N-aryl-piperidine-4-carboxamides as a novel class of potent inhibitors of MALT1 proteolytic activity

Starting from a weak screening hit, potent and selective inhibitors of the MALT1 protease function were elaborated. Advanced compounds displayed high potency in biochemical and cellular assays. Compounds showed activity in a mechanistic Jurkat T cell activation assay as well as in the B-cell lymphoma line OCI-Ly3, which suggests potential use of MALT1 inhibitors in the treatment of autoimmune diseases as well as B-cell lymphomas with a dysregulated NF-κB pathway. Initially, rat pharmacokinetic properties of this compound series were dominated by very high clearance which could be linked to amide cleavage. Using a rat hepatocyte assay a good in vitro-in vivo correlation could be established which led to the identification of compounds with improved PK properties.     

Chromosome Translocation Inflates Bacillus Forespores and Impacts Cellular Morphology

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Assessing genetic-based therapies for AIDS using the simian immunodeficiency virus

Abstract: A plasmid encoding the full-length infectious molecular proviral clone of SIV_mac239 was generated. Virus derived from cells transfected with this clone replicated to high levels and was cytopathic for some transformed human CD4⁺ cell lines and primary rhesus macaque peripheral blood mononuclear cells. Since replication of SIV requires the functional expression of the viral encoded rev protein, transient co-transfection studies were initiated with the infectious proviral clone and a well-characterized trans-dominant negative HIV-1 rev mutant.     

Virulence of La Crosse virus is under polygenic control.

To identify which RNA segments of the California serogroup bunyaviruses determine virulence, we prepared reassortant viruses by coinfecting BHK-21 cells with two wild-type parents, La Crosse/original and Tahyna/181-57 viruses, which differed about 30,000-fold in virulence. The progeny clones were screened by polyacrylamide gel electrophoresis to ascertain the phenotype of the M and S RNA segments, and RNA-RNA hybridization was used to determine the genotype of selected clones. Two or three clones of each of the six possible reassortant genotypes were characterized quantitatively for neuroinvasiveness by determining the PFU/50% lethal dose (LD50) ratio after subcutaneous injection into suckling mice. The reassortants fell into two groups. (i) Six of seven reassortants with a La Crosse M RNA segment were as virulent as the parent La Crosse virus (about 1 PFU/LD50); the one exception was strikingly different (about 1,000 PFU/LD50) and probably represents a spontaneous mutant. (ii) The seven reassortants with a Tahyna M RNA segment were about 10-fold more virulent than the parent Tahyna virus (median 1,600 PFU/LD50 for reassortants and 16,000 PFU/LD50 for Tahyna virus). A comparative pathogenesis study in suckling mice of one reassortant virus and the parent Tahyna virus confirmed the greater neuroinvasiveness of the reassortant virus. From these data it was concluded that the M RNA segment was the major determinant of virulence, but that the other two gene segments could modulate the virulence of a nonneuroinvasive California serogroup virus.     

Coordinate production by a T cell hybridoma of gamma interferon and three other lymphokine activities: multiple activities of a single lymphokine?

The T cell hybridoma FS7-20, produced by the fusion of normal B10.BR T cells to the AKR thymoma BW5147, was found when stimulated with concanavalin A (Con A) to produce the lymphokines: interleukin 2 (IL 2), interferon-gamma (IFN gamma), macrophage-activating factor (MAF), Ia induction factor IaIF), and the B cell helper factor interleukin X (IL X). The clones and subclones of FS7-20 varied dramatically in their ability to produce these lymphokines, presumably because of karyotypic variations. The ability to produce IL 2 segregated independently from the ability to produce the four other lymphokine activities; however, production of the latter activities showed a strong correlation. This coordinate production of IFN gamma, MAF, IaIF, and IL X was also observed with a cloned normal cytotoxic T cell line, cr15. These results suggest either that IFN gamma, MAF, IaIF, and IL X are all manifestations of a single molecular species or that, although these activities are different structurally, their production is controlled by a common genetic mechanism. In support of the first possibility, the IFN gamma, MAF, IaIF, and IL X activity produced by FS7-20 were all found to be equally sensitive to inactivation at pH 2. These results illustrate the usefulness of using T cell hybridomas for the study of lymphokines.     

An IL 2-secreting T cell hybridoma that responds to a self class I histocompatibility antigen in the H-2D region

A self-reactive T cell hybridoma that secretes IL-2 in response to H-2d haplotype cells resulted from a fusion of BALB/cBy lymph node cells with the AKR thymoma BW5147. The lymph node cells used had been enriched for cells reactive to (TG)-A--L, but neither this antigen nor fetal calf serum were required for stimulation of the hybridoma designated 3DT52.5. The gene product responsible for stimulation mapped to the H-2D region. Allogeneic cells of the b, f, k, q, and s haplotypes failed to stimulate. Not all H-2d haplotype cells were effective stimulators of 3DT52.5. Peritoneal cells and splenic B cells were much more stimulatory than splenic T cells. Most tumor cell lines of H-2d derivation and of B cell or macrophage/monocyte lineage were stimulatory, whereas H-2d T cell lines were not. The capacity to stimulate 3DT52.5 did not correlate with the ability to stimulate I region-restricted hybridomas, or with the ability to be induced to stimulate such hybridomas. Stimulatory cell lines did not apparently produce a soluble factor required for stimulation, and negative cell lines were not inhibitory. The monoclonal antibody 27-11-13, which reacts with H-2D of the b, d, and q haplotypes, inhibited stimulation of 3DT52.5 but did not inhibit stimulation of the sibling hybridoma 3DT18.11, which responds to (TG)-A--L plus I-Ad. Conversely, the monoclonal anti-I-Ad antibody MK-D6 inhibited stimulation of 3DT18.11 but not 3DT52.5. Although it is clear that 3DT52.5 recognizes a class I antigen coded for in the H-2D region, the precise molecular nature of the antigen is unknown. The structure of the antigen receptor on this hybridoma may prove to be of interest when it can be compared with receptors found on T cell hybridomas restricted by class II histocompatibility antigens.     

Diffusible Fraction of Serum Cholesterol and Atherogenesis

CHOLESTEROL is found in the blood as a structural component of lipoproteins concerned with the transport of other lipids¹. The high resolution nuclear magnetic resonance spectra of high density serum lipoproteins are similar to that observed when lipids are dissolved in organic solvents, or dispersed in water by bile salts or detergents, or in sonicated form. The lipid component in lipoproteins is therefore probably in an extremely fluid condition². If human serum is mixed with paraffin oil, some of the cholesterol diffuses into the oil without affecting the ultraviolet absorption spectrum of serum proteins. This procedure avoids any protein denaturing action used for cholesterol extraction^3–5. It therefore seems that serum cholesterol has two fractions, one strongly bound by lipoprotein structures and the other loosely bound and diffusible in an oil phase. In this article I designate the loosely bound fraction “diffusible”.     

Neuroattenuation of an avirulent bunyavirus variant maps to the L RNA segment.

The derivation and characterization of a neuroattenuated reassortant clone (RFC 25/B.5) of California serogroup bunyavirus was described previously (M. J. Endres, A. Valsamakis, F. Gonzalez-Scarano, and N. Nathanson, J. Virol. 64:1927-1933, 1990). To map the RNA segment responsible for this attenuation, a panel of reassortants was constructed between the attenuated clone B.5 (genotype TLL) and a virulent clone (B1-1a) of reciprocal genotype (LTT). Parent viruses and clones representing all of the six possible reassortants were examined for neurovirulence by intracerebral injection in adult mice. Reassortants bearing the large RNA segment from the virulent parent were almost as virulent as the virulent parent virus, while reassortants bearing the large RNA segment from the avirulent parent virus exhibited low or intermediate virulence. These results indicate that the large RNA segment is the major determinant of neuroattenuation of clone B.5. In addition to its neuroattenuation, clone B.5 was temperature sensitive and exhibited an altered plaque morphology. These phenotypes also segregated with the large RNA segment. The importance of the large RNA segment (which encodes the viral polymerase) in neurovirulence contrasts with prior studies which indicate that the ability to cause lethal encephalitis after peripheral injection of suckling mice (neuroinvasiveness) is primarily determined by the middle-sized RNA segment, which encodes the viral glycoproteins.