CCR4-/- mice developed AHR. Furthermore, adoptive transfer of wild-type but not CCR4-/- iNKT cells reconstituted AHR in iNKT cell-deficient mice. Moreover, we specifically tracked CCR4-/- vs wild-type iNKT cells in CCR4-/-:wild-type mixed BM chimeric mice in the resting state, and when AHR was induced by protein allergen or glycolipid. Using this unique model, we showed that both iNKT cells and conventional T cells required CCR4 for competitive localization into the bronchoalveolar lavage/airways compartment. These results establish for the first time that the pulmonary localization of iNKT cells critical for the induction of AHR requires CCR4 expression by iNKT cells.     

Investigation of protein expression in magnetic field-treated human glioma cells

We previously reported phenotypic changes in human breast cancer cells following low-level magnetic field (MF) exposure. Here proteomic methods were used to investigate the biochemical effect of MF exposure in SF767 human glioma cells. Protein alterations were studied after exposure to 1.2 microTesla (microT) MF [12 milliGauss (mG), 60 Hertz (Hz)] +/- epidermal growth factor (EGF). SF767 cells were exposed for 3 h to sham conditions (<0.2 microT ambient field strength) or 1.2 microT MF (+/-EGF; 10 ng/ml). Solubilized protein fractions (sham; 1.2 microT; sham + EGF; 1.2 microT + EGF) were loaded for electrophoresis by 2D-PAGE and stained using a colloidal Coomassie blue technique to resolve and characterize the proteins. Protein patterns were compared across groups via Student's t-test using PDQUEST software. Cell profiles revealed significant alterations in the spot density of a subset of treated cells. Automated spot excision and processing was performed prior to peptide mass fingerprinting proteins of interest. Fifty-seven proteins from the detectable pool were identified and/or found to differ significantly across treatment groups. The mean abundance of 10 identified proteins was altered following 1.2 microT exposure. In the presence of EGF six proteins were altered after low magnetic field treatment by increasing (4) or decreasing (2) in abundance. The results suggest that the analysis of differentially expressed proteins in SF767 cells may be useful as biomarkers for biological changes caused by exposure to magnetic fields.     

Final height in patients with idiopathic short stature and high growth hormone responses to stimulation tests

Children with idiopathic short stature (ISS) may have normal or increased growth hormone (GH) responses to provocation tests and achieve a final height (FH) below -2.0 standard deviation score (SDS) if untreated. FH of subjects with high stimulated GH levels has not been studied in detail. AIM: It was the aim of this study to analyse FH in ISS patients with high GH peak responses to the provocation test. PATIENTS AND METHODS: We studied 16 patients (9 pre-pubertal) with ISS and a GH peak >or=40 mU/l to insulin-induced hypoglycaemia. The patients were recalled at age 19.7 +/- 2.5 years for measurement of FH when blood samples were obtained for serum insulin-like growth factor (IGF)-I, IGF binding protein 3, acid-labile subunit and GH binding protein measurements. GH bioactivity was determined using the Nb2 bioassay. RESULTS: FH was -3.1 +/- 1.0 SDS, being significantly lower than target height (TH). At FH, IGF-I levels were within -1.5 and +1.5 SDS for age and sex in 10 patients and higher than +1.5 SDS in 6 patients. IGF binding protein 3, acid-labile subunit, GH binding protein levels and GH bioactivity values were normal. SUMMARY: These data suggest that patients with ISS and high GH levels during a GH stimulation test may have a more compromised FH. The association of severe ISS with a peak GH >40 mU/l might suggest a degree of insensitivity for the GH-IGF-I axis.     

Lethal combat and sex ratio evolution in a parasitoid wasp

Sex allocation theory provides excellent opportunities for testinghow behavior and life histories are adjusted in response toenvironmental variation. One of the most successful areas fromthis respect is Hamilton's local mate competition theory. Aspredicted by theory, a large number of animal species have beenshown to adjust their offspring sex ratios (proportion male)conditionally, laying less female-biased sex ratios as the numberof females that lay eggs on a patch increases. However, recentstudies have shown that this predicted pattern is not followedby 2 parasitoid species in the genus Melittobia, which alwaysproduce extremely female-biased sex ratios. A possible explanationfor this is that males fight fatally and that males producedby the first female to lay eggs on a patch have a competitiveadvantage over later emerging males. This scenario would negatethe advantage of later females producing a less female-biasedsex ratio. Here we examine fatal fighting and sex ratio evolutionin another species, Melittobia acasta. We show that femalesof this species also fail to adjust their offspring sex ratioin response to the number of females laying eggs on a patch.We then show that although earlier emerging males do have anadvantage in winning fights, this advantage 1) can be reducedby an interaction with body size, with larger males more likelyto win fights and 2) only holds for a brief period around thetime at which the younger males emerge from their pupae. Thissuggests that lethal male combat cannot fully explain the lackof sex ratio shift observed in Melittobia species. We discussalternative explanations.     

Setting the absolute tempo of biodiversity dynamics

Neutral biodiversity theory has the potential to contribute to our understanding of how macroevolutionary dynamics influence contemporary biodiversity, but there are issues regarding its dynamical predictions that must first be resolved. Here we address these issues by extending the theory in two ways using a novel analytical approach: (1) we set the absolute tempo of biodiversity dynamics by explicitly incorporating population-level stochasticity in abundance; (2) we allow new species to arise with more than one individual. Setting the absolute tempo yields quantitative predictions on biodiversity dynamics that can be tested using contemporary and fossil data. Allowing incipient-species abundances greater than one individual yields predictions on how these dynamics, and the form of the species-abundance distribution, are affected by multiple speciation modes. We apply this new model to contemporary and fossil data that encompass 30 Myr of macroevolution for planktonic foraminifera. By synthesizing the model with these empirical data, we present evidence that dynamical issues with neutral biodiversity theory may be resolved by incorporating the effects of environmental stochasticity and incipient-species abundance on biodiversity dynamics.     

Stable isotopes reveal links between human food inputs and urban ant diets

The amount of energy consumed within an average city block is an order of magnitude higher than that consumed in any other ecosystem over a similar area. This is driven by human food inputs, but the consequence of these resources for urban animal populations is poorly understood. We investigated the role of human foods in ant diets across an urbanization gradient in Manhattan using carbon and nitrogen stable isotopes. We found that some—but not all—ant species living in Manhattan''s most urbanized habitats had δ¹³C signatures associated with processed human foods. In particular, pavement ants (Tetramorium sp. E) had increased levels of δ¹³C similar to δ¹³C levels in human fast foods. The magnitude of this effect was positively correlated with urbanization. By contrast, we detected no differences in δ¹⁵N, suggesting Tetramorium feeds at the same trophic level despite shifting to human foods. This pattern persisted across the broader ant community; species in traffic islands used human resources more than park species. Our results demonstrate that the degree urban ants exploit human resources changes across the city and among species, and this variation could play a key role in community structure and ecosystem processes where human and animal food webs intersect.     

Composition of symbiotic bacteria predicts survival in Panamanian golden frogs infected with a lethal fungus

Symbiotic microbes can dramatically impact host health and fitness, and recent research in a diversity of systems suggests that different symbiont community structures may result in distinct outcomes for the host. In amphibians, some symbiotic skin bacteria produce metabolites that inhibit the growth of Batrachochytrium dendrobatidis (Bd), a cutaneous fungal pathogen that has caused many amphibian population declines and extinctions. Treatment with beneficial bacteria (probiotics) prevents Bd infection in some amphibian species and creates optimism for conservation of species that are highly susceptible to chytridiomycosis, the disease caused by Bd. In a laboratory experiment, we used Bd-inhibitory bacteria from Bd-tolerant Panamanian amphibians in a probiotic development trial with Panamanian golden frogs, Atelopus zeteki, a species currently surviving only in captive assurance colonies. Approximately 30% of infected golden frogs survived Bd exposure by either clearing infection or maintaining low Bd loads, but this was not associated with probiotic treatment. Survival was instead related to initial composition of the skin bacterial community and metabolites present on the skin. These results suggest a strong link between the structure of these symbiotic microbial communities and amphibian host health in the face of Bd exposure and also suggest a new approach for developing amphibian probiotics.