Critical role of the NOTCH ligand JAG2 in self-renewal of myeloma cells.

The purpose of this study was to identify the pathways associated with the ability of CD138(+) human myeloma cells to form colonies in a serum-free semi-solid human collagen-based assay. Only 26% (7 of 27) of human myeloma cell lines were able to spontaneously form colonies. This spontaneous clonogenic growth correlated with the expression of the NOTCH ligand JAG2 (p<0.001). Blocking JAG-NOTCH interactions with NOTCH-Fc chimeric molecules impaired self-colony formation, indicating a role for JAG-NOTCH pathway in colony formation. In two cell lines, silencing of JAG2 blocked both colony formation and in vivo tumor formation in immunocompromised mice. RT-PCR and flow cytometry analysis revealed that JAG2 is often expressed by CD138(+) primary cells. Our results indicate that spontaneous clonogenic growth of myeloma cells requires the expression of JAG2.

Human susceptibility and resistance to Norwalk virus infection.

Infectious diseases have influenced population genetics and the evolution of the structure of the human genome in part by selecting for host susceptibility alleles that modify pathogenesis. Norovirus infection is associated with approximately 90% of epidemic non-bacterial acute gastroenteritis worldwide. Here, we show that resistance to Norwalk virus infection is multifactorial. Using a human challenge model, we showed that 29% of our study population was homozygous recessive for the alpha(1,2)fucosyltransferase gene (FUT2) in the ABH histo-blood group family and did not express the H type-1 oligosaccharide ligand required for Norwalk virus binding. The FUT2 susceptibility allele was fully penetrant against Norwalk virus infection as none of these individuals developed an infection after challenge, regardless of dose. Of the susceptible population that encoded a functional FUT2 gene, a portion was resistant to infection, suggesting that a memory immune response or some other unidentified factor also affords protection from Norwalk virus infection.

Bile-salt-stimulated lipase and mucins from milk of 'secretor' mothers inhibit the binding of Norwalk virus capsids to their carbohydrate ligands.

Breast-feeding-associated protection against calicivirus diarrhoea is associated with the presence of high levels of 2-linked oligosaccharides in mother's milk, and human calicivirus strains including the NV (Norwalk virus) use gut 2-linked fucosylated glycans as receptors, suggesting the presence of decoy receptors in milk. Our aim was to analyse the ability of human milk to inhibit the attachment of rNV VLPs (recombinant NV-like particles) to their carbohydrate ligands and to characterize potential inhibitors found in milk. Milk from women with the secretor phenotype was strongly inhibitory, unlike milk from women that are non-secretors, which is devoid of 2-linked fucosylated structures. At least two fractions in human milk acted as inhibitors for the NV capsid attachment. The first fraction corresponded to BSSL (bile-salt-stimulated lipase) and the second to associated mucins MUC1 and MUC4. These proteins present tandem repeat O-glycosylated sequences that should act as decoy receptors for the NV, depending on the combined mother/child secretor status.

Novel tetrahydropyrido[1,2-a]isoindolone derivatives (valmerins): potent cyclin-dependent kinase/glycogen synthase kinase 3 inhibitors with antiproliferative activities and antitumor effects in human tumor xenografts.

The development of CDK and GSK3 inhibitors has been regarded as a potential therapeutic approach, and a substantial number of diverse structures have been reported to inhibit CDKs and GSK-3ß in recent years. Only a few molecules have gone through or are currently undergoing clinical trials as CDK and GSK inhibitors. In this paper, we prepared valmerins, a new family containing the tetrahydropyrido[1,2-a]isoindone core. The fused heterocycle was prepared with a straightforward synthesis that was functionalized by a (het)arylurea. Twelve valmerins inhibited the CDK5 and GSK3 with an IC(50) < 100 nM. A semiquantitative kinase scoring was realized, and a cellular screening was done. At the end of our study, we investigated the in vivo potency of one valmerin. Mice exhibited good tolerance to our lead, which proved its efficacy and clearly blocked tumor growth. Valmerins appear also as good candidates for further development as anticancer agents.

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.

Influence of the combined ABO, FUT2, and FUT3 polymorphism on susceptibility to Norwalk virus attachment.

The binding of Norwalk virus (NV) recombinant capsids was tested in a panel of saliva samples collected from 96 donors with different ABO, secretor, and Lewis phenotypes. As previously reported, binding occurred specifically to saliva from secretors, regardless of their Lewis phenotype status. Blood group B saliva was poorly recognized, whereas binding to blood group O saliva was higher and binding to blood group A saliva was highest. Transfection of either blood group A or B enzyme into H epitope-expressing cells showed that masking of H epitopes by the A and B antigens blocked the attachment of NV capsids. The high level of binding to blood group A secretor saliva could be explained by an optimal H type 1 ligand density, which was lower than that in blood group O saliva and much higher than that in blood group B saliva. Indeed, despite a higher ligand density, saliva from homozygotes with 2 functional FUT2 alleles was less strongly recognized than saliva from heterozygotes with 1 functional and 1 inactivated FUT2 allele. Partial fucosidase treatment of duodenal tissue sections and binding to a synthetic probe with varying densities of H type 1 trisaccharide indicated that optimal attachment occurred at medium ligand density.

Expression of histo-blood group A antigen increases resistance to apoptosis and facilitates escape from immune control of rat colon carcinoma cells.

A and B histo-blood group antigens are present on carcinoma cells at the early stages of cancerogenesis and tend to disappear at later stages, but it is not yet clear whether they take part to the process of tumor progression. To gain some insight into this issue, we used a rat colon carcinoma experimental model. To obtain expression of the A antigen, REG cells were cotransfected with the rat A enzyme cDNA and a rat alpha1,2fucosyltransferase cDNA, either FTA or FTB, whereas PRO cells that spontaneously have alpha1,2fucosyltransferase activity were only transfected with the A enzyme cDNA. All A antigen-expressing transfected cells derived from either REG FTA, REG FTB, or PRO parental cells were more resistant to apoptosis induced by either serum deprivation or heat shock than were their respective controls. When injected to syngeneic immunocompetent rats, A enzyme-transfected PRO cells formed tumors that grew faster than those formed by mock-transfected PRO cells. However, in immunodeficient SCID mice, no difference in growth could be observed between the two types of tumors, indicating that the faster tumor growth of the A antigen-positive cells in immunocompetent animals was due to their higher ability to escape immune control and that this was associated with their higher degree of resistance to apoptosis. These results might explain the slightly augmented incidence of carcinomas observed in A and B blood group individuals compared to O individuals.

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.

Noroviruses bind to human ABO, Lewis, and secretor histo-blood group antigens: identification of 4 distinct strain-specific patterns.

We characterized the binding of 8 Noroviruses (NORs) to histo-blood group antigens (HBGAs) in human saliva using recombinant NOR (rNOR) capsid proteins. Among the 8 rNORs tested, 6 formed viruslike particles (VLPs) when the capsid proteins were expressed in insect cells, all of which revealed variable binding activities with saliva; the remaining 2 rNORs did not form VLPs, and the proteins did not bind, or bound weakly, to saliva. Four distinct binding patterns were associated with different histo-blood types, defined by Lewis, secretor, and ABO types. Three patterns (VA387, NV, and MOH) recognized secretors, and 1 pattern (VA207) recognized Lewis-positive nonsecretors. The 3 secretor-recognizing patterns were defined as A/B (MOH), A/O (NV), and A/B/O (VA387) binders. Oligosaccharides containing the Lewis and ABH antigenic epitopes were involved in binding. Our findings suggest that different strains of NORs may recognize different human HBGAs on intestinal epithelial cells as receptors for infection.