On the Breeding Biology of Glyphidrilus annandalei MICHAELSON (Haplotaxida: Glossoscolecidae) and its Relation to the Various Physical and Chemical Factors

Glyphidrilus annandalei Michaelson exhibits a pronounced breeding period extending from September to November. Breeding individuals achieve their maximum frequency in September, when 50 per cent of the whole population was with fully developed clitellum. Studies on the physical and chemical conditions of the habitat revealed that breeding in this worm depends the monsoon of the area creating optimum conditions for the life and activities of the organism.     

Stereoselective synthesis of a ketohexofuranose from an aldohexopyranose by a [6+1-1] strategy

Ozonolysis of 2-acetoxymethyl-1,5-anhydro-3,4,6-tri-O-benzyl-2-deoxy-D-arabino-hex-1-enitol gave 1-O-acetyl-3,4,6-tri-O-benzyl-4-O-formyl-D-arabino-hex-2-ulose (5). Subsequent hydrolysis and acetylation of 5 provided 1,2-di-O-acetyl-3,4,6-tri-O-benzyl-D-fructofuranose 6 in excellent yield. This methodology allows specific deuteration at C-1 of a protected D-fructofuranose derivative. This approach therefore could serve as [6+1-1] formulation for hexose series inter-conversion, that is, aldohexopyranose to ketohexofuranose.     

Discovery of 3(S)-thiomethyl pyrrolidine ERK inhibitors for oncology

Compound 5 (SCH772984) was identified as a potent inhibitor of ERK1/2 with excellent selectivity against a panel of kinases (0/231 kinases tested @ 100?nM) and good cell proliferation activity, but suffered from poor PK (rat AUC PK @10?mpk?=?0?μM?h; F%?=?0) which precluded further development. In an effort to identify novel ERK inhibitors with improved PK properties with respect to 5, a systematic exploration of sterics and composition at the 3-position of the pyrrolidine led to the discovery of a novel 3(S)-thiomethyl pyrrolidine analog 28 with vastly improved PK (rat AUC PK @10?mpk?=?26?μM?h; F%?=?70).     

Predictive modeling of single pass tangential flow filtration for continuous biomanufacturing

Opportunities for process intensification have made continuous biomanufacturing an area of active research. While tangential flow filtration (TFF) is typically employed within the biologics purification train to increase drug substance concentration, single-pass TFF (SPTFF) modifies its format by enabling continuity of this process and achieving a multifold concentration factor through a single-pass over the filtration membranes. In continuous processes feed concentration and flow rate are determined by the preceding unit operations. Therefore, tight control of SPTFF output concentration must be achieved through precise design of the membrane configuration, unlike TFF. However, predictive modeling can be utilized to identify configurations that achieve a desired target concentration across ranges of possible feed conditions with minimal experimental data, hence enabling accelerated process development and design flexibility. We hereby describe the development of a mechanistic model predicting SPTFF performance across a wide design space using the well-established stagnant film model, which we demonstrate is more accurate at higher feed flow rates. The flux excursion dataset was generated within time constraints and with minimal material consumption, showing the method's ability to be quickly adapted. While this approach eliminates characterizing complex physicochemical model variables or the need for users with specialized training, the model and its assumptions become inaccurate at low flow rates, below 25 L/m²/h, and high conversions, above 0.9. As this low flow rate, high conversion operating regime is relevant for continuous biomanufacturing, we explore the assumptions and challenges involved in predicting and modeling SPTFF processes, while suggesting added characterization to gain further process insight.     

Footprinting of the spinach nitrite reductase gene promoter reveals the preservation of nitrate regulatory elements between fungi and higher plants

Nitrite reductase (NiR) is the second enzyme in the nitrate assimilatory pathway reducing nitrite to ammonium. The expression of the NiR gene is induced upon the addition of nitrate. In an earlier study, a 130 bp upstream region of the spinach NiR gene promoter, located between –330 to –200, was shown to be necessary for nitrate induction of -glucuronidase (GUS) expression in tissue-specific manner in transgenic tobacco plant [28]. To further delineate the cis-acting elements involved in nitrate regulation of NiR gene expression, transgenic tobacco plants were generated with 5 deletions in the–330 to –200 region of the spinach NiR gene promoter fused to the GUS gene. Plants with the NiR promoter deleted to –230 showed a considerable increase in GUS activity in the presence of nitrate, indicating that the 30 bp region between –230 to –200 is crucial for nitrate-regulated expression of NiR. In vivo DMS footprinting of the –300 to –130 region of the NiR promoter in leaf tissues from two independent transgenic lines revealed several nitrate-inducible footprints. Footprinting within the –230 to –181 region revealed factor binding to two adjacent GATA elements separated by 24 bp. This arrangement of GATA elements is analogous to cis-regulatory sequences found in the promoters of nitrate-inducible genes of Neurospora crassa, regulated by the NIT2 Zn-finger protein. The –240 to –110 fragment of the NiR promoter, which contains two NIT2 consensus core elements, bound in vitro to a fusion protein comprising the zinc finger domain of the N. crassa NIT2 protein. The data presented here show that nitrate-inducible expression of the NiR gene is mediated by nitrate-specific binding of trans-acting factors to sequences preserved between fungi and higher plants.     

Molecular Characterization of an Arabidopsis Gene Encoding Hydroperoxide Lyase, a Cytochrome P-450 That Is Wound Inducible

Hydroperoxide lyase (HPL) cleaves lipid hydroperoxides to produce volatile flavor molecules and also potential signal molecules. We have characterized a gene from Arabidopsis that is homologous to a recently cloned HPL from green pepper (Capsicum annuum). The deduced protein sequence indicates that this gene encodes a cytochrome P-450 with a structure similar to that of allene oxide synthase. The gene was cloned into an expression vector and expressed in Escherichia coli to demonstrate HPL activity. Significant HPL activity was evident when 13S-hydroperoxy-9(Z),11(E),15(Z)-octadecatrienoic acid was used as the substrate, whereas activity with 13S-hydroperoxy-9(Z),11(E)-octadecadienoic acid was approximately 10-fold lower. Analysis of headspace volatiles by gas chromatography-mass spectrometry, after addition of the substrate to E. coli extracts expressing the protein, confirmed enzyme-activity data, since cis-3-hexenal was produced by the enzymatic activity of the encoded protein, whereas hexanal production was limited. Molecular characterization of this gene indicates that it is expressed at high levels in floral tissue and is wound inducible but, unlike allene oxide synthase, it is not induced by treatment with methyl jasmonate.     

Analysis of cis-acting DNA elements mediating induction and repression of the spinach nitrite reductase gene

The expression of nitrite reductase (NiR; EC 1.7.7.1), the second enzyme in the nitrate assimilatory pathway, is regulated by nitrate as well as by end-products of nitrate assimilation, namely, glutamine (Gln) and asparagine (Asn). Nitrate induces expression of the NiR gene. Previously, using deletion analysis of the spinach (Spinacia oleracea L.) NiR gene promoter in transgenic tobacco (Nicotiana tabacum L.) and in-vivo dimethyl sulfate footprinting, we had identified the region between −230 bp and −180 bp as being critical for nitrate inducibility of this gene. In the present study, we show that the region from +1 to +67, which forms part of its untranslated leader, is important for minimal induction in the presence of nitrate. Electrophoretic mobility shift assays reveal concentration-dependent and competitive binding of a factor in tobacco nuclear extracts to this region. In the presence of Gln or Asn, the expression of spinach NiR is repressed. This repression is observed with the full-length NiR promoter (−3100 bp) as well as with the shortest promoter (−230 bp) that gives nitrate induction, which includes the +67 bp leader sequence. The repressed expression of the gene is not the result of reduced nitrate accumulation in the presence of the nitrogen metabolites. Received: 2 December 1997 / Accepted: 20 January 1998     

Discovery of 3-morpholino-imidazole[1,5-a]pyrazine BTK inhibitors for rheumatoid arthritis

8-Amino-imidazo[1,5-a]pyrazine-based Bruton’s tyrosine kinase (BTK) inhibitors, such as 6, exhibited potent inhibition of BTK but required improvements in both kinase and hERG selectivity (Liu et al., 2016; Gao et al., 2017). In an effort to maintain the inhibitory activity of these analogs and improve their selectivity profiles, we carried out SAR exploration of groups at the 3-position of pyrazine compound 6. This effort led to the discovery of the morpholine group as an optimized pharmacophore. Compounds 13, 23 and 38 displayed excellent BTK potencies, kinase and hERG selectivities, and pharmacokinetic profiles.     

Effects of Nutrients at Different Salinities on Growth of the Freshwater Green Alga Scenedesmus bijugatus in Water of Narendra Pond,Puri, Orissa

The growth and biochemical composition of Scenedesmus bijugatus grown in normal and nutrient enriched Narendra Pond water was evaluated at different levels of salinity. In normal pond water (NPW) the growth was enhanced when the initial salinity of the water was doubled (to 7.2 mmol/l) but further salinity increase caused growth retardation. In nutrient enriched pond water (NEW), however, the maximum growth occurred in the culture containing 29 mmol/l of NaCl. At growth retarding salt concentrations the chlorophyll, carbohydrate, protein and lipid contents per cell decreased in both NPW and NEW while the proline content first increased and was then reduced. All nutrients, taken separately or in combination, caused growth enhancement of the alga at the three selected salinity levels. The alga showed a better halotolerance when enriched with nitrogen or phosphorus.     

Discovery of novel BTK inhibitors with carboxylic acids

We report the design and synthesis of a series of novel Bruton’s Tyrosine Kinase (BTK) inhibitors with a carboxylic acid moiety in the ribose pocket. This series of compounds has demonstrated much improved off-target selectivities including adenosine uptake (AdU) inhibition compared to the piperidine amide series. Optimization of the initial lead compound 4 based on BTK enzyme inhibition, and human peripheral blood mononuclear cell (hPBMC) and human whole blood (hWB) activity led to the discovery of compound 40, with potent BTK inhibition, reduced off target activities, as well as favorable pharmacokinetic profile in both rat and dog.