22-Oxocholestane oximes as potential anti-inflammatory drug candidates.

22-Oxocholestanes bearing the oxime functionality in the side chain have been synthesized from diosgenin and evaluated in vivo as anti-inflammatory agents in an acute inflammation mouse ear model, against the commercial glucocorticoid dexamethasone. The final compounds were all regioselectively obtained with an E configuration at the oxime double bond. The title compounds reduced ear-induced inflammation and edema. The most active oximes repressed the expression of proinflammatory genes TNF-α, COX-2, and IL-6; including macrophage migration inhibitory factor. Overall, our data suggest that 22-oxocholestane oximes exert a strong in vivo anti-inflammatory activity.

Steroid-Based Amphiphiles for Membrane Protein Study: The Importance of Alkyl Spacers for Protein Stability.

Membrane proteins allow effective communication between cells and organelles and their external environments. Maintaining membrane protein stability in a non-native environment is the major bottleneck to their structural study. Detergents are widely used to extract membrane proteins from the membrane and to keep the extracted protein in a stable state for downstream characterisation. In this study, three sets of steroid-based amphiphiles-glyco-diosgenin analogues (GDNs) and steroid-based pentasaccharides either lacking a linker (SPSs) or containing a linker (SPS-Ls)-have been developed as new chemical tools for membrane protein research. These detergents were tested with three membrane proteins in order to characterise their ability to extract membrane proteins from the membrane and to stabilise membrane proteins long-term. Some of the detergents, particularly the SPS-Ls, displayed favourable behaviour with the tested membrane proteins. This result indicates the potential utility of these detergents as chemical tools for membrane protein structural study and a critical role of the simple alkyl spacer in determining detergent efficacy.

Catalytic cyclometallation in steroid chemistry IV: Efficient method for the synthesis of tetrahydrothiophene, tetrahydroselenophen and cyclopentanone derivatives of (5α)-cholestane.

Catalytic cycloalumination of (3ß,5α)-3-vinylcholestane and (3α,5α)-3-allylcholestane with Et3Al catalyzed by Cp2ZrCl2 was performed for the first time to give previously unknown aluminacyclopentanes in ∼90% yield; these products were converted in situ to carbo- and heterocyclic (5α)-cholestane derivatives.

In vitro cytotoxcity and interaction of new steroidal oxadiazinanones with calf thymus DNA using molecular docking, gel electrophoresis and spectroscopic techniques.

Herein we report synthesis of new steroidal oxadiazinanones from steroidal ketones. After characterization by spectral and analytical data, the interaction studies of compounds (4-6) with DNA were carried out by UV-vis, fluorescence spectroscopy and gel electrophoresis. The compounds bind to DNA preferentially through electrostatic and hydrophobic interactions with Kb; 1.8×10(4) M(-1), 2.2×10(4) M(-1) and 2.6×10(4) M(-1), respectively, indicating the higher binding affinity of compound 6 towards DNA. Gel electrophoresis showed the concentration dependent cleavage activity of compound 6 alone or in presence of Cu (II) causes the nicking of supercoiled pBR322 and it seems to follow the mechanistic pathway involving generation of hydroxyl radicals that are responsible for initiating DNA strand scission. Molecular simulations suggest that compounds binds through minor groove of DNA. MTT assay depicted promising anticancer activity of compound 5 and 6 particularly against HL-60 and MCF-7. The apoptotic degradation of DNA was analyzed by agarose gel electrophoresis and visualized by ethidium bromide staining (comet assay). The results revealed that compound 6 has better prospectus to act as cancer chemotherapeutic candidate which warrants further in vivo anticancer investigations.

22-Oxocholestanes as plant growth promoters.

The spirostanic steroidal side-chain of diosgenin and hecogenin was modified to produce 22-oxocholestane derivatives. This type of side-chain was obtained in good yields through a straightforward four-step pathway. These compounds show potent brassinosteroid-like growth promoting activity evaluated via the rice lamina joint inclination bioassay. This is the first report of steroidal skeletons bearing the 22-oxocholestane side-chain and preserving the basic structure (A-D rings) from their corresponding parent compounds acting as plant growth promoters.

Functional properties of Claramine: a novel PTP1B inhibitor and insulin-mimetic compound.

Protein tyrosine phosphatase 1B (PTP1B) inhibits insulin signaling, interfering with its control of glucose homeostasis and metabolism. PTP1B activity is elevated in obesity and type 2 diabetes and is a major cause of insulin resistance. Trodusquemine (MSI-1436) is a "first-in-class" highly selective inhibitor of PTP1B that can cross the blood-brain barrier to suppress feeding and promote insulin sensitivity and glycemic control. Trodusquemine is a naturally occurring cholestane that can be purified from the liver of the dogfish shark, Squalus acanthias, but it can also be manufactured synthetically by a fairly laborious process that requires several weeks. Here, we tested a novel easily and rapidly (2 days) synthesized polyaminosteroid derivative (Claramine) containing a spermino group similar to Trodusquemine for its ability to inhibit PTP1B. Like Trodusquemine, Claramine displayed selective inhibition of PTP1B but not its closest related phosphatase TC-PTP. In cultured neuronal cells, Claramine and Trodusquemine both activated key components of insulin signaling, with increased phosphorylation of insulin receptor-ß (IRß), Akt and GSK3ß. Intraperitoneal administration of Claramine or Trodusquemine effectively restored glycemic control in diabetic mice as determined by glucose and insulin tolerance tests. A single intraperitoneal dose of Claramine, like an equivalent dose of Trodusquemine, suppressed feeding and caused weight loss without increasing energy expenditure. In summary, Claramine is an alternative more easily manufactured compound for the treatment of type II diabetes.

Synthesis and evaluation of some new aza-B-homocholestane derivatives as anticancer agents.

Using analogues of some marine steroidal oximes as precursors, a series of aza-B-homocholestane derivatives possessing different substituted groups at the 3-position of the steroidal nucleus were synthesized. Their biological activity against cancer cell proliferation was determined with multiple cancer cell lines. Aza-B-homocholestane derivatives possessing 3-hydroxyl, 3-hydroximino and 3-thiosemicarbazone groups displayed remarkable cytotoxicity to cancer cells via apoptosis inducing mechanism. Compounds 5, 10, 12, 15 and 18 exhibited better potency to inhibit cancer cell proliferation. In addition, compound 15 was further evaluated with three dimensional (3D) multicellular spheroids assay to determine its potency against spheroid growth. The structure-activity relationship (SAR) generated in the studies is valuable for the design of novel chemotherapeutic agents.

Synthesis and biological studies of steroidal pyran based derivatives.

Steroid based cancer chemotherapeutic agents of the type 2'-amino-3'-cyanocholest-6-eno[5,7-de]4H-pyrans (1c-3c) have been synthesized and characterized by the various spectroscopic and analytical techniques. The DNA binding studies of compounds (1c-3c) with CT DNA were carried out by UV-vis and fluorescence spectroscopy and gel electrophoresis. The compounds (1c-3c) bind to DNA preferentially through electrostatic and hydrophobic interactions with Kb values found to be 5.4 × 10(3), 2.3 × 10(3)M(-1) and 1.97 × 10(3)M(-1), respectively indicating the higher binding affinity of compound (1c) towards DNA. The molecular docking study suggested that the electrostatic interaction of compounds (1c-3c) in between the nucleotide base pairs is due to the presence of pyran moiety in steroid molecule. All the compounds (1c-3c) cleave supercoiled pBR322 DNA via hydrolytic pathway, as validated by T4 DNA ligase assay. The compounds (1c-3c) were screened for in vitro cytotoxicity against the cancer and non-cancer cells SW480, A549, HepG2, HeLa, MCF-7, HL-60, DU-145, NL-20, HPC and HPLF by MTT assay. The compounds (1c-3c) were tested for genotoxicity (comet assay) involving apoptotic degradation of DNA and was analyzed by agarose gel electrophoresis and visualized by ethidium bromide staining. The results revealed that compound (1c) has better prospectus to act as cancer chemotherapeutic candidate which warrants further in vivo anticancer investigations.

Synthesis and anti-tumor evaluation of B-ring substituted steroidal pyrazoline derivatives.

The synthesis and anti-tumor activity screening of new steroidal derivatives (4-18) containing pharmacologically attractive pyrazoline moieties are performed. During in vitro anticancer evaluation, the newly synthesized compounds displayed moderate to good cytotoxicity on cervical and leukemia cancer cell lines. In addition these compounds were found to be nontoxic to normal cell (PBMCs) (IC50>50 µM). The structure-activity relationship is also discussed. The most effective anticancer compound 9 was found to be active with IC50 value of 10.6 µM. It demonstrated significant antiproliferative influence on Jurkat cell lines. The morphological changes and growth characteristics of HeLa cells treated with compound 4 were analyzed by means of SEM.

Catalytic cycloalumination in steroid chemistry: the introduction of a spirotetrahydrofuran or spirotetrahydroselenophene moiety into a 3'-methylene-(5α)-spirocholestane-3,1'-cyclobutane molecule.

Cycloalumination of 3'-methylene-(5α)-spirocholestane-3,1'-cyclobutane with triethylaluminum catalyzed by Cp(2)ZrCl(2) was accomplished for the first time to give (5α)-spirocholestane-3,1'-(6'-ethyl-6'-aluminaspiro [3.4] octane) in 89%. The latter, without isolation, was converted into spirotetrahydroselenophene or spirotetrahydrofuran.

Lipases in green chemistry: acylation and alcoholysis on steroids and nucleosides.

In this article, we describe the application of lipases in acylation and alcoholysis reactions on steroids and nucleosides. In the field of steroids, a variety of acetyl and fatty acid derivatives of androstanes, pregnanes, and cholestanes have been prepared through lipase-catalyzed acylation and alcoholysis reactions taking advantage of the high regio- and stereoselectivity of these enzymes. The substrates as well as the products show a high degree of biological activity as neurosteroids, hormones, and glucocorticoids. The regioselective preparation of diacylated nucleosides by means of an enzymatic alcoholysis allowed the synthesis of nucleosides prodrugs or modified nucleosides. The quantitative full deacylation and dealkoxycarbonylation of nucleosides and steroids is a mild synthetic method for the deprotection of these labile compounds. Some of the reported steroid and nucleoside products are novel, and it is not possible to obtain them satisfactorily by following traditional synthetic procedures. The advantages presented by this methodology, such as selectivity, mild reaction conditions, and low environmental impact, make the lipases an important tool in the application of the principles of Green Chemistry, offering a convenient way to prepare derivatives of natural compounds with a great potential in the pharmaceutical industry.

Synthesis and biological activity of 23-ethylidene-26-hydroxy-22-oxocholestane derivatives from spirostanic sapogenins.

The synthesis and biological evaluation of three new cholestane frameworks of the type: (25R)-3ß,16ß-Diacetoxy-23-ethylidene-26-hydroxy-22-oxocholestane, starting from spirostanic sapogenins of the 25R series, is described. The compounds were obtained by the reductive cleavage of the F ring of 22-oxo-23(1),26-epoxycholestane derivatives using 9-BBN. These modified derivatives exhibit cytotoxic activity against CEM and MCF7 cells and are able to induce apoptosis in them. Its effect on the cell cycle was determined.

Synthesis of [1,2,4]-triazolo-annulated 3-aza-A-homocholestanes--a novel class of pentacyclic compounds.

This study was performed to investigate the reactivity of azocarbenium salts derived from 5α-cholestan-3-one towards 1,3-dipolar cycloaddition reactions with inverse electron-demand to produce unprecedented steroidal heterocyclic derivatives, i.e. [1,2,4]-triazolo-annulated 3-aza-A-homocholestanes 8 and 11 and picrates 12. The synthetic steps were comprised of oxidizing hydrazones 3 with tert-butyl hypochlorite to germinal chloroazo compounds 4, generation of the 1-aza-2-azoniaallene cations 5 by action with equimolar antimony pentachloride and interception with nitrile and alkyne molecules by cycloaddition to the triple bond followed by ring enlargement. The structure of the compounds was principally established on the basis of the analytical and spectral data along with the previously published X-ray diffraction analysis on 8a.

Synthesis and antiproliferative activity of some steroidal lactone compounds.

Using cholesterol as starting material, some steroidal lactone compounds with the structures of 3-substituted-6-oxo-7-oxa-B-homo-cholestane or 3-substituted-7-oxo-6-oxa-B-homo-cholestane were synthesized by oxidation, reduction, Baeyer-Villiger reaction and condensation reaction. The cytotoxicity of these compounds against MGC 7901 (human gastric carcinoma), HeLa (human cervical carcinoma) and SMMC 7404 (human liver carcinoma) cells was investigated. Our results showed that the synthesized compounds displayed a distinct cytotoxicity against these cancer cells. In particular, compounds 8 and 9 have similar cytotoxic capability as cisplatin does. The information obtained from the studies may be useful for the design of novel chemotherapeutic drugs.

Synthesis of new 3,20-bispolyaminosteroid squalamine analogues and evaluation of their antimicrobial activities.

3,20-Amino- and polyaminosteroid analogues of squalamine and trodusquemine were synthesized involving a stereoselective titanium reductive amination reaction in high chemical yields in numerous cases. These derivatives were evaluated for their in vitro antimicrobial properties against references and clinical bacterial strains exhibiting minimum inhibitory concentrations of 2.5-40 µg/mL. The mechanism of action of these derivatives was determined using bioluminescence for ATP efflux measurements and fluorescence methods for membrane depolarization assays.

Synthesis of cholestane saponins as mimics of OSW-1 and their cytotoxic activities.

To fulfill the structure-activity relationship (SAR) of OSW-1, and aim at finding the simplest structural part while maintaining most of the biological activities, six cholestane saponins were synthesized by introducing OSW-1 disaccharide (2-O-4-methoxybenzoyl-ß-D-xylopyranosyl-(1→3)-2-O-acetyl-α-L-arabinopyranosyl) and its 1→4-linked analogue to the 7-hydroxy or 16-hydroxy of steroidal sapogenins. Cytotoxic activities of the products were tested. Compounds 1 and 3 exhibited potent cytotoxicities against five types of human tumor cells, with minimum IC(50) of 2.0 and 75 nM, respectively. And due to its high activity and easy accessibility compound 1 could be a potential candidate for new anti-tumor agents.

The synthesis of cholestane and furostan saponin analogues and the determination of sapogenin's absolute configuration at C-22.

A facile and efficient way for the synthesis of cholestane and furostan saponin analogues was established and adopted for the first time. Following this strategy, starting from diosgenin, three novel cholestane saponin analogues: (22S,25R)-3ß,22,26-trihydroxy-cholest-5-ene-16-one 22-O-[O-α-L-rhamnopyranosyl-(1→2)-ß-D-glucopyranoside] 11, (25R)-3ß,16ß,26-trihydroxy-cholest-5-ene-22-one 16-O-[O-α-L-rhamnopyranosyl-(1→2)-α-D-glucopyranoside] 14 and (25R)-3ß,16ß,26-trihydroxy-cholest-5-ene-22-one 16-O-[O-α-L-rhamnopyranosyl-(1→2)-ß-D-glucopyranoside] 17, three novel furostan saponin analogues: (22S,25R)-furost-5-ene-3ß,22,26-triol 22-O-(α-D-glucopyranoside) 23, (22R,25R)-furost-5-ene-3ß,22,26-triol 22-O-(α-D-glucopyranoside) 24 and (22S,25R)-furost-5-ene-3ß,22,26-triol 22-O-[O-α-L-rhamnopyranosyl-(1→2)-α-D-glucopyranoside] 26, were synthesized ultimately. The structures of all the synthesized analogues were confirmed by spectroscopic methods. The S-chirality at C-22 of cholestane was confirmed by Mosher's method. The absolute configuration at C-22 of furostan saponin analogues was distinguished by conformational analysis combined with the NMR spectroscopy. The cytotoxicities of the synthetic analogues toward four types of tumor cells were shown also.

Novel and efficient synthesis and antifungal evaluation of 2,3-functionalized cholestane and androstane derivatives.

Synthetic modifications of cholesterol and other traditional steroid molecules have become a promising area for the exploration and development of novel antifungal agents, especially with respect to the development of fatty-acid esters of steroids. In addition, 2,3-functionalized steroids are also compounds with potentially interesting biological properties and proper functionalization of 2,3-steroids can lead to the development of efficient syntheses of building blocks for novel fatty-acid esters of steroids. In this Letter, we outline a novel and efficient approach to the synthesis of 2,3-functionalized cholestane and androstane derivatives and present their promising preliminary antifungal activities against a number of fungal species.

Synthesis of cholestane glycosides bearing OSW-1 disaccharide or its 1-->4-linked analogue and their antitumor activities.

For further structure-activity relationship (SAR) research of OSW saponins, a cholestane glycoside, namely 3beta, 16beta, 26-trihydroxycholest-5-en-22-one 16-O-(2-O-4-methoxybenzoyl-beta-D-xylopyranosyl)-(1-->3)-2-O-acetyl-alpha-L-arabinopyranoside (1) together with two 1-->4-linked disaccharide analogues (2 and 3) were synthesized. Their cytotoxic activities were evaluated by the standard MTT assay. Compound 1 showed potent cytotoxicity against five types of human tumor cells, with IC50 ranging between 1.3 and 73 nM.

Synthesis, antibacterial and antifungal activities of 6,5 fused steroidal oxazoles in cholestane series.

Herein, we report a convenient one-pot synthesis of 2'-amino-5alpha-cholest-6-eno [6,5-d] oxazole derivatives (4-6). The synthesis involves the reaction of cholestan-6-ones (1-3) with urea and iodine. The structural assignment of the products was confirmed on the basis of IR, (1)H NMR, (13)C NMR, and Mass spectra which find support from comparison with authentic samples. The antibacterial activity of all the synthesized compounds was tested in vitro by the disk diffusion assay against three Gram-positive and three Gram-negative strains of bacteria. All the synthesized compounds were also tested for their inhibitory action against five strains of fungus and then the minimum inhibitory concentration (MIC) of all the synthesized compounds were determined. Compounds (4-6) showed inhibitory action against both types of the bacteria (Gram-positive and Gram-negative) and five strains of fungi are good antimicrobial agents. Chloramphenicol (30 microg) was used as standard drug in case of bacteria and nystatin was used as a standard drug in case of fungi.