Screening one bead one compound libraries against serum using a flow cytometer: Determination of the minimum antibody concentration required for ligand discovery.

One bead one compound (OBOC) libraries can be screened against serum samples to identify ligands to antibodies in this mixture. In this protocol, hit beads are identified by staining with a fluorescent labeled secondary antibody. When screens are conducted against two different sets of serum, antibodies, and ligands to them, can be discovered that distinguish the two populations. The application of DNA-encoding technology to OBOC libraries has allowed the use of 10 µm beads for library preparation and screening, which pass through a standard flow cytometer, allowing the fluorescent hit beads to be separated from beads displaying non-ligands easily. An important issue in using this approach for the discovery of antibody biomarkers is its analytical sensitivity. In other words, how abundant must an IgG be to allow it to be pulled out of serum in an unbiased screen using a flow cytometer? We report here a model study in which monoclonal antibodies with known ligands of varying affinities are doped into serum. We find that for antibody ligands typical of what one isolates from an unbiased combinatorial library, the target antibody must be present at 10-50 nM. True antigens, which bind with significantly higher affinity, can detect much less abundant serum antibodies.

Cytotoxic, Anti-inflammatory, and Leishmanicidal Activities of Diterpenes Isolated from the Roots of Caesalpinia pulcherrima.

A phytochemical investigation on the chloroform extract of Caesalpinia pulcherrima roots led to the isolation of ten known furanocassane diterpenoids, vouacapen-5α-ol (1), 8,9,11,14-didehydrovouacapen-5α-ol (2), 6ß-cinnamoyl-7ß-hydroxyvouacapen-5α-ol (3), pulcherrin A (4), pulcherrin B (5), pulcherrin J (6), pulcherrimin A (7), pulcherrimin B (8), pulcherrimin C (9), and pulcherrimin E (10). Chemical transformation of 3 and 7 gave compounds 6ß-hydroxyisovouacapenol C (11), 6ß-cinnamoyl-7ß-acetoxyvouacapen-5α-ol (12), and pulcherrimin D (13). Cytotoxicity of compounds 1-13 was evaluated against three cancer cell lines (MCF-7, HeLa, and PC-3). Anti-inflammatory potential of the compounds was evaluated via the oxidative burst assay using a luminol-amplified chemiluminescence technique. Leishmanicidal activity was tested against promastigotes of Leishmania major in vitro. Compounds 3, 4, 8, 9, and 10 were found active against all three cancer cell lines with IC50s ranging from 7.02 ± 0.31 to 36.49 ± 1.39 µM. Compounds 8 and 13 exhibited a potent inhibitory effect on reactive oxygen species generated from human whole blood phagocytes (IC50 = 15.30 ± 1.10 µM and 8.00 ± 0.80 µM, respectively). Compounds 3, 9, and 13 showed significant activity against promastigotes of L. major (IC50 = 65.30 ± 3.20, 58.70 ± 2.80, and 55.90 ± 2.40 µM, respectively).

Crystal structure of (3S*,4S*,4aS*,5R*,6R*,6aS*,7R*,11aS*,11bR*)-5,6-bis(benzo-yloxy)-3,4a-dihy-droxy-4,7,11b-trimethyl-1,2,3,4,4a,5,6,6a,7,11,11a,11b-dodeca-hydro-phenanthro[3,2-b]furan-4-carb-oxy-lic acid methanol monosolvate.

The title compound, C34H36O9·CH3OH, is a diterpenoid isolated from the roots of Caesalpinia pulcherrima (L.) Swartz. The three trans-fused six-membered rings are in chair, chair and half-chair conformations. The mean plane of this fused-ring system makes dihedral angles of 67.95 (15) and 83.72 (14)° with the two phenyl rings of the benzo-yloxy groups. An intra-molecular C-H⋯O hydrogen bond is observed. In the crystal, mol-ecules are linked via O-H⋯O hydrogen bonds, forming an infinite chain along the b-axis direction.

Anticancer activity of five forest crops used in African folklore: antiproliferative and pro-apoptotic effects.

Acalypha wilkesiana, Caesalpinia bonduc, Jatropha multifida, Momordica charantia and Picralima nitida used in African folklore for treating cancer were investigated. All extracts except J. multifida resulted in no significant alteration in cell cycle distribution and apoptosis in MCF-7 and BT-20. The J. multifilda (JMR-Ch) caused cell cycle arrest at G1 checkpoint and apoptosis in MCF-7. Slight changes in the integrin expression of MCF-7 after treatment with 1 and 10 µg/mL of JMR-Ch were observed. Fluorescence-activated confocal microscopy shows changes in cell morphology and ß1 integrin localisation within MCF-7 cells after exposure to 10 and 25 µg/mL of JMR-Ch. JMR-Ch (1 µg/mL) treatment resulted in time-dependent decrease in cell acidification and respiration in MCF-7 cells and a time-dependent decrease in BT-20 cell respiration, while in MCF-10A, there was an enhancement of acidification. These results revealed the probable application of JMR-Ch in cancer therapy.

Medicinal uses, phytochemistry and pharmacology of Picralima nitida (Apocynaceae) in tropical diseases: a review.

Picralima nitida Durand and Hook, (fam. Apocynaceae) is a West African plant with varied applications in African folk medicine. Various parts of the plant have been employed ethnomedicinally as remedy for fever, hypertension, jaundice, dysmenorrheal, gastrointestinal disorders and malaria. In order to reveal its full pharmacological and therapeutic potentials, the present review focuses on the current medicinal uses, phytochemistry, pharmacological and toxicological activities of this species. Literature survey on scientific journals, books as well as electronic sources have shown the isolation of alkaloids, tannins, polyphenols and steroids from different parts of the plant, pharmacological studies revealed that the extract or isolated compounds from this species posses analgesic, anti-inflammatory, hypoglyceamic, hypotensive, antiplasmodial, antimicrobial, antiulcer and antitumorigenic activities. Results from various scientific investigations to date have revealed the potential of the extract from the plant or isolated compounds for use in the treatment and prevention of various kinds of human diseases. However, further studies on the extracts and pure compounds from this species is required to completely assess its phytochemical, pharmacological and toxicological profile as well as the mechanism of action behind these pharmacological activities exhibited by the compounds isolated from this species.