Objective
This study was undertaken to describe the safety, tolerability, pharmacokinetics, and immunogenicity of elezanumab (ABT‐555), a fully human monoclonal antibody (mAb) directed against ...repulsive guidance molecule A (RGMa), in healthy and multiple sclerosis (MS) study participants.
Methods
The single‐center, first‐in‐human, single ascending dose (SAD) study evaluated elezanumab (50–1,600mg intravenous IV and 150mg subcutaneous) in 47 healthy men and women. The multicenter multiple ascending dose (MAD; NCT02601885) study evaluated elezanumab (150mg, 600mg, and 1,800mg) in 20 adult men and women with MS, receiving either maintenance or no immunomodulatory treatment.
Results
No pattern of study drug‐related adverse events was identified for either the SAD or MAD elezanumab regimens. Across both studies, the Tmax occurred within 4 hours of elezanumab IV infusion, and the harmonic mean of t1/2 ranged between 18.6 and 67.7 days. Following multiple dosing, elezanumab Cmax, area under the curve, and Ctrough increased dose‐proportionally and resulted in dose‐dependent increases in elezanumab cerebrospinal fluid (CSF) concentrations. Elezanumab CSF penetration was 0.1% to 0.4% across both studies, with CSF levels of free RGMa decreased by >40%. Changes in CSF interleukin‐10 (IL‐10) and free RGMa demonstrated dose/exposure‐dependence.
Interpretation
The elezanumab pharmacokinetic profile supports monthly, or bimonthly, administration of up to 1,800mg with the option of a loading dose of 3,600mg. Elezanumab partitioning into CSF is within the range expected for mAbs. Reduced CSF levels of free RGMa demonstrate central nervous system target binding of elezanumab with an apparent maximal effect at 1,800mg IV. Exposure‐associated increases in CSF IL‐10, an anti‐inflammatory cytokine with neuroprotective/neurorestorative properties, support potential pathway modulation in MS participants. ANN NEUROL 2023;93:285–296
Clinicopathological studies suggest that Alzheimer's disease (AD) pathology begins ∼10-15 years before the resulting cognitive impairment draws medical attention. Biomarkers that can detect AD ...pathology in its early stages and predict dementia onset would, therefore, be invaluable for patient care and efficient clinical trial design. We utilized a targeted proteomics approach to discover novel cerebrospinal fluid (CSF) biomarkers that can augment the diagnostic and prognostic accuracy of current leading CSF biomarkers (Aβ42, tau, p-tau181).
Using a multiplexed Luminex platform, 190 analytes were measured in 333 CSF samples from cognitively normal (Clinical Dementia Rating CDR 0), very mildly demented (CDR 0.5), and mildly demented (CDR 1) individuals. Mean levels of 37 analytes (12 after Bonferroni correction) were found to differ between CDR 0 and CDR>0 groups. Receiver-operating characteristic curve analyses revealed that small combinations of a subset of these markers (cystatin C, VEGF, TRAIL-R3, PAI-1, PP, NT-proBNP, MMP-10, MIF, GRO-α, fibrinogen, FAS, eotaxin-3) enhanced the ability of the best-performing established CSF biomarker, the tau/Aβ42 ratio, to discriminate CDR>0 from CDR 0 individuals. Multiple machine learning algorithms likewise showed that the novel biomarker panels improved the diagnostic performance of the current leading biomarkers. Importantly, most of the markers that best discriminated CDR 0 from CDR>0 individuals in the more targeted ROC analyses were also identified as top predictors in the machine learning models, reconfirming their potential as biomarkers for early-stage AD. Cox proportional hazards models demonstrated that an optimal panel of markers for predicting risk of developing cognitive impairment (CDR 0 to CDR>0 conversion) consisted of calbindin, Aβ42, and age.
Using a targeted proteomic screen, we identified novel candidate biomarkers that complement the best current CSF biomarkers for distinguishing very mildly/mildly demented from cognitively normal individuals. Additionally, we identified a novel biomarker (calbindin) with significant prognostic potential.
A major feature of septic shock is the development of a vascular crisis characterized by nonresponsiveness to sympathetic vasoconstrictor agents and the subsequent irreversible fall in blood ...pressure. In addition, sepsis, like other inflammatory conditions, results in a large increase in the production of free radicals, including superoxide anions (O$_{2}^{\overline{\cdot}}$) within the body. Here we show that O$_{2}^{\overline{\cdot}}$reacts with catecholamines deactivating them in vitro. Moreover, this deactivation would appear to account for the hyporeactivity to exogenous catecholamines observed in sepsis, because administration of a superoxide dismutase (SOD) mimetic to a rat model of septic shock to remove excess O$_{2}^{\overline{\cdot}}$restored the vasopressor responses to norepinephrine. This treatment with the SOD mimetic also reversed the hypotension in these animals; suggesting that deactivation of endogenous norepinephrine by O$_{2}^{\overline{\cdot}}$contributes significantly to this aspect of the vascular crisis. Indeed, the plasma concentrations of both norepinephrine and epinephrine in septic rats treated with the SOD mimetic were significantly higher than in untreated rats. Interestingly, the plasma concentrations for norepinephrine and epinephrine were inversely related to the plasma concentrations of adrenochromes, the product of the autoxidation of catecholamines initiated by O$_{2}^{\overline{\cdot}}$. We propose, therefore, that the use of a SOD mimetic represents a new paradigm for the treatment of septic shock. By removing O$_{2}^{\overline{\cdot}}$, exogenous and endogenous catecholamines are protected from autoxidation. As a result, both hyporeactivity and hypotension are reversed, generation of potentially toxic adrenochromes is reduced, and survival rate is improved.
Measurement of nitrotyrosine levels in biological fluids can serve as a biomarker for oxidative/nitrative damage arising from formation of reactive nitrogen species, including peroxynitrite. ...Peroxynitrite is formed by the reaction of the superoxide radical
(
O
2
.
-
)
with the nitric oxide radical (
.NO) that is generated by nitric oxide synthase (NOS). This article describes an immunoaffinity liquid chromatography–tandem mass spectrometry (LC–MS/MS) method to measure 3-nitrotyrosine at very low (picomolar) levels. Incorporation of a pronase digestion step prior to the immunoaffinity LC–MS/MS allowed for measuring not only free amino acid but also protein 3-nitrotyrosine in biological fluids. The use of an in-line antibody column allowed for increased specificity as compared with previously reported assays. The assay is linear over a range of 5 to 500 pg/ml (0.022–2.20 nM,
r
2
=
0.9987), with the lower detection limit being 5 pg/ml. In addition to its increased sensitivity and specificity, this assay showed great nitrotyrosine recovery from biological fluids when either nitrotyrosine or nitrotyrosine-containing peptides were added exogenously. The utility of this assay for nitrotyrosine as a clinically translatable biomarker was demonstrated by quantifying both free and total nitrotyrosine levels in various biological fluids, including urine, plasma, serum, cerebrospinal fluid (CSF), and synovial fluid (SF) from both preclinical species and human subjects. Thus, whether in an animal model of human disease or in a clinical setting, the quantification of nitrotyrosine levels should provide support for NOS-driven pathology and its blockade following therapeutic intervention.
Many human diseases are associated with the overproduction of oxygen free radicals that inflict cell damage. A manganese(II) complex with a bis(cyclohexylpyridine)-substituted macrocyclic ligand ...(M40403) was designed to be a functional mimic of the superoxide dismutase (SOD) enzymes that normally remove these radicals. M40403 had high catalytic SOD activity and was chemically and biologically stable in vivo. Injection of M40403 into rat models of inflammation and ischemia-reperfusion injury protected the animals against tissue damage. Such mimics may result in better clinical therapies for diseases mediated by superoxide radicals.
Inflamed tissue is often characterized by the production of NO and superoxide. These radicals react at diffusion-limited rates to form the powerful oxidant peroxynitrite (PN). When protonated, PN ...decomposes into either nitrate or reactive intermediates capable of mediating tissue damage by oxidation of protein, lipid, and nucleic acid. We recently have identified porphyrin derivatives capable of catalyzing an increase in nitrate formation with a concomitant decrease in the HO· -like and NO2
·-like reactivity of PN. Here, we present evidence for the efficacy of these PN decomposition catalysts both in vitro and in vivo. Cells in culture were protected from exogenously added PN by the catalyst 5,10,15,20-tetrakis(2,4,6-trimethyl-3,5-disulfonatophenyl)-porphyrinato iron (III), whereas free iron and the structurally related compound without iron 5,10,15,20-tetrakis(2,4,6-trimethyl-3,5-disulfonatophenyl)porphyrin did not protect. Cytoprotection correlated well with a reduction in the nitrotyrosine content of released cytosolic proteins, a biochemical marker for PN formation. Carrageenan-induced paw edema is a model of acute inflammation in which PN may play a major role. When tested in this system, both 5,10,15,20-tetrakis(2,4,6-trimethyl-3,5-disulfonatophenyl)porphyrinato iron (III) and 5,10,15,20-tetrakis(N-methyl-4′-pyridyl) porphyrinato iron (III) caused a dose-dependent reduction in swelling and lactate dehydrogenase release as well as a detectable shift to nitrate formation in paw tissue. In addition, the catalysts did not elevate mean arterial pressure, suggesting a lack of interaction with NO. Taken together, our data provide compelling evidence supporting the therapeutic value of manipulating PN pharmacologically. Thus, PN decomposition catalysts may represent a unique class of antiinflammatory agents.
Summary Objective Statins possess anti-inflammatory properties. This study was undertaken to characterize the mechanism of action of statin drugs on collagenase expression in primary human ...osteoarthritic cartilage tissue. Method Human articular chondrocytes and cartilage explants from osteoarthritic donors were exposed to simvastatin in the presence or absence of interleukin-1 beta (IL-1β). Collagenase expression was determined by quantifying levels of matrix metalloproteinase 13 (MMP-13) and MMP-1 mRNA and MMP-13 protein. The mechanism of statin action was tested by addition of farnesyl pyrophosphate (FPP) and geranylgeranyl pyrophosphate (GGPP) or by using inhibitors of farnesyl transferase (FT) and geranylgeranyl transferase (GGT-1). Results Treatment of osteoarthritic chondrocytes with simvastatin decreased mRNA levels of MMP-13 and MMP-1 whether under basal conditions or during stimulation with IL-1β. MMP-13 protein secreted into the culture media was also decreased. Genes involved in cartilage synthesis (type II collagen and aggrecan) were not down-regulated by simvastatin. Exogenous addition of GGPP completely reversed the statin-mediated decrease in MMP-13 mRNA and protein levels whereas FPP partially reversed the statin-mediated effect. An inhibitor of GGT-1 mimicked the simvastatin-mediated reduction in MMP-13 expression by chondrocytes. Finally, consistent with impacts on MMP-13 and MMP-1 expression, simvastatin as well as the GGT-1 inhibitor both blocked type II collagen degradation in primary human articular cartilage explants. Conclusion These results suggest that statins modulate chondrocyte metabolism by reducing prenylation of key signaling molecules that control the expression of collagen-degrading enzymes. Our results strongly support the hypothesis that protein prenyltransferases including geranylgeranyl transferase regulate chondrocyte collagenase expression in osteoarthritis.
Standard measurements used to assess murine models of rheumatoid arthritis, notably paw thickness and clinical score, do not align well with certain aspects of disease severity as assessed by ...histopathology. We tested the hypothesis that non-invasive optical tomographic imaging of molecular biomarkers of inflammation and bone turnover would provide a superior quantitative readout and would discriminate between a disease-modifying anti-rheumatic drug (DMARD) and a non-DMARD treatment.
Using two protease-activated near-infrared fluorescence imaging agents to detect inflammation-associated cathepsin and matrix metalloprotease activity, and a third agent to detect bone turnover, we quantified fluorescence in paws of mice with collagen antibody-induced arthritis. Fluorescence molecular tomographic (FMT) imaging results, which provided deep tissue detection and quantitative readouts in absolute picomoles of agent fluorescence per paw, were compared with paw swelling, clinical scores, a panel of plasma biomarkers, and histopathology to discriminate between steroid (prednisolone), DMARD (p38 mitogen-activated protein kinase (MAPK) inhibitor) and non-DMARD (celecoxib, cyclooxygenase-2 (COX-2) inhibitor) treatments.
Paw thickness, clinical score, and plasma biomarkers failed to discriminate well between a p38 MAPK inhibitor and a COX-2 inhibitor. In contrast, FMT quantification using near-infrared agents to detect protease activity or bone resorption yielded a clear discrimination between the different classes of therapeutics. FMT results agreed well with inflammation scores, and both imaging and histopathology provided clearer discrimination between treatments as compared with paw swelling, clinical score, and serum biomarker readouts.
Non-invasive optical tomographic imaging offers a unique approach to monitoring disease pathogenesis and correlates with histopathology assessment of joint inflammation and bone resorption. The specific use of optical tomography allowed accurate three-dimensional imaging, quantitation in picomoles rather than intensity or relative fluorescence, and, for the first time, showed that non-invasive imaging assessment can predict the pathologist's histology inflammation scoring and discriminate DMARD from non-DMARD activity.
Nitric Oxide Activates Cyclooxygenase Enzymes Salvemini, Daniela; Misko, Thomas P.; Masferrer, Jaime L. ...
Proceedings of the National Academy of Sciences - PNAS,
08/1993, Letnik:
90, Številka:
15
Journal Article
Recenzirano
Odprti dostop
We have evaluated the role of nitric oxide (NO) on the activity of the constitutive and induced forms of cyclooxygenase (COX; COX-1 and COX-2, respectively). Induction of NO synthase (NOS) and COX ...(COX-2) in the mouse macrophage cell line RAW264.7 by Escherichia coli lipopolysaccharide (1 μ g/ml, 18 h) caused an increase in the release of nitrite (NO-
2) and prostaglandin E2(PGE2), products of NOS and COX, respectively. Production of both NO-
2and PGE2was blocked by the NOS inhibitors NG-monomethyl-L-arginine or aminoguanidine. The effects of NG-monomethyl-L-arginine or aminoguanidine were reversed by coincubation with L-Arg, the precursor for NO synthesis, but not by D-Arg. RAW264.7 cells stimulated for 18 h with lipopolysaccharide in L-Arg-free medium (to reduce NO generation by the endogenous NOS pathway) failed to release NO-
2and accumulated at least 4-fold less PGE2when compared to cells in the presence of L-Arg. PGE2production elicited by a 15-min arachidonic acid treatment of lipopolysaccharide-induced RAW264.7 cells in L-Arg-deficient medium was decreased 3-fold when compared to the release obtained with cells induced in medium containing L-Arg. To examine the NO activation of the induced form of COX in the absence of an endogenous L-Arg, human fetal fibroblasts were first stimulated for 18 h with interleukin 1β. These cells released PGE2but not NO-
2, consistent with the induction of COX but not NOS in the fibroblast. Exogenous NO either as a gaseous solution or released by a NO donor, sodium nitroprusside or glyceryl trinitrate, increased COX activity in the interleukin 1β-stimulated fibroblasts by 5-fold; these effects were abolished by coincubation with hemoglobin (10 μ M), which binds and inactivates NO, but not by methylene blue, an inhibitor of the soluble guanylate cyclase. Furthermore, sodium nitroprusside (0.25-1 mM) increased arachidonic acid-stimulated PGE2production by murine recombinant COX-1 and COX-2. These results demonstrate that NO enhances COX activity through a mechanism independent of cGMP and suggest that, in conditions in which both the NOS and COX systems are present, there is an NO-mediated increase in the production of proinflammatory prostaglandins that may result in an exacerbated inflammatory response. The data suggest that NO directly interacts with COX to cause an increase in the enzymatic activity.
Overproduction of the free radical nitric oxide (NO) has been implicated in the pathogenesis of a variety of inflammatory and immunologically mediated diseases as well as complications of diabetes. ...In the present study we have demonstrated that aminoguanidine selectively inhibits the cytokine-inducible isoform of NO synthase which appears to be responsible for the excess production of NO linked to these disease states. By using organ, cell, and enzyme-based measurements we have shown that aminoguanidine is equipotent to NG-monomethyl-L-arginine (L-NMA) as an inhibitor of the cytokine-induced isoform of NO synthase but is 10 to 100-fold less potent as an inhibitor of the constitutive isoform. Thus, aminoguanidine may be useful as a selective inhibitor of the inducible NO synthase in the treatment of disease states characterized by the pathological overproduction of NO.
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