Life Sciences

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We examined the effects of an inhibitor of PI3K, XL147, against human breast cancer cell lines with constitutive PI3K activation. Treatment with XL147 resulted in dose-dependent inhibition of cell growth and levels of pAKT and pS6, signal transducers in the PI3K/AKT/TOR pathway. In HER2-overexpressing cells, inhibition of PI3K was followed by up-regulation of expression and phosphorylation of multiple receptor tyrosine kinases, including HER3. Knockdown of FoxO1 and FoxO3a transcription factors suppressed the induction of HER3, InsR, IGF1R, and FGFR2 mRNAs upon inhibition of PI3K. In HER2 + cells, knockdown of HER3 with siRNA or cotreatment with the HER2 inhibitors trastuzumab or lapatinib enhanced XL147-induced cell death and inhibition of pAKT and pS6. Trastuzumab and lapatinib each synergized with XL147 for inhibition of pAKT and growth of established BT474 xenografts. These data suggest that PI3K antagonists will inhibit AKT and relieve suppression of receptor tyrosine kinase expression and their activity. Relief of this feedback limits the sustained inhibition of the PI3K/AKT pathway and attenuates the response to these agents. As a result, PI3K pathway inhibitors may have limited clinical activity overall if used as single agents. In patients with HER2- overexpressing breast cancer, PI3K inhibitors should be used in combination with HER2/HER3 antagonists.

(SoNS) School of Natural Sciences

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Proceedings of the National Academy of Sciences of the United States of America

Feedback upregulation of HER3 (ErbB3) expression and activity attenuates antitumor effect of PI3K inhibitors

Forty eight mango hybrids were analyzed using 17 simple sequence repeat (SSR) markers. Which detected 59 scorable loci, of which 45 were polymorphic. The size of the alleles detected ranged from 100 to 480 bp. SSR markers was highly polymorphic with an average of 3.47 alleles per primer. SSRs gave moderate values of Polymorphis Information Content (PIC) and heterozygosity. The genetic relationship among mango hybrids, based on Jaccard's Similarity Coefficient values ranged from 0.38 (between H-1-13 and H-6-8) to 0.97 (between H-13-4 and H-13-7). The dendrogram, based on UPGMA cluster analysis, grouped the mango hybrids into three major groups. Cluster 'A' comprised of the five most diverse hybrids, namely, H-12-2, H-9-6, Pusa Arunima, H-9-5 and H-1-13. Two unique fingerprints were identified in hybrid H-9-6. The size of unique fingerprints ranged from 90 (MiSHRS-18) to 350 bp (MiSHRS-39). The Principal Coordinate Analysis also exhibited more or less similar distribution of mango hybrids. The tendency of clustering among mango hybrids revealed that they had stronger affinity towards female parent Amrapali.

Indian Journal of Horticulture

SSR markers reveal genetic diversity in closely related mango hybrids

Excellent studies have been carried by Indian scientists to understand paleoceanographic changes in the Indian Ocean including die Arabian Sea and Bay of Bengal using Ocean Drilling Program cores. These studies revealed both short-term and long-term changes in the Indian Ocean circulation and Indian monsoon system. In general, summer monsoon was intense during warm intervals and weaker during cold intervals. Changes in deep-sea were also observed. The above studies were based on planktic and benthic foraminiferal faunal and geochemical data. Changes in deep sea oxygenation, oxygen minimum zone and surface productivity in the Indian Ocean including the Arabian Sea and Bay of Bengal during the Neogene and Quatemay were driven by the Indian summer monsoon. These changes in nutrient and oxygen levels drove major changes in the fauna and flora of the Indian Ocean. Indian monsoon variability has been suggested as contemporaneous with the southeast Monsoon variability during the last glacial and the Holocene. © Printed in India.

Proceedings of the Indian National Science Academy

Indian contributions in the field of palaeoceanography (2006-2012)

HisC2 from Mycobacterium tuberculosis was overexpressed in M. smegmatis and purified to homogeneity using nickel-nitrilotriacetic acid metal-affinity and gel-filtration chromatography. Diffraction-quality crystals were grown using the hanging-drop vapour-diffusion technique from a condition consisting of 7 mg ml-1 HisC2 (in 20 mM Tris pH 8.8, 50 mM NaCl and 5% glycerol), 1 M succinic acid pH 7.0, 0.1 M HEPES pH 7.0 and 1%(w/v) polyethylene glycol monomethyl ether 2000. The crystals belonged to the orthorhombic space group P21212, with unit-cell parameters a = 255.98, b = 77.09, c = 117.97 {\AA}. X-ray diffraction data were recorded to 2.45 {\AA} resolution from a single crystal using the in-house X-ray facility. {\textcopyright} 2012 International Union of Crystallography. All rights reserved.

Acta Crystallographica Section F: Structural Biology and Crystallization Communications

Molecular cloning, overexpression, purification, crystallization and preliminary X-ray diffraction studies of histidinol phosphate aminotransferase (HisC2) from Mycobacterium tuberculosis

Aspartate-semialdehyde dehydrogenase (Asd; ASADH; EC 1.2.1.11) is the enzyme that lies at the first branch point in the biosynthetic pathway of important amino acids including lysine and methionine and the cell-wall component diamino-pimelate (DAP). The enzymatic reaction of ASADH is the reductive dephosphorylation of aspartyl-$\beta$-phosphate (ABP) to aspartate $\beta$-semialdehyde (ASA). Since the aspartate pathway is absolutely essential for the survival of many microbes and is absent in humans, the enzymes involved in this pathway can be considered to be potential antibacterial drug targets. In this work, the structure of ASADH from Myco-bacterium tuberculosis H37Rv (Mtb-ASADH) has been determined in complex with glycerol and sulfate at 2.18 {\AA} resolution and in complex with S-methyl-l-cysteine sulfoxide (SMCS) and sulfate at 1.95 {\AA} resolution. The overall structure of Mtb-ASADH is similar to those of its orthologues. However, in the Mtb-ASADH-glycerol complex structure the glycerol molecule is noncovalently bound to the active-site residue Cys130, while in the Mtb-ASADH-SMCS complex structure the SMCS (Cys) is covalently linked to Cys130. The Mtb-ASADH-SMCS complex structurally mimics one of the intermediate steps in the proposed mechanism of ASADH enzyme catalysis. Comparison of the two complex structures revealed that the amino acids Glu224 and Arg249 undergo conformational changes upon binding of glycerol. Moreover, the structures reported here may help in the development of species-specific antibacterial drug molecules against human pathogens. {\textcopyright} 2012 International Union of Crystallography Printed in Singapore - all rights reserved.

Acta Crystallographica Section D: Biological Crystallography

Structures of ternary complexes of aspartate-semialdehyde dehydrogenase (Rv3708c) from Mycobacterium tuberculosis H37Rv

Adhesive forces at endothelial cell-cell borders maintain vascular integrity. cAMP enhances barrier properties and controls cellular processes through protein kinase A bound to A-kinase anchoring proteins (AKAPs). It also activates exchange protein directly activated by cAMP (Epac1), an exchange factor for Ras-related protein 1 (Rap1) GTPases that promotes cadherin- and integrin-mediated adhesion through effects on the actin cytoskeleton. We demonstrate that AKAP9 facilitates the microtubule polymerization rate in endothelial cells, interacts with Epac1, and is required for Epac1-stimulated microtubule growth. AKAP9 is not required for maintaining barrier properties under steady-state conditions. Rather, it is essential when the cell is challenged to make new adhesive contacts, as is the case when Epac activation enhances barrier function through a mechanism that, surprisingly, requires integrin adhesion at cell-cell contacts. In the present study, defects in Epac-induced responses in AKAP9-silenced cells were evident despite an intact Epac-induced increase in Rap activation, cortical actin, and vascular endothelial-cadherin adhesion. We describe a pathway that integrates Epac-mediated signals with AKAP9-dependent microtubule dynamics to coordinate integrins at lateral borders. © 2011 by The American Society of Hematology.

Blood

AKAP9 regulation of microtubule dynamics promotes Epac1-induced endothelial barrier properties

VLF chorus emissions recorded at Indian Antarctic station, Maitri (Lat = 70.46’CS, Long = 11.50’CE, L = 4.5) during a geomagnetic quiet period on 3 Feb. 2001 is reported. The detailed spectral analysis of recorded chorus emissions shows that each chorus element originates from the upper edge of the underlying hiss band. The observed mean chorus element parameters are as follows: lower band frequency fmin = 3.1 kHz, upper band frequency ƒUB = 4.8 kHz, frequency sweep rate df/dt = 2.1 kHz/s and repetition period T = 1 s. To explain the observed dynamic spectra of these chorus emissions, a possible generation mechanism is presented based on the recent nonlinear theory. On the basis of this theory, frequency sweep rate of chorus emission is computed and compared with that of our experimentally observed values, which shows, in general, good agreements. © 2011 by World Scientific Publishing Co. Pte. Ltd. All rights reserved.

Advances in Geosciences: Volume 27: Solar Terrestrial (ST)

Nonlinear generation mechanism of VLF chorus emissions observed at Antarctica during a quiet period

The ErbB receptor family member ErbB3 has been implicated in breast cancer growth, but it has yet to be determined whether its disruption is therapeutically valuable. In a mouse model of mammary carcinoma driven by the polyomavirus middle T (PyVmT) oncogene, the ErbB2 tyrosine kinase inhibitor lapatinib reduced the activation of ErbB3 and Akt as well as tumor cell growth. In this phosphatidylinositol-3 kinase (PI3K)-dependent tumor model, ErbB2 is part of a complex containing PyVmT, p85 (PI3K), and ErbB3, that is disrupted by treatment with lapatinib. Thus, full engagement of PI3K/Akt by ErbB2 in this oncogene-induced mouse tumor model may involve its ability to dimerize with and phosphorylate ErbB3, which itself directly binds PI3K. In this article, we report that ErbB3 is critical for PI3K/Akt-driven tumor formation triggered by the PyVmT oncogene. Tissue-specific, Cre-mediated deletion of ErbB3 reduced Akt phosphorylation, primary tumor growth, and pulmonary metastasis. Furthermore, EZN-3920, a chemically stabilized antisense oligonucleotide that targets the ErbB3 mRNA in vivo, produced similar effects while causing no toxicity in the mouse model. Our findings offer further preclinical evidence that ErbB3 ablation may be therapeutically effective in tumors where ErbB3 engages PI3K/Akt signaling. ©2011 AACR.

Cancer Research

ErbB3 ablation impairs PI3K/Akt-dependent mammary tumorigenesis

The phosphatidylinositol 3-kinase (PI3K)/AKT/mammalian target of rapamycin (mTOR) axis regulates essential cellular functions including cell survival, proliferation, metabolism, migration, and angiogenesis. The PI3K pathway is activated in human cancers by mutation, amplification, and deletion of genes encoding components of this pathway. The critical role of PI3K in cancer has led to the development of drugs targeting the effector mechanisms of this signaling network. Recent studies have shown that inhibition at multiple levels of the PI3K pathway results in FOXOdependent feedback reactivation of several receptor tyrosine kinases (RTKs) which, in turn, limit the sustained inhibition of this pathway and attenuates the action of therapeutic antagonists. This suggests that if used as single agents, PI3K pathway inhibitors may have limited clinical activity. We propose herein that to successfully target the output of the PI3K pathway in cancer cells, combination therapies that hinder these compensatory mechanisms should be used. Thus, combination therapies that target RTKs, PI3K, and mTOR activities may be required to maximize the clinical benefit derived from treatment with these inhibitors. © Garrett et al.

Oncotarget

Will PI3K pathway inhibitors be effective as single agents in patients with cancer?

HisB, encoded by open reading frame Rv1601, possesses enzymatic activity as an imidazoleglycerol-phosphate dehydratase in the histidine-biosynthetic pathway of Mycobacterium tuberculosis. A recombinant form of HisB was crystallized in three crystal forms: crystals grown using 20% PEG 1500 as a precipitant belonged to either the cubic space group P432 or the tetragonal space group P4, while an orthorhombic crystal form belonging to space group P21212 was obtained using 15% PEG 5000 and 10 mM MnCl2 as precipitant. The structure of HisB in the orthorhombic crystal form was solved by the molecular-replacement method using the crystal structure of its Arabidopsis thaliana counterpart, which shares 47% sequence identity with Rv1601, as the search model. {\textcopyright} 2011 International Union of Crystallography. All rights reserved.

HisB from Mycobacterium tuberculosis: Cloning, overexpression in Mycobacterium smegmatis, purification, crystallization and preliminary X-ray crystallographic analysis