T-cell activation and B-cell depletion in chimpanzees treated with a bispecific anti-CD19/anti-CD3 single-chain antibody construct

BscCD19xCD3 is a bispecific single-chain antibody construct with exceptional cytotoxic potency in vitro and in vivo. Here, we have investigated the biological activity of bscCD19xCD3 in chimpanzee, the only animal species identified in which bscCD19xCD3 showed bispecific binding, redirected B-cell lysis and cytokine production comparable to human cells. Pharmacokinetic analysis following 2-h intravenous infusion of 0.06, 0.1 or 0.12 μg/kg of bscCD19xCD3 as part of a dose escalation study in a single female chimpanzee revealed a half-life of approximately 2 h and elimination of the bispecific antibody from circulation within approximately 8 h after the end of infusion. This short exposure to bscCD19xCD3 elicited a transient increase in serum levels of IFNγ, IL-6, IL-2, soluble CD25, and transiently upregulated expression of CD69 and MHC class II on CD8-positive cells. Cytokine release and upregulation of T-cell activation markers were not observed with vehicle controls. A multiple-dose study using 5 weekly doses of 0.1 μg/kg in two animals also showed transient cytokine release and an activation of peripheral T cells with a first-dose effect, accompanied by a transient lymphopenia. While oscillations of T-cell counts were relatively even during repeated treatments, the amplitudes of peripheral B cells declined with every infusion, which was not observed in a vehicle control animal. Our data show that bscCD19xCD3 can be safely administered to chimpanzees at dose levels that cause fully reversible T-cell activation and, despite a very short exposure time, cumulative loss of peripheral B lymphocytes. A clinical trial testing prolonged administration of bscCD19xCD3 (MT103) for improving efficacy is currently ongoing.     

Observations of a new source of coral mortality along the Kenyan coast

In early 2002 coral mortality occurred along 600 km of coastline from Tanzania to Kenya. Astreopora, Echinopora , and Montipora species were severely affected, with Montipora being nearly eliminated from Kenyan reefs. Acropora , Platygyra , Goniopora , and massive Porites were also affected; however, Porites and Goniopora rarely died and often recovered, whereas death for most other species occurred within 2 weeks. In Echinopora and Montipora , a dull ashy tissue color and brittle skeletons characterized the early stages of this event with a mucus layer on the tissue surface in intermediate stages. Mucus and embedded debris then disappeared and surfaces were left covered in a white calcareous dust that sometimes capped a black layer. Astreopora tissues became dull and pale, and seldom produced mucus; eventually the skeleton became bare and white. Either a colorless translucent or brownish thin margin of tissue was visible between living tissue and bare skeleton, depending on species. Scanning electron micrographs of affected corals revealed the presence of fungi. Histology and staining showed that the fungi were mostly in the three genera that died from the syndrome and it may be that fungi invaded and killed corals weakened by another unidentified pathogen.     

Neurovirulence of polytropic murine retrovirus is influenced by two separate regions on opposite sides of the envelope protein receptor binding domain

Changes in the envelope proteins of retroviruses can alter the ability of these viruses to infect the central nervous system (CNS) and induce neurological disease. In the present study, nine envelope residues were found to influence neurovirulence of the Friend murine polytropic retrovirus Fr98. When projected on a three-dimensional model, these residues were clustered in two spatially separated groups, one in variable region B of the receptor binding site and the other on the opposite side of the envelope. Further studies indicated a role for these residues in virus replication in the CNS, although the residues did not affect viral entry.     

A new class of bradykinin B1 receptor antagonists with high oral bioavailability and minimal PXR activity

The design and synthesis of a novel class of human bradykinin B1 antagonists featuring difluoroethyl ether and isoxazole carboxamide moieties are disclosed. Compound 7g displayed excellent pharmacokinetic properties, efficient ex vivo receptor occupancy, and low potential for P450 induction via PXR activation.     

Neural correlates of Early Stone Age toolmaking: technology, language and cognition in human evolution

Archaeological and palaeontological evidence from the Early Stone Age (ESA) documents parallel trends of brain expansion and technological elaboration in human evolution over a period of more than 2Myr. However, the relationship between these defining trends remains controversial and poorly understood. Here, we present results from a positron emission tomography study of functional brain activation during experimental ESA (Oldowan and Acheulean) toolmaking by expert subjects. Together with a previous study of Oldowan toolmaking by novices, these results document increased demands for effective visuomotor coordination and hierarchical action organization in more advanced toolmaking. This includes an increased activation of ventral premotor and inferior parietal elements of the parietofrontal praxis circuits in both the hemispheres and of the right hemisphere homologue of Broca's area. The observed patterns of activation and of overlap with language circuits suggest that toolmaking and language share a basis in more general human capacities for complex, goal-directed action. The results are consistent with coevolutionary hypotheses linking the emergence of language, toolmaking, population-level functional lateralization and association cortex expansion in human evolution.     

Display of a peptide mimotope on a crystalline bacterial cell surface layer (S-layer) lattice for diagnosis of Epstein-Barr virus infection

Fusion proteins based on the crystalline bacterial cell surface layer (S-layer) proteins SbpA from Bacillus sphaericus CCM 2177 and SbsB from Geobacillus stearothermophilus PV72/p2 and a peptide mimotope F1 that mimics an immunodominant epitope of Epstein-Barr virus (EBV) were designed and overexpressed in Escherichia coli. Constructs were designed such that the peptide mimotope was presented either at the C-terminus (SbpA/F1) or at the N-terminus (SbsB/F1) of the respective S-layer proteins. The resulting S-layer fusion proteins, SbpA/F1 and SbsB/F1, fully retained the intrinsic self-assembly capability of the S-layer moiety into monomolecular lattices. As determined by immunodot assays and ELISAs using monoclonal antibodies, the F1 mimotope was well-presented on the outer surface of the S-layer lattices and accessible for antibody binding. Further comparison of the two S-layer fusion proteins showed that the S-layer fusion protein SbpA/F1 had a higher antibody binding capacity than SbsB/F1 in aqueous solution and in immune sera, illustrating the importance of epitope orientation on the performance of solid-phase immunoassays. To assess the diagnostic values of S-layer mimotope fusion protein SbpA/F1, we screened a panel of 83 individual EBV IgM-positive, EBV negative, and potential cross-reactive sera for their reactivities. This resulted in 98.2% specificity and 89.3% sensitivity, and furthermore no cross-reactivity with related viral disease states including rheumatoid factor was observed. This study shows the potential of S-layer fusion proteins as a matrix for site-directed immobilization of small ligands in solid-phase immunoassays using EBV diagnostics as a model system.     

Characterization of a complex glycoprotein whose variable metabolic clearance in humans is dependent on terminal N-acetylglucosamine content.

Glycoproteins can be cleared from circulation if they carry oligosaccharide structures that are recognized by specific receptors. High-mannose type and asialo complex oligosaccharides are cleared by the mannose and asialoglycoprotein receptors, respectively. This paper presents the protein and terminal saccharide characterization for nine batches of a glycoprotein developed for pharmaceutical use. Each of these batches was evaluated in human pharmacokinetic (PK) studies, and had similar terminal elimination half-lives, but the initial clearance of this glycoprotein varied between batches. The protein is lenercept, an immunoadhesin comprising the Fc domain of human IgG1 and two tumor necrosis factor (TNF) binding domains derived from the extracellular portion of the TNFR1(p55). Lenercept is manufactured in Chinese hamster ovary (CHO) cells and is extensively N-glycosylated but is devoid of high-mannose glycans. The pharmacokinetic variability between these lots only correlated with terminal N-acetylglucosamine and not with terminal galactose, sialic acid or any polypeptide related parameter. The data emphasize the need for appropriate analytical methods for the characterization of glycoproteins, especially those designed for long half-lives, and show that assessment of the content of all three terminal saccharides is sufficient to ensure consistency of their PK performance properties.     

Origin of beta-cells in regenerating pancreas

The origin of insulin-expressing beta-cells in the adult mammalian pancreas is controversial. During normal tissue turnover and following injury, beta-cells may be replaced by duplication of existing beta-cells.1 However, an alternative source of beta-cells has recently been proposed based on neogenesis from a Ngn3-positive population present in regenerating pancreatic ducts.2 The appearance of beta-cells from Ngn3-positive progenitors is reminiscent of normal pancreas development, and Ngn3-expressing cells isolated from regenerating pancreas can generate the full repertoire of endocrine phenotypes. The isolation and characterisation of the equivalent human progenitors may represent a significant step forward in the hunt for a cure for diabetes.