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1.
J Biol Chem ; 295(33): 11379-11387, 2020 08 14.
Article in English | MEDLINE | ID: mdl-32571879

ABSTRACT

Systemic amyloidosis caused by extracellular deposition of insoluble fibrils derived from the pathological aggregation of circulating proteins, such as transthyretin, is a severe and usually fatal condition. Elucidation of the molecular pathogenic mechanism of the disease and discovery of effective therapies still represents a challenging medical issue. The in vitro preparation of amyloid fibrils that exhibit structural and biochemical properties closely similar to those of natural fibrils is central to improving our understanding of the biophysical basis of amyloid formation in vivo and may offer an important tool for drug discovery. Here, we compared the morphology and thermodynamic stability of natural transthyretin fibrils with those of fibrils generated in vitro either using the common acidification procedure or primed by limited selective cleavage by plasmin. The free energies for fibril formation were -12.36, -8.10, and -10.61 kcal mol-1, respectively. The fibrils generated via plasmin cleavage were more stable than those prepared at low pH and were thermodynamically and morphologically similar to natural fibrils extracted from human amyloidotic tissue. Determination of thermodynamic stability is an important tool that is complementary to other methods of structural comparison between ex vivo fibrils and fibrils generated in vitro Our finding that fibrils created via an in vitro amyloidogenic pathway are structurally similar to ex vivo human amyloid fibrils does not necessarily establish that the fibrillogenic pathway is the same for both, but it narrows the current knowledge gap between in vitro models and in vivo pathophysiology.


Subject(s)
Amyloid Neuropathies, Familial/pathology , Amyloid/chemistry , Prealbumin/chemistry , Amyloid/genetics , Amyloid/ultrastructure , Amyloid Neuropathies, Familial/genetics , Humans , Mutation , Prealbumin/genetics , Protein Aggregates , Protein Aggregation, Pathological/genetics , Protein Aggregation, Pathological/pathology , Protein Stability , Thermodynamics
2.
J Biol Chem ; 293(37): 14192-14199, 2018 09 14.
Article in English | MEDLINE | ID: mdl-30018138

ABSTRACT

Systemic amyloidosis is a usually fatal disease caused by extracellular accumulation of abnormal protein fibers, amyloid fibrils, derived by misfolding and aggregation of soluble globular plasma protein precursors. Both WT and genetic variants of the normal plasma protein transthyretin (TTR) form amyloid, but neither the misfolding leading to fibrillogenesis nor the anatomical localization of TTR amyloid deposition are understood. We have previously shown that, under physiological conditions, trypsin cleaves human TTR in a mechano-enzymatic mechanism that generates abundant amyloid fibrils in vitro In sharp contrast, the widely used in vitro model of denaturation and aggregation of TTR by prolonged exposure to pH 4.0 yields almost no clearly defined amyloid fibrils. However, the exclusive duodenal location of trypsin means that this enzyme cannot contribute to systemic extracellular TTR amyloid deposition in vivo Here, we therefore conducted a bioinformatics search for systemically active tryptic proteases with appropriate tissue distribution, which unexpectedly identified plasmin as the leading candidate. We confirmed that plasmin, just as trypsin, selectively cleaves human TTR between residues 48 and 49 under physiological conditions in vitro Truncated and full-length protomers are then released from the native homotetramer and rapidly aggregate into abundant fibrils indistinguishable from ex vivo TTR amyloid. Our findings suggest that physiological fibrinolysis is likely to play a critical role in TTR amyloid formation in vivo Identification of this surprising intersection between two hitherto unrelated pathways opens new avenues for elucidating the mechanisms of TTR amyloidosis, for seeking susceptibility risk factors, and for therapeutic innovation.


Subject(s)
Amyloidosis/metabolism , Plasminogen/metabolism , Prealbumin/metabolism , Amyloid/metabolism , Databases, Protein , Fibrinolysin/metabolism , Humans , Hydrogen-Ion Concentration , In Vitro Techniques , Protein Denaturation , Protein Folding , Proteolysis , Trypsin/metabolism
3.
N Engl J Med ; 373(12): 1106-14, 2015 Sep 17.
Article in English | MEDLINE | ID: mdl-26176329

ABSTRACT

BACKGROUND: The amyloid fibril deposits that cause systemic amyloidosis always contain the nonfibrillar normal plasma protein, serum amyloid P component (SAP). The drug (R)-1-[6-[(R)-2-carboxy-pyrrolidin-1-yl]-6-oxo-hexanoyl]pyrrolidine-2-carboxylic acid (CPHPC) efficiently depletes SAP from the plasma but leaves some SAP in amyloid deposits that can be specifically targeted by therapeutic IgG anti-SAP antibodies. In murine amyloid A type amyloidosis, the binding of these antibodies to the residual SAP in amyloid deposits activates complement and triggers the rapid clearance of amyloid by macrophage-derived multinucleated giant cells. METHODS: We conducted an open-label, single-dose-escalation, phase 1 trial involving 15 patients with systemic amyloidosis. After first using CPHPC to deplete circulating SAP, we infused a fully humanized monoclonal IgG1 anti-SAP antibody. Patients with clinical evidence of cardiac involvement were not included for safety reasons. Organ function, inflammatory markers, and amyloid load were monitored. RESULTS: There were no serious adverse events. Infusion reactions occurred in some of the initial recipients of larger doses of antibody; reactions were reduced by slowing the infusion rate for later patients. At 6 weeks, patients who had received a sufficient dose of antibody in relation to their amyloid load had decreased liver stiffness, as measured with the use of transient elastography. These patients also had improvements in liver function in association with a substantial reduction in hepatic amyloid load, as shown by means of SAP scintigraphy and measurement of extracellular volume by magnetic resonance imaging. A reduction in kidney amyloid load and shrinkage of an amyloid-laden lymph node were also observed. CONCLUSIONS: Treatment with CPHPC followed by an anti-SAP antibody safely triggered clearance of amyloid deposits from the liver and some other tissues. (Funded by GlaxoSmithKline; ClinicalTrials.gov number, NCT01777243.).


Subject(s)
Amyloidosis/drug therapy , Antibodies, Monoclonal/administration & dosage , Carboxylic Acids/administration & dosage , Pyrrolidines/administration & dosage , Serum Amyloid P-Component/antagonists & inhibitors , Adult , Aged , Amyloidosis/diagnostic imaging , Antibodies, Monoclonal/adverse effects , Antibodies, Monoclonal/pharmacokinetics , Carboxylic Acids/adverse effects , Carboxylic Acids/pharmacokinetics , Dose-Response Relationship, Drug , Humans , Immunoglobulin G , Immunoglobulin Light-chain Amyloidosis , Infusions, Intravenous , Liver/diagnostic imaging , Liver/drug effects , Liver/pathology , Middle Aged , Pyrrolidines/adverse effects , Pyrrolidines/pharmacokinetics , Radionuclide Imaging , Serum Amyloid P-Component/analysis , Serum Amyloid P-Component/immunology
4.
Proc Natl Acad Sci U S A ; 111(4): 1539-44, 2014 Jan 28.
Article in English | MEDLINE | ID: mdl-24474780

ABSTRACT

The Ser52Pro variant of transthyretin (TTR) produces aggressive, highly penetrant, autosomal-dominant systemic amyloidosis in persons heterozygous for the causative mutation. Together with a minor quantity of full-length wild-type and variant TTR, the main component of the ex vivo fibrils was the residue 49-127 fragment of the TTR variant, the portion of the TTR sequence that previously has been reported to be the principal constituent of type A, cardiac amyloid fibrils formed from wild-type TTR and other TTR variants [Bergstrom J, et al. (2005) J Pathol 206(2):224-232]. This specific truncation of Ser52Pro TTR was generated readily in vitro by limited proteolysis. In physiological conditions and under agitation the residue 49-127 proteolytic fragment rapidly and completely self-aggregates into typical amyloid fibrils. The remarkable susceptibility to such cleavage is likely caused by localized destabilization of the ß-turn linking strands C and D caused by loss of the wild-type hydrogen-bonding network between the side chains of residues Ser52, Glu54, Ser50, and a water molecule, as revealed by the high-resolution crystallographic structure of Ser52Pro TTR. We thus provide a structural basis for the recently hypothesized, crucial pathogenic role of proteolytic cleavage in TTR amyloid fibrillogenesis. Binding of the natural ligands thyroxine or retinol-binding protein (RBP) by Ser52Pro variant TTR stabilizes the native tetrameric assembly, but neither protected the variant from proteolysis. However, binding of RBP, but not thyroxine, inhibited subsequent fibrillogenesis.


Subject(s)
Amyloid/metabolism , Prealbumin/metabolism , Proline/metabolism , Serine/metabolism , Amino Acid Sequence , Amyloidosis/genetics , Amyloidosis/pathology , Crystallography, X-Ray , Humans , Hydrogen Bonding , Molecular Conformation , Molecular Sequence Data , Phenotype , Prealbumin/chemistry , Prealbumin/genetics , Proteolysis
5.
Eur Heart J ; 42(23): 2280-2283, 2021 06 14.
Article in English | MEDLINE | ID: mdl-33778886
7.
Circ Res ; 114(4): 672-6, 2014 Feb 14.
Article in English | MEDLINE | ID: mdl-24337102

ABSTRACT

RATIONALE: Baseline circulating concentrations of C-reactive protein (CRP) are significantly associated with cardiovascular disease risk in general populations. This modest association has been inappropriately conflated with causality, and it has been claimed that CRP is proatherogenic. Most of the known causative factors for atherosclerosis stimulate increased CRP production, but comprehensive genetic epidemiology studies provide no support for a pathogenic role of CRP. The reported proinflammatory effects of human CRP preparations on healthy cells in vitro and in healthy animals in vivo have all been produced by poorly characterized CRP preparations, demonstrably caused by impurities, or elicited by CRP made in recombinant Escherichia coli not by humans. None of the in vitro or animal findings have been reproduced with pure natural human CRP. Nevertheless, the strong proinflammatory effects of infusing recombinant bacterial CRP into humans have still been inappropriately ascribed to CRP. OBJECTIVE: To investigate the effects of infusion into healthy adult human volunteers of pure natural human CRP. METHODS AND RESULTS: Comprehensively characterized, pharmaceutical-grade, endotoxin-free, purified CRP, prepared to GMP standard from pooled normal human donor plasma was infused as an intravenous bolus in 7 healthy adult human volunteers at ≤2 mg/kg to provide circulating CRP concentrations ≤44 mg/L. No recipient showed any significant clinical, hematologic, coagulation, or biochemical changes, or any increase in proinflammatory cytokines or acute phase proteins. CONCLUSIONS: The human CRP molecule itself is not proinflammatory in healthy human adults.


Subject(s)
C-Reactive Protein/administration & dosage , Cardiovascular Physiological Phenomena/drug effects , Inflammation/immunology , Adolescent , Adult , Atherosclerosis/blood , Atherosclerosis/immunology , C-Reactive Protein/immunology , C-Reactive Protein/isolation & purification , Dose-Response Relationship, Drug , Healthy Volunteers , Humans , Inflammation/blood , Infusions, Intravenous , Male , Serum Amyloid A Protein/metabolism , Young Adult
8.
Nature ; 468(7320): 93-7, 2010 Nov 04.
Article in English | MEDLINE | ID: mdl-20962779

ABSTRACT

Accumulation of amyloid fibrils in the viscera and connective tissues causes systemic amyloidosis, which is responsible for about one in a thousand deaths in developed countries. Localized amyloid can also have serious consequences; for example, cerebral amyloid angiopathy is an important cause of haemorrhagic stroke. The clinical presentations of amyloidosis are extremely diverse and the diagnosis is rarely made before significant organ damage is present. There is therefore a major unmet need for therapy that safely promotes the clearance of established amyloid deposits. Over 20 different amyloid fibril proteins are responsible for different forms of clinically significant amyloidosis and treatments that substantially reduce the abundance of the respective amyloid fibril precursor proteins can arrest amyloid accumulation. Unfortunately, control of fibril-protein production is not possible in some forms of amyloidosis and in others it is often slow and hazardous. There is no therapy that directly targets amyloid deposits for enhanced clearance. However, all amyloid deposits contain the normal, non-fibrillar plasma glycoprotein, serum amyloid P component (SAP). Here we show that administration of anti-human-SAP antibodies to mice with amyloid deposits containing human SAP triggers a potent, complement-dependent, macrophage-derived giant cell reaction that swiftly removes massive visceral amyloid deposits without adverse effects. Anti-SAP-antibody treatment is clinically feasible because circulating human SAP can be depleted in patients by the bis-d-proline compound CPHPC, thereby enabling injected anti-SAP antibodies to reach residual SAP in the amyloid deposits. The unprecedented capacity of this novel combined therapy to eliminate amyloid deposits should be applicable to all forms of systemic and local amyloidosis.


Subject(s)
Amyloid/drug effects , Amyloidosis/prevention & control , Antibodies/immunology , Antibodies/pharmacology , Serum Amyloid P-Component/antagonists & inhibitors , Serum Amyloid P-Component/immunology , Amyloidosis/therapy , Animals , Antibodies/therapeutic use , Disease Models, Animal , Humans , Mice , Mice, Inbred C57BL , Mice, Transgenic , Serum Amyloid P-Component/genetics
9.
Proc Natl Acad Sci U S A ; 110(40): 16115-20, 2013 Oct 01.
Article in English | MEDLINE | ID: mdl-23959890

ABSTRACT

Systemic amyloid A (AA) amyloidosis is a serious complication of chronic inflammation. Serum AA protein (SAA), an acute phase plasma protein, is deposited extracellularly as insoluble amyloid fibrils that damage tissue structure and function. Clinical AA amyloidosis is typically preceded by many years of active inflammation before presenting, most commonly with renal involvement. Using dose-dependent, doxycycline-inducible transgenic expression of SAA in mice, we show that AA amyloid deposition can occur independently of inflammation and that the time before amyloid deposition is determined by the circulating SAA concentration. High level SAA expression induced amyloidosis in all mice after a short, slightly variable delay. SAA was rapidly incorporated into amyloid, acutely reducing circulating SAA concentrations by up to 90%. Prolonged modest SAA overexpression occasionally produced amyloidosis after long delays and primed most mice for explosive amyloidosis when SAA production subsequently increased. Endogenous priming and bulk amyloid deposition are thus separable events, each sensitive to plasma SAA concentration. Amyloid deposits slowly regressed with restoration of normal SAA production after doxycycline withdrawal. Reinduction of SAA overproduction revealed that, following amyloid regression, all mice were primed, especially for rapid glomerular amyloid deposition leading to renal failure, closely resembling the rapid onset of renal failure in clinical AA amyloidosis following acute exacerbation of inflammation. Clinical AA amyloidosis rarely involves the heart, but amyloidotic SAA transgenic mice consistently had minor cardiac amyloid deposits, enabling us to extend to the heart the demonstrable efficacy of our unique antibody therapy for elimination of visceral amyloid.


Subject(s)
Amyloid/metabolism , Amyloidosis/physiopathology , Inflammation/complications , Serum Amyloid A Protein/metabolism , Amyloidosis/etiology , Animals , Congo Red , DNA Primers/genetics , Doxycycline/pharmacology , Female , Gene Expression Regulation/drug effects , Mice , Mice, Transgenic , Real-Time Polymerase Chain Reaction
10.
N Engl J Med ; 366(24): 2276-83, 2012 Jun 14.
Article in English | MEDLINE | ID: mdl-22693999

ABSTRACT

We describe a kindred with slowly progressive gastrointestinal symptoms and autonomic neuropathy caused by autosomal dominant, hereditary systemic amyloidosis. The amyloid consists of Asp76Asn variant ß(2)-microglobulin. Unlike patients with dialysis-related amyloidosis caused by sustained high plasma concentrations of wild-type ß(2)-microglobulin, the affected members of this kindred had normal renal function and normal circulating ß(2)-microglobulin values. The Asp76Asn ß(2)-microglobulin variant was thermodynamically unstable and remarkably fibrillogenic in vitro under physiological conditions. Previous studies of ß(2)-microglobulin aggregation have not shown such amyloidogenicity for single-residue substitutions. Comprehensive biophysical characterization of the ß(2)-microglobulin variant, including its 1.40-Å, three-dimensional structure, should allow further elucidation of fibrillogenesis and protein misfolding.


Subject(s)
Amyloidosis, Familial/genetics , beta 2-Microglobulin/genetics , Amyloidosis, Familial/complications , Diarrhea/etiology , Female , Genes, Dominant , Humans , Male , Middle Aged , Pedigree , Protein Structure, Quaternary , Proteome/genetics , Sjogren's Syndrome/complications , Sjogren's Syndrome/genetics , beta 2-Microglobulin/chemistry
11.
J Biol Chem ; 288(43): 30917-30, 2013 Oct 25.
Article in English | MEDLINE | ID: mdl-24014031

ABSTRACT

Systemic amyloidosis is a fatal disease caused by misfolding of native globular proteins, which then aggregate extracellularly as insoluble fibrils, damaging the structure and function of affected organs. The formation of amyloid fibrils in vivo is poorly understood. We recently identified the first naturally occurring structural variant, D76N, of human ß2-microglobulin (ß2m), the ubiquitous light chain of class I major histocompatibility antigens, as the amyloid fibril protein in a family with a new phenotype of late onset fatal hereditary systemic amyloidosis. Here we show that, uniquely, D76N ß2m readily forms amyloid fibrils in vitro under physiological extracellular conditions. The globular native fold transition to the fibrillar state is primed by exposure to a hydrophobic-hydrophilic interface under physiological intensity shear flow. Wild type ß2m is recruited by the variant into amyloid fibrils in vitro but is absent from amyloid deposited in vivo. This may be because, as we show here, such recruitment is inhibited by chaperone activity. Our results suggest general mechanistic principles of in vivo amyloid fibrillogenesis by globular proteins, a previously obscure process. Elucidation of this crucial causative event in clinical amyloidosis should also help to explain the hitherto mysterious timing and location of amyloid deposition.


Subject(s)
Amyloid/chemistry , Mutation, Missense , Protein Folding , alpha-Crystallins/chemistry , beta 2-Microglobulin/chemistry , Amino Acid Substitution , Amyloid/genetics , Amyloid/metabolism , Amyloidosis, Familial/genetics , Amyloidosis, Familial/metabolism , Humans , Protein Structure, Quaternary , alpha-Crystallins/genetics , alpha-Crystallins/metabolism , beta 2-Microglobulin/genetics , beta 2-Microglobulin/metabolism
12.
Acta Crystallogr D Biol Crystallogr ; 70(Pt 8): 2232-40, 2014 Aug.
Article in English | MEDLINE | ID: mdl-25084341

ABSTRACT

Under physiological conditions, the pentameric human plasma protein serum amyloid P component (SAP) binds hexanoyl bis(D-proline) (R-1-{6-[R-2-carboxy-pyrrolidin-1-yl]-6-oxo-hexanoyl}pyrrolidine-2-carboxylic acid; CPHPC) through its D-proline head groups in a calcium-dependent interaction. Cooperative effects in binding lead to a substantial enhancement of affinity. Five molecules of the bivalent ligand cross-link and stabilize pairs of SAP molecules, forming a decameric complex that is rapidly cleared from the circulation by the liver. Here, it is reported that X-ray analysis of the SAP complex with CPHPC and cadmium ions provides higher resolution detail of the interaction than is observed with calcium ions. Conformational isomers of CPHPC observed in solution by HPLC and by X-ray analysis are compared with the protein-bound form. These are discussed in relation to the development of CPHPC to provide SAP depletion for the treatment of amyloidosis and other indications.


Subject(s)
Proline/chemistry , Serum Amyloid P-Component/chemistry , Calcium/chemistry , Calorimetry , Chromatography, High Pressure Liquid , Crystallography, X-Ray , Proline/analogs & derivatives , Protein Conformation , Thermodynamics
13.
Immunology ; 142(3): 414-20, 2014 Jul.
Article in English | MEDLINE | ID: mdl-24673624

ABSTRACT

No deficiency of human C-reactive protein (CRP), or even structural polymorphism of the protein, has yet been reported so its physiological role is not known. Here we show for the first time that CRP-deficient mice are remarkably susceptible to Streptococcus pneumoniae infection and are protected by reconstitution with isolated pure human CRP, or by anti-pneumococcal antibodies. Autologous mouse CRP is evidently essential for innate resistance to pneumococcal infection before antibodies are produced. Our findings are consistent with the significant association between clinical pneumococcal infection and non-coding human CRP gene polymorphisms which affect CRP expression. Deficiency or loss of function variation in CRP may therefore be lethal at the first early-life encounter with this ubiquitous virulent pathogen, explaining the invariant presence and structure of CRP in human adults.


Subject(s)
C-Reactive Protein/immunology , Immunity, Innate , Pneumococcal Infections/immunology , Streptococcus pneumoniae/immunology , Animals , C-Reactive Protein/deficiency , C-Reactive Protein/genetics , Humans , Mice , Mice, Knockout , Phenotype
14.
Open Biol ; 14(7): 230419, 2024 Jul.
Article in English | MEDLINE | ID: mdl-39013416

ABSTRACT

The mechanisms responsible for neuronal death causing cognitive loss in Alzheimer's disease (AD) and many other dementias are not known. Serum amyloid P component (SAP) is a constitutive plasma protein, which is cytotoxic for cerebral neurones and also promotes formation and persistence of cerebral Aß amyloid and neurofibrillary tangles. Circulating SAP, which is produced exclusively by the liver, is normally almost completely excluded from the brain. Conditions increasing brain exposure to SAP increase dementia risk, consistent with a causative role in neurodegeneration. Furthermore, neocortex content of SAP is strongly and independently associated with dementia at death. Here, seeking genomic evidence for a causal link of SAP with neurodegeneration, we meta-analysed three genome-wide association studies of 44 288 participants, then conducted cis-Mendelian randomization assessment of associations with neurodegenerative diseases. Higher genetically instrumented plasma SAP concentrations were associated with AD (odds ratio 1.07, 95% confidence interval (CI) 1.02; 1.11, p = 1.8 × 10-3), Lewy body dementia (odds ratio 1.37, 95%CI 1.19; 1.59, p = 1.5 × 10-5) and plasma tau concentration (0.06 log2(ng l-1) 95%CI 0.03; 0.08, p = 4.55 × 10-6). These genetic findings are consistent with neuropathogenicity of SAP. Depletion of SAP from the blood and the brain, by the safe, well tolerated, experimental drug miridesap may thus be neuroprotective.


Subject(s)
Genome-Wide Association Study , Neurodegenerative Diseases , Serum Amyloid P-Component , Humans , Neurodegenerative Diseases/genetics , Neurodegenerative Diseases/etiology , Neurodegenerative Diseases/metabolism , Serum Amyloid P-Component/metabolism , Serum Amyloid P-Component/genetics , Alzheimer Disease/genetics , Alzheimer Disease/metabolism , Alzheimer Disease/etiology , Polymorphism, Single Nucleotide , Genetic Predisposition to Disease , Mendelian Randomization Analysis , Biomarkers , tau Proteins/metabolism , tau Proteins/genetics , Lewy Body Disease/genetics , Lewy Body Disease/metabolism , Male , Female
15.
Proc Natl Acad Sci U S A ; 107(47): 20483-8, 2010 Nov 23.
Article in English | MEDLINE | ID: mdl-21059958

ABSTRACT

Transthyretin (TTR) amyloidosis is a fatal disease for which new therapeutic approaches are urgently needed. We have designed two palindromic ligands, 2,2'-(4,4'-(heptane-1,7-diylbis(oxy))bis(3,5-dichloro-4,1-phenylene)) bis(azanediyl)dibenzoic acid (mds84) and 2,2'-(4,4'-(undecane-1,11-diylbis(oxy))bis(3,5-dichloro-4,1-phenylene)) bis(azanediyl)dibenzoic acid (4ajm15), that are rapidly bound by native wild-type TTR in whole serum and even more avidly by amyloidogenic TTR variants. One to one stoichiometry, demonstrable in solution and by MS, was confirmed by X-ray crystallographic analysis showing simultaneous occupation of both T4 binding sites in each tetrameric TTR molecule by the pair of ligand head groups. Ligand binding by native TTR was irreversible under physiological conditions, and it stabilized the tetrameric assembly and inhibited amyloidogenic aggregation more potently than other known ligands. These superstabilizers are orally bioavailable and exhibit low inhibitory activity against cyclooxygenase (COX). They offer a promising platform for development of drugs to treat and prevent TTR amyloidosis.


Subject(s)
Amyloid/biosynthesis , Amyloidosis/metabolism , Fenamates/metabolism , Ligands , Prealbumin/metabolism , Amyloid/metabolism , Amyloidosis/drug therapy , Animals , Calorimetry, Differential Scanning , Chromatography, Gel , Crystallography, X-Ray , Fenamates/chemical synthesis , Fenamates/chemistry , Fenamates/pharmacokinetics , Fluorometry , Mass Spectrometry , Mice , Models, Molecular , Molecular Structure , Ultracentrifugation
16.
Open Biol ; 13(12): 230253, 2023 Dec.
Article in English | MEDLINE | ID: mdl-38052249

ABSTRACT

The mechanisms underlying neurodegenerative sequelae of traumatic brain injury (TBI) are poorly understood. The normal plasma protein, serum amyloid P component (SAP), which is normally rigorously excluded from the brain, is directly neurocytotoxic for cerebral neurones and also binds to Aß amyloid fibrils and neurofibrillary tangles, promoting formation and persistence of Aß fibrils. Increased brain exposure to SAP is common to many risk factors for dementia, including TBI, and dementia at death in the elderly is significantly associated with neocortical SAP content. Here, in 18 of 30 severe TBI cases, we report immunohistochemical staining for SAP in contused brain tissue with blood-brain barrier disruption. The SAP was localized to neurofilaments in a subset of neurones and their processes, particularly damaged axons and cell bodies, and was present regardless of the time after injury. No SAP was detected on astrocytes, microglia, cerebral capillaries or serotoninergic neurones and was absent from undamaged brain. C-reactive protein, the control plasma protein most closely similar to SAP, was only detected within capillary lumina. The appearance of neurocytotoxic SAP in the brain after TBI, and its persistent, selective deposition in cerebral neurones, are consistent with a potential contribution to subsequent neurodegeneration.


Subject(s)
Brain Injuries, Traumatic , Dementia , Humans , Aged , Serum Amyloid P-Component/chemistry , Serum Amyloid P-Component/metabolism , Brain Injuries, Traumatic/metabolism , Brain/metabolism , Blood Proteins/metabolism , Dementia/metabolism , Amyloid beta-Peptides/metabolism
17.
medRxiv ; 2023 Aug 16.
Article in English | MEDLINE | ID: mdl-37645746

ABSTRACT

The direct causes of neurodegeneration underlying Alzheimer's disease (AD) and many other dementias, are not known. Here we identify serum amyloid P component (SAP), a constitutive plasma protein normally excluded from the brain, as a potential drug target. After meta-analysis of three genome-wide association studies, comprising 44,288 participants, cis-Mendelian randomization showed that genes responsible for higher plasma SAP values are significantly associated with AD, Lewy body dementia and plasma tau concentration. These genetic findings are consistent with experimental evidence of SAP neurotoxicity and the strong, independent association of neocortex SAP content with dementia at death. Depletion of SAP from the blood and from the brain, as is provided by the safe, well tolerated, experimental drug, miridesap, may therefore contribute to treatment of neurodegeneration.

18.
Nature ; 440(7088): 1217-21, 2006 Apr 27.
Article in English | MEDLINE | ID: mdl-16642000

ABSTRACT

Complement-mediated inflammation exacerbates the tissue injury of ischaemic necrosis in heart attacks and strokes, the most common causes of death in developed countries. Large infarct size increases immediate morbidity and mortality and, in survivors of the acute event, larger non-functional scars adversely affect long-term prognosis. There is thus an important unmet medical need for new cardioprotective and neuroprotective treatments. We have previously shown that human C-reactive protein (CRP), the classical acute-phase protein that binds to ligands exposed in damaged tissue and then activates complement, increases myocardial and cerebral infarct size in rats subjected to coronary or cerebral artery ligation, respectively. Rat CRP does not activate rat complement, whereas human CRP activates both rat and human complement. Administration of human CRP to rats is thus an excellent model for the actions of endogenous human CRP. Here we report the design, synthesis and efficacy of 1,6-bis(phosphocholine)-hexane as a specific small-molecule inhibitor of CRP. Five molecules of this palindromic compound are bound by two pentameric CRP molecules, crosslinking and occluding the ligand-binding B-face of CRP and blocking its functions. Administration of 1,6-bis(phosphocholine)-hexane to rats undergoing acute myocardial infarction abrogated the increase in infarct size and cardiac dysfunction produced by injection of human CRP. Therapeutic inhibition of CRP is thus a promising new approach to cardioprotection in acute myocardial infarction, and may also provide neuroprotection in stroke. Potential wider applications include other inflammatory, infective and tissue-damaging conditions characterized by increased CRP production, in which binding of CRP to exposed ligands in damaged cells may lead to complement-mediated exacerbation of tissue injury.


Subject(s)
C-Reactive Protein/antagonists & inhibitors , C-Reactive Protein/metabolism , Cardiovascular Diseases/drug therapy , Cardiovascular Diseases/metabolism , Hexanes/pharmacology , Hexanes/therapeutic use , Phosphorylcholine/analogs & derivatives , Animals , C-Reactive Protein/chemistry , C-Reactive Protein/pharmacology , Cardiovascular Diseases/pathology , Cardiovascular Diseases/physiopathology , Complement System Proteins/immunology , Drug Design , Hexanes/administration & dosage , Hexanes/chemistry , Humans , Male , Models, Molecular , Molecular Conformation , Myocardial Infarction/drug therapy , Myocardial Infarction/metabolism , Myocardial Infarction/pathology , Myocardial Infarction/physiopathology , Phosphorylcholine/administration & dosage , Phosphorylcholine/chemistry , Phosphorylcholine/pharmacology , Phosphorylcholine/therapeutic use , Rats , Rats, Wistar
19.
Proc Natl Acad Sci U S A ; 106(18): 7619-23, 2009 May 05.
Article in English | MEDLINE | ID: mdl-19372378

ABSTRACT

New therapeutic approaches in Alzheimer's disease are urgently needed. The normal plasma protein, serum amyloid P component (SAP), is always present in cerebrospinal fluid (CSF) and in the pathognomonic lesions of Alzheimer's disease, cerebrovascular and intracerebral Abeta amyloid plaques and neurofibrillary tangles, as a result of its binding to amyloid fibrils and to paired helical filaments, respectively. SAP itself may also be directly neurocytotoxic. Here, in this unique study in Alzheimer's disease of the bis(d-proline) compound, (R)-1-[6-[(R)-2-carboxy-pyrrolidin-1-yl]-6-oxo-hexanoyl]pyrrolidine-2-carboxylic acid (CPHPC), we observed depletion of circulating SAP and also remarkable, almost complete, disappearance of SAP from the CSF. We demonstrate that SAP depletion in vivo is caused by CPHPC cross-linking pairs of SAP molecules in solution to form complexes that are immediately cleared from the plasma. We have also solved the structure of SAP complexed with phosphothreonine, its likely ligand on hyperphosphorylated tau protein. These results support further clinical study of SAP depletion in Alzheimer's disease and potentially other neurodegenerative diseases.


Subject(s)
Alzheimer Disease/metabolism , Carboxylic Acids/administration & dosage , Pyrrolidines/administration & dosage , Serum Amyloid P-Component/antagonists & inhibitors , Alzheimer Disease/blood , Alzheimer Disease/cerebrospinal fluid , Circular Dichroism , Crystallography, X-Ray , Humans , Mass Spectrometry , Middle Aged , Pilot Projects , Protein Conformation , Serum Amyloid P-Component/cerebrospinal fluid , Serum Amyloid P-Component/chemistry
20.
Lancet ; 375(9709): 132-40, 2010 Jan 09.
Article in English | MEDLINE | ID: mdl-20031199

ABSTRACT

BACKGROUND: Associations of C-reactive protein (CRP) concentration with risk of major diseases can best be assessed by long-term prospective follow-up of large numbers of people. We assessed the associations of CRP concentration with risk of vascular and non-vascular outcomes under different circumstances. METHODS: We meta-analysed individual records of 160 309 people without a history of vascular disease (ie, 1.31 million person-years at risk, 27 769 fatal or non-fatal disease outcomes) from 54 long-term prospective studies. Within-study regression analyses were adjusted for within-person variation in risk factor levels. RESULTS: Log(e) CRP concentration was linearly associated with several conventional risk factors and inflammatory markers, and nearly log-linearly with the risk of ischaemic vascular disease and non-vascular mortality. Risk ratios (RRs) for coronary heart disease per 1-SD higher log(e) CRP concentration (three-fold higher) were 1.63 (95% CI 1.51-1.76) when initially adjusted for age and sex only, and 1.37 (1.27-1.48) when adjusted further for conventional risk factors; 1.44 (1.32-1.57) and 1.27 (1.15-1.40) for ischaemic stroke; 1.71 (1.53-1.91) and 1.55 (1.37-1.76) for vascular mortality; and 1.55 (1.41-1.69) and 1.54 (1.40-1.68) for non-vascular mortality. RRs were largely unchanged after exclusion of smokers or initial follow-up. After further adjustment for fibrinogen, the corresponding RRs were 1.23 (1.07-1.42) for coronary heart disease; 1.32 (1.18-1.49) for ischaemic stroke; 1.34 (1.18-1.52) for vascular mortality; and 1.34 (1.20-1.50) for non-vascular mortality. INTERPRETATION: CRP concentration has continuous associations with the risk of coronary heart disease, ischaemic stroke, vascular mortality, and death from several cancers and lung disease that are each of broadly similar size. The relevance of CRP to such a range of disorders is unclear. Associations with ischaemic vascular disease depend considerably on conventional risk factors and other markers of inflammation. FUNDING: British Heart Foundation, UK Medical Research Council, BUPA Foundation, and GlaxoSmithKline.


Subject(s)
C-Reactive Protein/analysis , Coronary Disease/blood , Risk Assessment , Stroke/blood , Alcohol Drinking/blood , Alcohol Drinking/epidemiology , Biomarkers/blood , Blood Pressure , Body Mass Index , Cholesterol/blood , Coronary Disease/mortality , Coronary Disease/prevention & control , Databases, Factual , Diabetes Mellitus/blood , Diabetes Mellitus/epidemiology , Female , Fibrinogen/analysis , Humans , Interleukin-6/blood , Leukocyte Count , Lung Diseases/blood , Lung Diseases/mortality , Male , Middle Aged , Motor Activity , Neoplasms/blood , Neoplasms/mortality , Regression Analysis , Risk Factors , Serum Albumin , Sex Factors , Smoking/blood , Smoking/epidemiology , Stroke/mortality , Stroke/prevention & control , Triglycerides/blood
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