Crossing the blood–brain barrier in primates is a major obstacle for gene delivery to the brain. Adeno-associated viruses (AAVs) promise robust, non-invasive gene delivery from the bloodstream to the brain. However, unlike in rodents, few neurotropic AAVs efficiently cross the blood–brain barrier in non-human primates. Here we report on AAV.CAP-Mac, an engineered variant identified by screening in adult marmosets and newborn macaques, which has improved delivery efficiency in the brains of multiple non-human primate species: marmoset, rhesus macaque and green monkey. CAP-Mac is neuron biased in infant Old World primates, exhibits broad tropism in adult rhesus macaques and is vasculature biased in adult marmosets. We demonstrate applications of a single, intravenous dose of CAP-Mac to deliver functional GCaMP for ex vivo calcium imaging across multiple brain areas, or a cocktail of fluorescent reporters for Brainbow-like labelling throughout the macaque brain, circumventing the need for germline manipulations in Old World primates. As such, CAP-Mac is shown to have potential for non-invasive systemic gene transfer in the brains of non-human primates.

Adeno-associated viruses (AAVs) promise robust gene delivery to the brain through non-invasive, intravenous delivery. However, unlike in rodents, few neurotropic AAVs efficiently cross the blood-brain barrier in non-human primates (NHPs). Here we describe AAV.CAP-Mac, an engineered variant identified by screening in adult marmosets and newborn macaques with improved efficiency in the brain of multiple NHP species: marmoset, rhesus macaque, and green monkey. CAP-Mac is neuron-biased in infant Old World primates, exhibits broad tropism in adult rhesus macaques, and is vasculature-biased in adult marmosets. We demonstrate applications of a single, intravenous dose of CAP-Mac to deliver (1) functional GCaMP for ex vivo calcium imaging across multiple brain areas, and (2) a cocktail of fluorescent reporters for Brainbow-like labeling throughout the macaque brain, circumventing the need for germline manipulations in Old World primates. Given its capabilities for systemic gene transfer in NHPs, CAP-Mac promises to help unlock non-invasive access to the brain.

Adeno-associated viruses (AAVs) can enable robust and safe gene delivery to the mammalian central nervous system (CNS). While the scientific community has developed numerous neurotropic AAV variants for systemic gene-transfer to the rodent brain, there are few AAVs that efficiently access the CNS of higher order primates. We describe here AAV.CAP-Mac, an engineered AAV variant that enables systemic, brain-wide gene delivery in infants of two Old World primate species—the rhesus macaque ( Macaca mulatta ) and the green monkey ( Chlorocebus sabaeus ). We identified CAP-Mac using a multi-species selection strategy, initially screening our library in the adult common marmoset ( Callithrix jacchus ) and narrowing our pool of test-variants for another round of selection in infant macaques. In individual characterization, CAP-Mac robustly transduces human neurons in vitro and Old World primate neurons in vivo , where it targets all lobes of cortex, the cerebellum, and multiple subcortical regions of disease relevance. We use CAP-Mac for Brainbow-like multicolor labeling of macaque neurons throughout the brain, enabling morphological reconstruction of both medium spiny neurons and cortical pyramidal cells. Because of its broad distribution throughout the brain and high neuronal efficiency in infant Old World primates compared to AAV9, CAP-Mac shows promise for researchers and clinicians alike to unlock novel, noninvasive access to the brain for efficient gene transfer.

A screening campaign of a diverse collection of approximately 250,000 small molecule compounds was performed to identify inhibitors of proline-rich tyrosine kinase 2 (Pyk2) with potential osteogenic activity in osteoblast cells. Compounds were prioritized based on selectivity following a counter-screen against focal adhesion kinase (FAK), a closely related kinase. 4-Amino and 5-aryl substituted pyridinone series were identified that showed strong biochemical potency against Pyk2 and up to 3700-fold selectivity over FAK. Modeling analysis suggested that structural differences in the substrate binding cleft could explain the high selectivity of these chemical series against FAK. Representative compounds from each series showed inhibition of Pyk2 autophosphorylation in 293T cells (IC(50) approximately 0.11 microM), complete inhibition of endogenous Pyk2 in A7r5 cells and increased levels of osteogenic markers in MC3T3 osteoblast cells (EC(50)'s approximately 0.01 microM). These results revealed a new class of compounds with osteogenic-inducing activity in osteoblast cells and a starting point for the development of more potent and selective Pyk2 inhibitors.

Calcimimetic compounds, which activate the parathyroid cell Ca2+ receptor (CaR) and inhibit parathyroid hormone (PTH) secretion, are under experimental study as a treatment for hyperparathyroidism. This report describes the salient pharmacodynamic properties, using several test systems, of a new calcimimetic compound, cinacalcet HCl. Cinacalcet HCl increased the concentration of cytoplasmic Ca2+ ([Ca2+](i)) in human embryonic kidney 293 cells expressing the human parathyroid CaR. Cinacalcet HCl (EC50 = 51 nM) in the presence of 0.5 mM extracellular Ca2+ elicited increases in [Ca2+](i) in a dose- and calcium-dependent manner. Similarly, in the presence of 0.5 mM extracellular Ca2+, cinacalcet HCl (IC50 = 28 nM) produced a concentration-dependent decrease in PTH secretion from cultured bovine parathyroid cells. Using rat medullary thyroid carcinoma 6-23 cells expressing the CaR, cinacalcet HCl (EC50 = 34 nM) produced a concentration-dependent increase in calcitonin secretion. In vivo studies in rats demonstrated cinacalcet HCl is orally bioavailable and displays approximately linear pharmacokinetics over the dose range of 1 to 36 mg/kg. Furthermore, this compound suppressed serum PTH and blood-ionized Ca2+ levels and increased serum calcitonin levels in a dose- dependent manner. Cinacalcet was about 30-fold more potent at lowering serum levels of PTH than it was at increasing serum calcitonin levels. The S-enantiomer of cinacalcet (S-AMG 073) was at least 75-fold less active in these assay systems. The present findings provide compelling evidence that cinacalcet HCl is a potent and stereoselective activator of the parathyroid CaR and, as such, might be beneficial in the treatment of hyperparathyroidism.

This study examines the effects of an IL-6-producing murine multiple myeloma cell line on trabecular and cortical mouse bone, and evaluates the efficacy of interleukin-1 receptor antagonist (IL-1ra) in mitigating bone destruction. Six-week-old BALB/c mice were assigned to two groups: normal controls and myeloma animals (5 x 10(7) MPC-11 cells on day 0). Myeloma animals were further assigned to three unique groups: MPC-11 only; MPC-11 treated with hyaluronic acid (HA); and MPC-11 + IL-1ra/HA (100 mg/kg). Disease development was assessed at 14 and 21 days via spleen, liver, and proximal tibia histology; histomorphometry at the femoral middiaphysis; and long bone composition and mechanical testing. Histologic analysis revealed marked myeloma infiltration into organs and bone marrow and gross bone resorption of the proximal tibia. IL-1ra tended to decrease bone resorption at the proximal tibia; however, it had no effect on quantitatively measured bone parameters. Whole femur and tibia, and tibial epiphysis, percent mineralization was decreased (3.0%, 2.9%, and 6.3%, respectively) in all MPC-11 groups. The presence of myeloma did not affect long bone stiffness, strength, or length over the 3 week study. The percent of the femoral endosteal perimeter showing excessive resorption ( approximately 60%) in the MPC-11 groups increased significantly after 21 days. MPC-11 cell presence caused no change in bone formation or morphology. Normal growth mechanisms were not impacted, as the bones lengthened and increased in size and mass despite the presence of myeloma. IL-1 does not appear to be a primary factor in in vivo bone destruction caused by the MPC-11 cell line. These findings reveal the stochastic nature of bone lesions in multiple myeloma and suggest that IL-1 is not a cytokine critical to this disease pathology.

OBJECTIVE:To determine the potential for additive or synergistic effects of combination therapy with the recombinant anticytokine agents interleukin-1 receptor antagonist (IL-1Ra) and PEGylated soluble tumor necrosis factor receptor type I (PEG sTNFRI) in established type H collagen-induced arthritis (CIA) and developing adjuvant-induced arthritis (AIA) in rats.METHODS:Rats with established CIA or developing AIA were treated with various doses of IL-1Ra in a slow-release hyaluronic acid vehicle or with PEG sTNFRI, either alone or in combination with the IL-1Ra. The effects of treatment were monitored by sequential caliper measurements of the ankle joints or hind paw volumes, final paw weights, and histologic evaluation with particular emphasis on bone and cartilage lesions.RESULTS:Combination therapy with IL-1Ra and PEG sTNFRI in rats with CIA resulted in an additive effect on clinical and histologic parameters when moderately to highly efficacious doses of each protein were administered. Greater-than-additive effects were seen when an inactive dose of IL-1Ra was given in combination with moderately to minimally active doses of PEG sTNFRI. Plasma levels associated with the latter effect (for both proteins) were similar to those seen in rheumatoid arthritis (RA) patients in clinical trials with these agents. Combination therapy in the AIA model generally resulted in additive effects, but some parameters showed a greater-than-additive benefit.CONCLUSION:The results provide preclinical support for the hypothesis that IL-1Ra administered in combination with PEG sTNFRI might provide substantially more clinical benefit to RA patients than either agent alone at blood levels that are currently achievable in patients.

Purpose. To determine the effect of hylan fluid (HA), a model slow release vehicle on the pharmacokinetic profile and efficacy of interleukin-1 receptor antagonist (IL-lra) in rats with established type II collagen arthritis.

ADVERTISEMENT RETURN TO ISSUEPREVCommunicationNEXTMorpholino-Linked Ribozymes: A Convergent Synthetic ApproachLaurent Bellon, Christopher Workman, Jon Scherrer, Nassim Usman, and Francine WincottView Author Information Department of Chemistry and Biochemistry Ribozyme Pharmaceuticals, Inc. 2950 Wilderness Place, Boulder, Colorado 80301 Cite this: J. Am. Chem. Soc. 1996, 118, 15, 3771–3772Publication Date (Web):April 17, 1996Publication History Received22 January 1996Published online17 April 1996Published inissue 1 January 1996https://pubs.acs.org/doi/10.1021/ja9602049https://doi.org/10.1021/ja9602049rapid-communicationACS PublicationsCopyright © 1996 American Chemical SocietyRequest reuse permissionsArticle Views224Altmetric-Citations28LEARN ABOUT THESE METRICSArticle Views are the COUNTER-compliant sum of full text article downloads since November 2008 (both PDF and HTML) across all institutions and individuals. These metrics are regularly updated to reflect usage leading up to the last few days.Citations are the number of other articles citing this article, calculated by Crossref and updated daily. Find more information about Crossref citation counts.The Altmetric Attention Score is a quantitative measure of the attention that a research article has received online. Clicking on the donut icon will load a page at altmetric.com with additional details about the score and the social media presence for the given article. Find more information on the Altmetric Attention Score and how the score is calculated. Share Add toView InAdd Full Text with ReferenceAdd Description ExportRISCitationCitation and abstractCitation and referencesMore Options Share onFacebookTwitterWechatLinked InRedditEmail Other access optionsGet e-Alertsclose SUBJECTS:Adducts,Crystal cleavage,High-performance liquid chromatography,Organic reactions,Sodium Get e-Alerts

In general, protein conformational changes are beststudied in the appropriate native environment, whetherit be aqueous solution or a biological membrane. 19Fnuclear magnetic resonance (NMR) provides a valuabletool with which to carry out such studies, even for avarietyofproteinstoolargeorinsoluble for multidimen­sional NMRtechniques. Recombinant DNAtechniquessignificantly extend the usefulness of19F NMR, both byfacilitating resonance assignments, and by enablingseparation ofmultidomain structures into smaller, moreaccessible units. Thepresentstudyillustrates the type ofinformation provided by this combined approach usingtwo receptorsfrom theE. coli chemosensory pathway asexamples: the periplasmic o-galactose and o-glucosereceptorand the transmembrane aspartate transducer.