Synthesis of Ofornine mimics from natural product l-vasicine as anti-hypertensive agents.

We report the chemical synthesis of Ofornine mimics from l-vasicine, structure-activity relationship studies and their in vivo screening for anti-hypertensive action in Wistar rats. It was observed that most of the analogs possessed anti-hypertensive effect; however, the duration of the effect was variable and mostly transient. The results demonstrated that the analogs 12, 13, 14, 15, and 16 showed a sharp and significant decrease in systolic and diastolic blood pressure for 30-60min after intravenous administration. Analog (S)-(3-hydroxypyrrolidin-1-yl)(2-(pyridin-4-ylamino)phenyl)methanone (8) showed a significant decrease in blood pressure in a dose dependent manner whose maximal response lowered to 79.29±4.26mmHg of SBP and 62.55±2.9 of DBP at 10mg/kg intravenous dose. Further, the significant anti-hypertensive effect of 8 lasted for about 2.5h at 10mg/kg dose. We also evaluated the acute toxicity of the analog 8 as per the OECD guidelines and the compound was found to be safe up to the dose of 2000mg/kg body weight. These preclinical findings suggest that the analog 8 could be considered as a promising lead and a durable anti-hypertensive drug candidate and deserves further investigation. The SAR studies clearly showed that the amide, hydroxyl and pyridine ring plays important role in showing the activity.

Reversal of boswellic acid analog BA145 induced caspase dependent apoptosis by PI3K inhibitor LY294002 and MEK inhibitor PD98059.

PI3K/Akt and ERK pathways are important for growth and proliferation of many types of cancers. Therefore, PI3K inhibitor LY294002 (LY) and MEK1/2 inhibitor PD98059 (PD) are used to sensitize many types of cancer cell lines to chemotherapeutic agents, where AKT and ERK pathways are over activated. However, in this study, we show for the first time that PD could protect the leukemia cells independent of ERK pathway inhibition, besides, we also report a detailed mechanism for antiapoptotic effect of LY in HL-60 cells against the cytotoxicity induced by a boswellic acid analog BA145. Apoptosis induced by BA145 is accompanied by downregulation of PI3K/Akt and ERK pathways in human myelogenous leukemia HL-60 cells, having activating N-Ras mutation. Both LY and PD protected the cells against mitochondrial stress caused by BA145, and reduced the release of cytochrome c and consequent activation of caspase-9. LY and PD also diminished the activation of caspase-8 without affecting the death receptors. Besides, LY and PD also reversed the caspase dependent DNA damage induced by BA145. Further studies revealed that LY and PD significantly reversed the inhibitory effect of BA145 on cell cycle regulatory proteins by upregulating hyperphosphorylated retinoblastoma, pRB (S795) and downregulating p21 and cyclin E. More importantly, all these events were reversed by caspase inhibition by Z-VAD-fmk, suggesting that both LY and PD act at the level of caspases to diminish the apoptosis induced by BA145. These results indicate that inhibitors of PI3K/Akt and ERK pathways can play dual role and act against chemotherapeutic agents.

Tubulin inhibitors from an endophytic fungus isolated from Cedrus deodara.

From an endophytic fungus, a close relative of Talaromyces sp., found in association with Cedrus deodara, four compounds including two new ones (2 and 4) were isolated and characterized. The structures of two compounds (1 and 4) were confirmed by X-ray crystallography. The compounds displayed a range of cytotoxicities against human cancer cell lines (HCT-116, A-549, HEP-1, THP-1, and PC-3). All the compounds were found to induce apoptosis in HL-60 cells, as evidenced by fluorescence and scanning electron microscopy studies. Also, the compounds caused significant microtubule inhibition in HL-60 cells.

A novel cyano derivative of 11-keto-ß-boswellic acid causes apoptotic death by disrupting PI3K/AKT/Hsp-90 cascade, mitochondrial integrity, and other cell survival signaling events in HL-60 cells.

Intervention of apoptosis is a promising strategy for discovery of novel anti-cancer therapeutics. In this study, we examined the ability of a novel cyano derivative of 11-keto-ß-boswellic acid, that is, butyl 2-cyano-3,11-dioxours-1,12-dien-24-oate (BCDD) to induce apoptosis in cancer cells. BCDD inhibited cell proliferation with 48 h IC(50) of 0.67 µM in HL-60, 1 µM in Molt4, and 1.5 µM in THP1 cells. The mechanism of cell death was investigated in HL-60 cells where it caused apoptosis by acting against several potential apoptosis suppressive targets. It inhibited phosphatidylinositol-3-kinase (PI3K)/AKT activity, NF-κB, Hsp-90, and survivin which may enhance the sensitivity of cells to apoptosis. Also, BCDD decreased the activity of Bid and Bax in cytosol, caused ΔΨ(mt) loss, releasing pro-apoptotic cytochrome c, SMAC/DIABLO leading to caspase-9-mediated down stream activation of caspase-3, ICAD, and PARP1 cleavage. Translocation of apoptotis-inducing factor (AIF) from mitochondria to the nucleus indicated some caspases-independent apoptosis. Though it upregulated DR-5 and caspase-8, the caspase inhibitor yet had no effect on apoptosis as against 75% inhibition by caspase-9 inhibitor. Attempts were made to examine any acclaimed role of AIF in the activation of caspase-8 using siRNA where it had no effect on caspase-8 activity while the Bax-siRNA inhibited caspase-3 activation suggesting predominance of intrinsic signaling. Our studies thus demonstrated that BCDD exerts multi-focal action in cancer cells while it required 10-fold higher the concentration to produce cytotoxicity in normal human PBMC and gingival cell line, and therefore, may find usefulness in the management of human leukemia.

Acyl derivatives of boswellic acids as inhibitors of NF-κB and STATs.

Boswellic acid acylates including their epimers were synthesized and screened against a panel of human cancer cell lines. They exhibited a range of cytotoxicity against various human cancer cell lines thereby leading to the development of a possible SAR. One of the identified lead compounds was found to be an inhibitor of the NF-κB and STAT proteins, warranting further investigations to be developed into a potential anticancer lead.

PARP cleavage and perturbance in mitochondrial membrane potential by 3-α-propionyloxy-ß-boswellic acid results in cancer cell death and tumor regression in murine models.

BACKGROUND: Apoptotic induction in cancer cells has become a major focus of anticancer therapeutics. In this regard, ß-boswellic acids, naturally occurring pentacyclic triterpenes, have demonstrated antiproliferative and cytotoxic effects against different types of cancers. Surprisingly, not much has been reported regarding the chemical modifications or preparation of structural analogs of the key constituents of ß-boswellic acid. AIM: The anticancer activity of 3-α-propionyloxy-ß-boswellic acid (POBA) was investigated and this article reports for the first time that the triterpenoid ring of the boswellic acid derivative POBA is targeting the PI3K pathway. MATERIALS & METHODS: Induction of apoptosis of the semi-synthetic derivative of ß-boswellic acid-POBA in vitro was analyzed using a battery of human cancer cell lines followed by cell cycle phase distribution, further validated by DNA fragmentation, and was found to cause mitochondrial membrane potential loss with ultrastructural changes, as observed by electron microscopy studies and expression study using PARP cleavage, as well as validated by in vivo anti-tumor activity. RESULTS: The cytotoxicity data revealed the sensitivity of various human cancer cell lines of varied tissue origin to ß-boswellic acid, which robustly induced cell cycle arrest, DNA fragmentation and loss of mitochondrial membrane potential. Morphological studies of the effects of POBA revealed loss of surface projections, chromatin condensation, apoptotic body formation and POBA-mediated PARP cleavage. For in vivo therapeutic experiments, murine tumor models were treated with POBA and the treatment resulted in a significantly higher level of growth inhibition and apoptosis was significantly induced. CONCLUSION: These findings demonstrate that acyl substituents/groups in the main skeleton of ß-boswellic acid have the potential to be potent chemotherapeutic agents.

1,2,3,4,5,6-Hexa-O-acetyl-scyllo-inositol.

The title mol-ecule, C(18)H(24)O(12), has crystallographic 2/m symmetry with two acetate group located on a mirror plane. The H-Csp(3)-O-Csp(2) torsion angles characterizing orientation of the acetyl groups with respect to the cyclo-hexane ring are 0.0, 23.9 and -23.9°. The cyclo-hexane ring is in a chair conformation with all substituents in equatorial positions. In the crystal, mol-ecules are connected through C-H⋯O hydrogen bonds into a chain extending along the c axis.

Antistaphylococcal and biofilm inhibitory activities of acetyl-11-keto-ß-boswellic acid from Boswellia serrata.

BACKGROUND: Boswellic acids are pentacyclic triterpenes, which are produced in plants belonging to the genus Boswellia. Boswellic acids appear in the resin exudates of the plant and it makes up 25-35% of the resin. ß-boswellic acid, 11-keto-ß-boswellic acid and acetyl-11-keto-ß-boswellic acid have been implicated in apoptosis of cancer cells, particularly that of brain tumors and cells affected by leukemia or colon cancer. These molecules are also associated with potent antimicrobial activities. The present study describes the antimicrobial activities of boswellic acid molecules against 112 pathogenic bacterial isolates including ATCC strains. Acetyl-11-keto-ß-boswellic acid (AKBA), which exhibited the most potent antibacterial activity, was further evaluated in time kill studies, postantibiotic effect (PAE) and biofilm susceptibility assay. The mechanism of action of AKBA was investigated by propidium iodide uptake, leakage of 260 and 280 nm absorbing material assays. RESULTS: AKBA was found to be the most active compound showing an MIC range of 2-8 µg/ml against the entire gram positive bacterial pathogens tested. It exhibited concentration dependent killing of Staphylococcus aureus ATCC 29213 up to 8 × MIC and also demonstrated postantibiotic effect (PAE) of 4.8 h at 2 × MIC. Furthermore, AKBA inhibited the formation of biofilms generated by S. aureus and Staphylococcus epidermidis and also reduced the preformed biofilms by these bacteria. Increased uptake of propidium iodide and leakage of 260 and 280 nm absorbing material by AKBA treated cells of S aureus indicating that the antibacterial mode of action of AKBA probably occurred via disruption of microbial membrane structure. CONCLUSIONS: This study supported the potential use of AKBA in treating S. aureus infections. AKBA can be further exploited to evolve potential lead compounds in the discovery of new anti-Gram-positive and anti-biofilm agents.

2,3-Unsaturated allyl glycosides as glycosyl donors for selective α-glycosylation.

In the presence of NBS and a catalytic amount of a Lewis acid, 2,3-unsaturated allyl glycosides [6-(allyloxy)-3,6-dihydro-2-(hydroxymethyl)-2H-pyran-3-ol] have been successfully used as versatile glycosyl donors for the stereoselective α-glycosylation of a variety of alcohols comprising sensitive functions such as acetonide, keto, nitro, and ester in 50-90% yields. The methodology offers an equally facile alternative to 4-pentenyl replacement in unsaturated sugars.

Entrapment and kinetic resolution of stabilized axial and equatorial conformers of spiro-ß-lactams.

The facile synthesis of the stabilized axial and equatorial conformers of spiro-ß-lactams was achieved via entrapment of cyclohexanone imines (Schiff bases) with acetoxyacetyl chloride in a [2 + 2]-cycloaddition reaction followed by their kinetic resolution. The immobilization of the racemic substrates on an inert solid support significantly reduced the reaction time and improved the enantioselectivity of conformers during kinetic resolution. The mechanism of the formation of the spiro-ß-lactams was explored using B3LYP/6-31+G* level quantum chemical calculations.

Semi-synthetic analogs of pinitol as potential inhibitors of TNF-alpha cytokine expression in human neutrophils.

Semi-synthetic analogs of pinitol were subjected to screening by determining TNF-alpha expression in human neutrophils using flowcytometry. Among the tested compounds, three derivatives displayed more than 50% inhibition of TNF-alpha cytokine secretion in LPS induced stimulated neutrophils and can be considered as potent anti-inflammatory moieties.

Arthrobacter sp. lipase immobilization for preparation of enantiopure masked beta-amino alcohols.

Recent reports on immobilization of lipase from Arthrobacter sp. (ABL, MTCC 5125; IIIM isolate) on insoluble polymers have shown altered properties including stability and enantioselectivity. Present work demonstrates a facile method for the preparation of enantiopure beta-amino alcohols by modulation of ABL enzyme properties via immobilization on insoluble as well as soluble supports using entrapment/covalent binding techniques. Efficacies of immobilized ABL on insoluble supports prepared from tetraethylorthosilicate/aminopropyltriethoxy silane and soluble supports derived from copolymerization of N-vinyl pyrrolidone-allylglycidyl ether (ANP type)/N-vinyl pyrrolidone-glycidyl methacrylate (GNP type) for kinetic resolution of masked beta-amino alcohols have been studied vis-à-vis free ABL enzyme/wet cell biomass. The immobilized lipase on different insoluble/soluble supports has shown 21-110 mg/g protein binding and 30-700 U/g activity for hydrolyzing tributyrin substrate. The findings have shown a significant enhancement in enantioselectivity (ee 99%) vis-à-vis wet cell biomass providing ee 70-90% for resolution of beta-amino alcohols.

Medicinal plants and cancer chemoprevention.

Cancer is the second leading cause of death worldwide. Although great advancements have been made in the treatment and control of cancer progression, significant deficiencies and room for improvement remain. A number of undesired side effects sometimes occur during chemotherapy. Natural therapies, such as the use of plant-derived products in cancer treatment, may reduce adverse side effects. Currently, a few plant products are being used to treat cancer. However, a myriad of many plant products exist that have shown very promising anti-cancer properties in vitro, but have yet to be evaluated in humans. Further study is required to determine the efficacy of these plant products in treating cancers in humans. This review will focus on the various plant-derived chemical compounds that have, in recent years, shown promise as anticancer agents and will outline their potential mechanism of action.

An essential oil and its major constituent isointermedeol induce apoptosis by increased expression of mitochondrial cytochrome c and apical death receptors in human leukaemia HL-60 cells.

An essential oil from a lemon grass variety of Cymbopogon flexuosus (CFO) and its major chemical constituent sesquiterpene isointermedeol (ISO) were investigated for their ability to induce apoptosis in human leukaemia HL-60 cells because dysregulation of apoptosis is the hallmark of cancer cells. CFO and ISO inhibited cell proliferation with 48 h IC50 of approximately 30 and 20 microg/ml, respectively. Both induced concentration dependent strong and early apoptosis as measured by various end-points, e.g. annexinV binding, DNA laddering, apoptotic bodies formation and an increase in hypo diploid sub-G0 DNA content during the early 6h period of study. This could be because of early surge in ROS formation with concurrent loss of mitochondrial membrane potential observed. Both CFO and ISO activated apical death receptors TNFR1, DR4 and caspase-8 activity. Simultaneously, both increased the expression of mitochondrial cytochrome c protein with its concomitant release to cytosol leading to caspase-9 activation, suggesting thereby the involvement of both the intrinsic and extrinsic pathways of apoptosis. Further, Bax translocation, and decrease in nuclear NF-kappaB expression predict multi-target effects of the essential oil and ISO while both appeared to follow similar signaling apoptosis pathways. The easy and abundant availability of the oil combined with its suggested mechanism of cytotoxicity make CFO highly useful in the development of anti-cancer therapeutics.

Cytotoxic and apoptotic activities of novel amino analogues of boswellic acids.

4-Amino analogues prepared from beta-boswellic acid and 11-keto-beta-boswellic acid, wherein the carboxyl group in ursane nucleus was replaced by an amino function via Curtius reaction, displayed improved cytotoxicity than the parent molecules. The same molecules also exhibited apoptotic activity by inducing DNA fragmentation.

Molecular cloning of enantioselective ester hydrolase from Bacillus pumilus DBRL-191.

A gene from Bacillus pumilus expressed under its native promoter was cloned in Escherichia coli. Recombinant B. pumilus esterase (BPE) affects the kinetic resolution of racemic mixtures such as unsubstituted and substituted 1-(phenyl)ethanols (E approximately 33-103), ethyl 3-hydroxy-3-phenylpropanoate (E approximately 45-71), trans-4-fluorophenyl-3-hydroxymethyl-N-methylpiperidine (E approximately 10-13) and ethyl 2-hydroxy-4-phenylbutyrate (E approximately 7). The enzyme is composed of a 34-amino acid signal peptide and a 181-amino acid mature protein corresponding to a molecular weight of approximately 19.2kD and pI approximately 9.4. 3-D the structural model of the enzyme built by homology modelling using the atomic coordinates from the crystal structure of B. subtilis lipase (LipA) showed a compact minimal alpha/beta hydrolase fold.

Capsaicin production by Alternaria alternata, an endophytic fungus from Capsicum annum; LC-ESI-MS/MS analysis.

Alternaria alternata, an endophytic fungus capable of producing capsaicin (1) was isolated from Capsicum annum. The endophyte was found to produce capsaicin upto three generations. Upscaling of the fermentation broth led to the isolation of one known and one compound characterized as 2,4-di-tert-butyl phenol (2) and alternariol-10-methyl ether (3) respectively. Compound 1 and 3 were identified and quantified using liquid chromatography-electrospray ionization tandem mass spectrometry (LC-ESI-MS/MS) system through multiple reaction monitoring (MRM). Furthermore, compound 3 displayed a range of cytotoxicity against a panel of human cancer cell lines and was found to induce apoptosis evidenced by Hoechst staining and loss of mitochondrial-membrane potential in HL-60 cells.

NF-κB down-regulation and PARP cleavage by novel 3-α-butyryloxy-ß-boswellic acid results in cancer cell specific apoptosis and in vivo tumor regression.

The present study relates to the induction of apoptosis thereof cytotoxicity and anti-cancer activity displayed by semi-synthetic analog of Boswellic acid i.e. 3-α-Butyryloxy-ß-boswellic acid (BOBA). The cytotoxicity data revealed the differential sensitivity of cancer cell lines towards BOBA which may display its impact against different types of cancers. Considering the inhibitory potential of BOBA, we further sought to understand the target for BOBA deciphering the mechanism of action leading to apoptotic cell death and it was for the first time reported about the triterpenoid ring especially the ß-boswellic acid derivative is targeting PI3K pathway. Our data revealed that BOBA treatment provides evidence about the apoptotic nature showing the potential of targeting mitochondria dependent pathways during apoptosis in HL-60 cells. BOBA induced hypo-diploid sub-G(1) DNA population in HL-60 cells as was also evident from the pattern of DNA fragmentation and mitochondrial membrane potential (ΛΨm) loss. Morphological analysis under fluorescent and scanning electron microscopy displayed typical features such as cell shrinkage, membrane blebbing, chromatin condensation and nuclear fragmentation. These events paralleled with the down-regulation of NF-κB and induced PARP cleavage. Furthermore, it is noteworthy that BOBA also depicted significant growth inhibition in Ehrlich Ascitic Tumour (EAT), Ehrlich Ascitic Carcinoma (EAC) and Sarcoma- 180 tumour models. Taken together, BOBA treatment may represent as potential agent to the currently available anticancer agents in both prophylactic and/or therapeutic applications. Also, our findings may open up a new perspective in the construction of novel anticancer agents based on boswellic acids that will facilitate the development of these agents for anticancer therapeutics.

Highly regio- and stereoselective one-pot synthesis of carbohydrate-based butyrolactones.

The use of manganese(III) acetate allows the direct synthesis of diverse arrays of [4.3.0] bicyclic carbohydrate-based γ-lactone building blocks from glycals. A mechanism to explain the high regio- and stereoselectivity is proposed. The new reaction has the potential to generate libraries for biological screening.

A novel parthenin analog exhibits anti-cancer activity: activation of apoptotic signaling events through robust NO formation in human leukemia HL-60 cells.

This study describes the anti-cancer activity of P19, an analog of parthenin. P19 induced apoptosis in HL-60 cells and inhibited cell proliferation with 48h IC50 of 3.5µM. At 10mg/kg dose, it doubled the median survival time of L1210 leukemic mice and at 25mg/kg it inhibited Ehrlich ascites tumor growth by 60%. Investigation of the mechanism of P19 induced apoptosis in HL-60 cells revealed that N-acetyl-l-cysteine (NAC) and s-methylisothiourea (sMIT) could reverse several molecular events that lead to cell death by inhibiting nitric oxide (NO) formation. It selectively produced massive NO in cells while quenching the basal ROS levels with concurrent elevation of GSH. P19 disrupted mitochondrial integrity leading to cytochrome c release and caspase-9 activation. P19 also caused caspase-8 activation by selectively elevating the expression of DR4 and DR5. All these events lead to the activation of caspase-3 leading to PARP-1 cleavage and DNA fragmentation. However, knocking down of AIF by siRNA also suppressed the apoptosis substantially thus indicating caspase independent apoptosis, too. Further, contrary to enhanced iNOS expression, its transcription factor, NF-κB (p65) was cleaved with a simultaneous increase in cytosolic IκB-alpha. In addition, P19 potently inhibited pro-survival proteins pSTAT3 and survivin. The multi-modal pro-apoptotic activity of P19 raises its potential usefulness as a promising anti-cancer therapeutic.