Antimalarial activity of axidjiferosides, new ß-galactosylceramides from the African sponge Axinyssa djiferi.

The marine sponge, Axinyssa djiferi, collected on mangrove tree roots in Senegal, was investigated for glycolipids. A mixture containing new glycosphingolipids, named axidjiferoside-A, -B and -C, accounted for 0.07% of sponge biomass (dry weight) and for 2.16% of total lipids. It showed a significant antimalarial activity, with a 50% inhibitory concentration (IC50) of 0.53 ± 0.2 µM against a chloroquine-resistant strain of Plasmodium falciparum. They were identified as homologous ß-galactopyranosylceramides composed of 2-amino-(6E)-octadec-6-en-1,3,4-triol, and the major one, axidjiferoside-A (around 60%), contained 2-hydroxytetracosanoic acid. Cytotoxicity was studied in vitro on human cancer cell lines (multiple myeloma, colorectal adenocarcinoma, glioblastoma and two lung cancer NSCLC-N6 and A549). Results of this investigation showed that axidjiferosides are of interest, because they proved a good antiplasmodial activity, with only a low cytotoxicity against various human cell lines and no significant antitrypanosomal and antileishmanial activity. Thus, it seems that galactosylceramides with a ß anomeric configuration may be suitable in searching for new antimalarial drugs.

Synthesis of bile acid derivatives and in vitro cytotoxic activity with pro-apoptotic process on multiple myeloma (KMS-11), glioblastoma multiforme (GBM), and colonic carcinoma (HCT-116) human cell lines.

The novelty of this work derives from the use of nitrogenous heterocycles as building block in the synthesis of conjugate bile acid derivatives. New piperazinyl bile acid derivatives were synthesized and tested in vitro against various human cancer cells (GBM, KMS-11, HCT-116). The best pro-apoptotic activity was obtained with N-[4N-cinnamylpiperazin-1-yl)-3alpha,7beta-dihydroxy-5beta-cholan-24-amide (7b) and N-[4N-cinnamyllpiperazin-1-yl)- 3alpha,7alpha-dihydroxy-5beta-cholan-24-amide (7c) on these human cancer cell lines (IC(50): 8.5-31.4microM). This activity was associated with nuclear and DNA fragmentation, demonstrating that 7b induces cell death by an apoptotic process as 7c. This study shows the possibility of hydrid heterocycle-steroids as new anticancer agents with improved bioactivity and easy to synthesize.

Inhibition of mTORC1 activity by REDD1 induction in myeloma cells resistant to bortezomib cytotoxicity.

The combination of bortezomib and dexamethasone is becoming the reference induction treatment for multiple myeloma patients younger than 65 years. Despite its advantage over vincristin adryamicin dexamethasone induction treatment, bortezomib does not benefit all patients. We hypothesize that heterogeneity of the response experienced by myeloma patients is, at least in part, due to genomic variations in the malignant plasma cells. To test this hypothesis we used gene expression profiling to identify early responsive genes induced by bortezomib in resistant myeloma cells. Our study revealed: (i) a dramatic induction of REDD1, a negative regulator of mammalian target of rapamycin kinase complex 1 (mTORC1) activity, in these cells; (ii) a transient cell size decrease associated with REDD1 overexpression; and (iii) partial restoration of bortezomib sensitivity in REDD1 knockdown bortezomib-resistant myeloma cells. Together, these results identify a possible novel mechanism of bortezomib resistance in myeloma patients mediated by REDD1 overexpression involving inhibition of mTORC1 activity and suggest that the use of mammalian target of rapamycin inhibitors in myeloma patients could be deleterious.

Cytotoxicity on human cancer cells of ophidiacerebrosides isolated from the African starfish Narcissia canariensis.

The starfish Narcissia canariensis harvested from the coasts off Dakar, Senegal, was investigated for glycolipids (GL). This report deals with the isolation, characterization and biological activity of a fraction F13-3 separated from the GL mixture and selected according to its ability to inhibit KB cell proliferation after 72 hours of treatment. Firstly, a GL mixture F13 was obtained that accounted for 1.36% of starfish biomass (dry weight) and 0.36% of total lipids. The fraction F13-3 obtained from F13 contained three homologous GL identified as peracetylated derivatives on the basis of chemical and spectroscopic evidence. These contained a ß-glucopyranoside as sugar head, a 9-methyl-branched 4,8,10-triunsaturated long-chain aminoalcohol as sphingoid base and amide-linked 2-hydroxy fatty acid chains. The majority (63%) had an amide-linked 2-hydroxydocosanoic acid chain and was identified as the ophidiacerebroside-C, firstly isolated from the starfish Ophidiaster ophidiamus. The minor components of F13-3 differed by one more or one less methylene group, and corresponded to ophidiacerebroside-B and -D. We found that F13-3 displayed an interesting cytotoxic activity over 24 hours on various adherent human cancerous cell lines (multiple myeloma, colorectal adenocarcinoma and glioblastoma multiforme) with an IC(50) of around 20 µM.

Focus on the controversial activation of human iNKT cells by 4-deoxy analogue of KRN7000.

4-Deoxy-alpha-GalCer analogues are considered weaker agonists than KRN7000 for the stimulation of human iNKT cells, but this remains strongly debated. In this work, we described a strategy toward 4-deoxy-alpha-GalCers with, as a key step, a metathesis reaction allowing sphingosine modifications from a single ethylenic alpha-galactoside precursor. The 4-deoxy-KRN7000 derivative 2, described here, induced potent cytokinic responses, comparable to those of KRN7000, both from human iNKT cells in vitro and from their murine counterpart in vivo.

New lithocholic and chenodeoxycholic piperazinylcarboxamides with antiproliferative and pro-apoptotic effects on human cancer cell lines.

Six new synthetic bile acid derivatives were synthesized and tested in vitro against various human cancer cells (glioblastoma multiforme (GBM), multiple myeloma (KMS-11), and colonic carcinoma (HCT-116) cell lines. The best activity was obtained with compound IIIb on multiple myeloma cells (LD(50): 8.5+/-0.5 microM). This activity was associated with Mcl-1 and PARP-1 cleavage, inhibition of NFkappaB signaling, and DNA fragmentation, demonstrating an apoptotic cell death signaling pathway.

Inhibition of the interaction between the SARS-CoV spike protein and its cellular receptor by anti-histo-blood group antibodies.

Severe acute respiratory syndrome coronavirus (SARS-CoV) is a highly pathogenic emergent virus which replicates in cells that can express ABH histo-blood group antigens. The heavily glycosylated SARS-CoV spike (S) protein binds to angiotensin-converting enzyme 2 which serves as a cellular receptor. Epidemiological analysis of a hospital outbreak in Hong Kong revealed that blood group O was associated with a low risk of infection. In this study, we used a cellular model of adhesion to investigate whether natural antibodies of the ABO system could block the S protein and angiotensin-converting enzyme 2 interaction. To this aim, a C-terminally EGFP-tagged S protein was expressed in chinese hamster ovary cells cotransfected with an alpha1,2-fucosyltransferase and an A-transferase in order to coexpress the S glycoprotein ectodomain and the A antigen at the cell surface. We observed that the S protein/angiotensin-converting enzyme 2-dependent adhesion of these cells to an angiotensin-converting enzyme 2 expressing cell line was specifically inhibited by either a monoclonal or human natural anti-A antibodies, indicating that these antibodies may block the interaction between the virus and its receptor, thereby providing protection. In order to more fully appreciate the potential effect of the ABO polymorphism on the epidemiology of SARS, we built a mathematical model of the virus transmission dynamics that takes into account the protective effect of ABO natural antibodies. The model indicated that the ABO polymorphism could contribute to substantially reduce the virus transmission, affecting both the number of infected individuals and the kinetics of the epidemic.

Meriolins (3-(pyrimidin-4-yl)-7-azaindoles): synthesis, kinase inhibitory activity, cellular effects, and structure of a CDK2/cyclin A/meriolin complex.

We report the synthesis and biological characterization of 3-(pyrimidin-4-yl)-7-azaindoles (meriolins), a chemical hybrid between the natural products meridianins and variolins, derived from marine organisms. Meriolins display potent inhibitory activities toward cyclin-dependent kinases (CDKs) and, to a lesser extent, other kinases (GSK-3, DYRK1A). The crystal structures of 1e (meriolin 5) and variolin B (Bettayeb, K.; Tirado, O. M.; Marionneau-Lambert, S.; Ferandin, Y.; Lozach, O.; Morris, J.; Mateo-Lozano, S.; Drückes, P.; Schächtele, C.; Kubbutat, M.; Liger, F.; Marquet, B.; Joseph, B.; Echalier, A.; Endicott, J.; Notario, V.; Meijer, L. Cancer Res. 2007, 67, 8325-8334) in complex with CDK2/cyclin A reveal that the two inhibitors are orientated in very different ways inside the ATP-binding pocket of the kinase. A structure-activity relationship provides further insight into the molecular mechanism of action of this family of kinase inhibitors. Meriolins are also potent antiproliferative and proapoptotic agents in cells cultured either as monolayers or in spheroids. Proapoptotic efficacy of meriolins correlates best with their CDK2 and CDK9 inhibitory activity. Meriolins thus constitute a promising class of pharmacological agents to be further evaluated against the numerous human diseases that imply abnormal regulation of CDKs including cancers, neurodegenerative disorders, and polycystic kidney disease.

Expression of sialyl-Tn epitopes on beta1 integrin alters epithelial cell phenotype, proliferation and haptotaxis.

Sialyl-Tn (STn) is a tumor-associated carbohydrate antigen overexpressed in various carcinomas. To obtain its expression, murine carcinoma cells were transfected with the cDNA encoding ST6GalNAc I, a glycosyltransferase that acts exclusively on O-glycans. Overexpression of this enzyme led to the expected expression of cell surface STn epitopes. Surprisingly, the transfectants (STn+ cells) presented dramatic morphological changes and altered behavior. These STn+ cells lost the epithelial appearance of parental cells, became larger, more elongated and presented disorganized actin stress fibers. Additionally, their proliferation was impaired and their ability to migrate on fibronectin and hyaluronic acid was severely reduced. By contrast their adhesion on fibronectin remained unchanged. The major glycoprotein carrying the STn epitope was shown to be the integrin beta1 subunit. Anti-STn antibodies could restore migration of STn+ cells on fibronectin. A constitutively active permeant form of RhoA (TAT-RhoA(Val-14)) also restored motility on fibronectin of STn+ cells as well as a parental STn-cellular phenotype. These observations indicate that overexpression of ST6GalNAc I leads to a major change of the O-glycosylation of the integrin beta1 chain which in turn impairs the integrin-mediated signalling and leads to major alterations in morphology and cell behavior.

Butyrate restores motile function and actin cytoskeletal network integrity in apc mutated mouse colon epithelial cells.

Loss of function of the Apc gene product is an early and frequent event in colorectal carcinogenesis. Altered migration of intestinal epithelial cells has been described in vivo in the Min mouse Apc+/Min model. Using cell lines established from this model we show in vitro that Apc+/Min cells are less motile than Apc+/+ cells and exhibit a disordered actin cytoskeletal network. This would increase the probabilities of the initiated cell to acquire additional genetic alterations leading to neoplasia. Butyrate, a product of indigestible carbohydrate fermentation by the colonic flora, is able to restore both motility and actin cytoskeletal organization. This feature may contribute to explain the protective effect exerted by butyrogenic diets on colon carcinogenesis in animal models.

Norwalk virus binds to histo-blood group antigens present on gastroduodenal epithelial cells of secretor individuals.

BACKGROUND & AIMS: Norwalk Virus (NV) is a member of the Caliciviridae family, which causes acute epidemic gastroenteritis in humans of all ages and its cellular receptors have not yet been characterized. Another calicivirus, Rabbit Hemorrhagic Disease Virus, attaches to H type 2 histo-blood group oligosaccharide present on rabbit epithelial cells. Our aim was to test if, by analogy, recombinant NV-like particles (rNV VLPs) use carbohydrates present on human gastroduodenal epithelial cells as ligands. METHODS: Attachment of rNV VLPs was tested on tissue sections of the gastroduodenal junction and on saliva from individuals of known ABO, Lewis, and secretor phenotypes. It was also tested on human Caco-2 cells and on animal cell lines transfected with glycosyltransferases complementary DNA (cDNA). Competition experiments were performed with synthetic oligosaccharides and anticarbohydrate antibodies. Internalization was monitored by confocal microscopy. RESULTS: Attachment of rNV VLPs to surface epithelial cells of the gastroduodenal junction as well as to saliva was detected, yet only from secretor donors. It was abolished by alpha1,2fucosidase treatment, and by competition with the H types 1 and 3 trisaccharides or with anti-H type 1 and anti-H types (3/4) antibodies. Transfection of CHO and TS/A cells with an alpha1,2fucosyltransferase cDNA allowed attachment of VLPs. These transfectants as well as differentiated Caco-2 cells expressing H type 1 structures internalized the bound particles. CONCLUSIONS: rNV VLPs use H type 1 and/or H types (3/4) as ligands on gastroduodenal epithelial cells of secretor individuals.