Search

Background Thyroid eye disease is a debilitating, disfiguring, and potentially blinding periocular condition for which no Food and Drug Administration–approved medical therapy is available...

Background Risankizumab is a humanised IgG1 monoclonal antibody that binds to the p19 subunit of interleukin-23, inhibiting this key cytokine and its role in psoriatic inflammation. We aimed to assess the efficacy and safety of risankizumab compared with placebo or ustekinumab in patients with moderate-to-severe chronic plaque psoriasis. Methods UltIMMa-1 and UltIMMa-2 were replicate phase 3, randomised, double-blind, placebo-controlled and active comparator-controlled trials done at 139 sites in Australia, Austria, Belgium, Canada, Czech Republic, France, Germany, Japan, Mexico, Poland, Portugal, South Korea, Spain, and the USA. Eligible patients were 18 years or older, with moderate-to-severe chronic plaque psoriasis. In each study, patients were stratified by weight and previous exposure to tumour necrosis factor inhibitor and randomly assigned (3:1:1) by use of interactive response technology to receive 150 mg risankizumab, 45 mg or 90 mg ustekinumab (weight-based per label), or placebo. Following the 16-week double-blind treatment period (part A), patients initially assigned to placebo switched to 150 mg risankizumab at week 16; other patients continued their originally randomised treatment (part B, double-blind, weeks 16-52). Study drug was administered subcutaneously at weeks 0 and 4 during part A and at weeks 16, 28, and 40 during part B. Co-primary endpoints were proportions of patients achieving a 90% improvement in the Psoriasis Area Severity Index (PASI 90) and a static Physician's Global Assessment (sPGA) score of 0 or 1 at week 16 (non-responder imputation). All efficacy analyses were done in the intention-to-treat population. These trials are registered with ClinicalTrials.gov, numbers NCT02684370 (UltIMMa-1) and NCT02684357 (UltIMMa-2), and have been completed. Findings Between Feb 24, 2016, and Aug 31, 2016, 506 patients in UltIMMa-1 were randomly assigned to receive 150 mg risankizumab (n= 304), 45 mg or 90 mg ustekinumab (n= 100), or placebo (n= 102). Between March 1, 2016, and Aug 30, 2016, 491 patients in UltIMMa-2 were randomly assigned to receive 150 mg risankizumab (n= 294), 45 mg or 90 mg ustekinumab (n= 99), or placebo (n= 98). Co-primary endpoints were met for both studies. At week 16 of UltIMMa-1, PASI 90 was achieved by 229 (75.3%) patients receiving risankizumab versus five (4.9%) receiving placebo (placebo-adjusted difference 70.3% [95% CI 64.0-76.7]) and 42 (42.0%) receiving ustekinumab (ustekinumab-adjusted difference 33.5% [22.7-44.3]; p

Background In this phase 2, randomized, double-blind, placebo-controlled, dose-ranging study, we assessed the efficacy and safety of brodalumab (AMG 827), a human anti–interleukin-17–receptor monoclonal antibody, for the treatment of moderate-to-severe plaque psoriasis. Methods We randomly assigned patients with a score of 12 or higher on the psoriasis area-and-severity index (PASI, on which scores range from 0 to 72, with higher scores indicating more severe disease) and with 10% or more of their body-surface area affected by psoriasis to receive brodalumab (70 mg, 140 mg, or 210 mg at day 1 and weeks 1, 2, 4, 6, 8, and 10 or 280 mg monthly) or placebo. The primary end point was the percentage improvement from baseline in the PASI score at week 12. Secondary end points included improvement of at least 75% and at least 90% in the PASI score and the score on the static physician's global assessment at week 12. Results A total of 198 patients underwent randomization. At week 12, the mean percentage improvem...

Aminopurine (2-AP), a fluorescent analog of ade- nine, has been widely used as a probe for local DNA conformation, since excitation and emission charac- teristics and fluoresence lifetimes of 2-AP vary in a sequence-dependent manner within DNA. Using steady-state and time-resolved fluorescence tech- niques, we report that 2-AP appears to be unusually stacked in the internal positions of ATAT and TATA in duplex DNA. The excitation wavelength maxima for 2-AP within these contexts were red shifted, indi- cating reduced solvent exposure for the fluoro- phore. Furthermore, in these contexts, 2-AP fluorescence was resistant to acrylamide-dependent collisional quenching, suggesting that the fluoro- phore is protected by its stacked position within the duplex. This conclusion was further reinforced by the presence of a secondary peak at 275 nm in the fluorescence excitation spectra that is indicative of efficient excitation energy transfer from nearby non- fluorescent DNA bases. Fluorescence anisotropy decay and internal angular 'wobbling' motion meas- urements of 2-AP within these alternating AT con- texts were also consistent with the fluorophore being highly constrained and immobile within the base stack. When these fluorescence characteris- tics are compared with those of 2-AP within other duplex DNA sequence contexts, they are unique.

The solvent-dependent ground-state conformational equilibrium and excited-state dynamics of 2-(2‘-hydroxyphenyl)benzoxazole have been characterized in several solvents on the femtosecond to nanosecond time scales. The only observable ground-state tautomer is the enol, which exists in equilibrium between the syn- and anti-rotational isomers. In the anti-enol isomer, the phenyl hydroxyl group appears to not interact strongly with solvent but rather forms a strong intramolecular hydrogen bond with the benzoxazole oxygen atom. In the syn-enol isomer, the phenyl hydroxyl proton may interact with solvent or form an internal hydrogen bond with the benzoxazole nitrogen atom. Upon excitation, the proton is transferred from the oxygen atom to the nitrogen atom of the internally hydrogen bonded syn-enol isomer in 170 fs, regardless of the solvent. The lifetime of the resulting excited keto tautomer is solvent dependent and on the order of picoseconds. In addition to these dynamics, several additional dynamic process...

The mechanism of the 3'-5' exonuclease activity of the Klenow fragment of DNA polymerase I has been investigated with a combination of biochemical and spectroscopic techniques. Site-directed mutagenesis was used to make alanine substitutions of side chains that interact with the DNA substrate on the 5' side of the scissile phosphodiester bond. Kinetic parameters for 3'-5' exonuclease cleavage of single- and double-stranded DNA substrates were determined for each mutant protein in order to probe the role of the selected side chains in the exonuclease reaction. The results indicate that side chains that interact with the penultimate nucleotide (Q419, N420, and Y423) are important for anchoring the DNA substrate at the active site or ensuring proper geometry of the scissile phosphate. In contrast, side chains that interact with the third nucleotide from the DNA terminus (K422 and R455) do not participate directly in exonuclease cleavage of single-stranded DNA. Alanine substitutions of Q419, Y423, and R455 have markedly different effects on the cleavage of single- and double-stranded DNA, causing a much greater loss of activity in the case of a duplex substrate. Time-resolved fluorescence anisotropy decay measurements with a dansyl-labeled primer/template indicate that the Q419A, Y423A, and R455A mutations disrupted the ability of the Klenow fragment to melt duplex DNA and bind the frayed terminus at the exonuclease site. In contrast, the N420A mutation stabilized binding of a duplex terminus to the exonuclease site, suggesting that the N420 side chain facilitates the 3'-5' exonuclease reaction by introducing strain into the bound DNA substrate. Together, these results demonstrate that protein side chains that interact with the second or third nucleotides from the terminus can participate in both the chemical step of the exonuclease reaction, by anchoring the substrate in the active site or by ensuring proper geometry of the scissile phosphate, and in the prechemical steps of double-stranded DNA hydrolysis, by facilitating duplex melting.

Background Novel therapies are needed for difficult-to-treat populations of patients with psoriasis. Objective We sought to assess the efficacy and safety of the interleukin-17 Receptor A inhibitor brodalumab in patients with psoriasis with or without a self-reported history of psoriatic arthritis (PsA) and with or without a history of biologic use. Methods Subset analyses of a phase II, randomized, double-blind, placebo-controlled study of brodalumab in patients with moderate-to-severe plaque psoriasis were performed. Improvement from baseline in Psoriasis Area and Severity Index score of 75%, 90%, and 100% at week 12; static Physician Global Assessment (0/1) score; Dermatology Life Quality Index response; and Psoriasis Symptom Inventory response were evaluated within subgroups. Results Efficacy and quality-of-life measures were generally similar between subgroups of patients with or without a history of PsA and with or without a history of biologic use across brodalumab doses and were significantly higher among patients who received brodalumab 140 mg every 2 weeks or 210 mg every 2 weeks versus placebo. Limitations Differences between subgroups were not compared statistically, PsA was self-reported, only skin involvement/symptoms were reported, and reasons for discontinuation of prior biologic were not captured. Conclusion Brodalumab is efficacious in patients with psoriasis with or without a history of PsA or biologic use.

The Klenow fragment of Escherichia coli DNA polymerase I catalyzes template-directed synthesis of DNA and uses a separate 3'-5' exonuclease activity to edit misincorporated bases. The polymerase and exonuclease activities are contained in separate structural domains. In this study, nine Klenow fragment derivatives containing mutations within the polymerase domain were examined for their interaction with model primer-template duplexes. The partitioning of the DNA primer terminus between the polymerase and 3'-5' exonuclease active sites of the mutant proteins was assessed by time-resolved fluorescence anisotropy, utilizing a dansyl fluorophore attached to the DNA. Mutation of N845 or R668 disrupted favorable interactions between the Klenow fragment and a duplex containing a matched terminal base pair but had little effect when the terminus was mismatched. Thus, N845 and R668 are required for recognition of correct terminal base pairs in the DNA substrate. Mutation of N675, R835, R836, or R841 resulted in tighter polymerase site binding of DNA, suggesting that the side chains of these residues induce strain in the DNA and/or protein backbone. A double mutant (N675A/R841A) showed an even greater polymerase site partitioning than was displayed by either single mutation, indicating that such strain is additive. In both groups of mutant proteins, the ability to discriminate between duplexes containing matched or mismatched base pairs was impaired. In contrast, mutation of K758 or Q849 had no effect on partitioning relative to wild type, regardless of DNA mismatch character. These results demonstrate that DNA mismatch recognition is dependent on specific amino acid residues within the polymerase domain and is not governed solely by thermodynamic differences between correct and mismatched base pairs. Moreover, this study suggests a mechanism whereby the Klenow fragment is able to recognize polymerase errors following a misincorporation event, leading to their eventual removal by the 3'-5' exonuclease activity.

Prior to undergoing postsynthetic 3'-5' editing (proofreading), a defective DNA primer terminus must be transferred from the 5'-3' polymerase active site to a remote 3'-5' exonuclease site. To elucidate the mechanisms by which this occurs, we have used time-resolved fluorescence spectroscopy to study the interaction of dansyl-labeled DNA primer/templates with the Klenow fragment of Escherichia coli DNA polymerase I. The dansyl probe is positioned such that when the DNA substrate occupies the polymerase active site, the probe is solvent-exposed and possesses a short average fluorescence lifetime (4.7 ns) and extensive angular diffusion (42.5 degrees). Conversely, when the DNA substrate occupies the exonuclease active site, the probe becomes buried within the protein, resulting in an increase in the average lifetime (14.1 ns) and a decrease in the degree of angular diffusion (14.4 degrees ). If both polymerase and exonuclease binding modes are populated (lower limit approximately 5%), their markedly different fluorescence properties cause the anisotropy to decay with a characteristic "dip and rise" shape. Nonlinear least-squares analysis of these data recovers the ground-state mole fractions of exposed (x(e)) and buried (x(b)) probes, which are equivalent to the equilibrium proportions of the DNA substrate bound at the polymerase and exonuclease sites, respectively. The distribution between the polymerase and exonuclease binding modes is given by the equilibrium partitioning constant K(pe) (equal to x(b)/x(e)). The important determinants of the proofreading process can therefore be identified by changes made to either the protein or DNA that perturb the partitioning equilibrium and hence alter the magnitude of K(pe).

The forte of catalytic antibodies has resided in the control of the ground-state reaction coordinate. A principle and method are now described in which antibodies can direct the outcome of photophysical and photochemical events that take place on excited-state potential energy surfaces. The key component is a chemically reactive optical sensor that provides a direct report of the dynamic interplay between protein and ligand at the active site. To illustrate the concept, we used a trans -stilbene hapten to elicit a panel of monoclonal antibodies that displayed a range of fluorescent spectral behavior when bound to a trans -stilbene substrate. Several antibodies yielded a blue fluorescence indicative of an excited-state complex or “exciplex” between trans -stilbene and the antibody. The antibodies controlled the isomerization coordinate of trans -stilbene and dynamically coupled this manifold with an active-site residue. A step was taken toward the use of antibody-based photochemical sensors for diagnostic and clinical applications.

Introduction Le brodalumab (AMG 827), anticorps monoclonal inhibiteur du recepteur de l’interleukine 17 (IL17), a demontre son efficacite dans le traitement du psoriasis en plaque modere a severe. Cette etude a evalue l’efficacite a long terme et la tolerance du brodalumab chez les sujets issus d’un essai de phase 2 ayant recu du placebo ou du brodalumab (70, 140 ou 210 mg/2 semaines ou 280 mg/4 semaines). Patients et methodes Dans cette etude d’extension en ouvert, les patients ont initialement recu 210 mg de brodalumab toutes les 2 semaines. Suivant un amendement au protocole mis en place environ un an apres l’inclusion, les sujets de ≤ 100 kg ont change de traitement et recu une dose de 140 mg/2 semaines a la visite suivante ; les sujets de > 100 kg ont ete maintenus sous le traitement a la dose de 210 mg/2 semaines. Les criteres d’evaluation ont inclus les scores PASI 75, PASI 90, PASI 100 et les evenements indesirables (EI). Observations Parmi les 181 patients inclus dans l’extension, 165 etaient encore dans l’etude a 48 semaines (groupe 140 mg, n = 48 ; groupe 210 mg, n = 17) et 153 a 96 semaines (groupe 140 mg, n = 105 ; groupe 210 mg, n = 48). A 12 semaines, 95,4 % des patients (IC 95 % : 91,2 ; 98,0) ont atteint le PASI 75, 85,1 % (79,0 ; 90,1) le PASI 90 et 62,9 % (55,2 ; 70,0) le PASI 100. A 48 semaines, 93,3 % (IC 95 % : 88,4 ; 96,6) ont atteint le PASI 75, 83,0 % (76,4 ; 88,4) le PASI 90 et 61,8 % (53,9 ; 69,3) le PASI 100. A 96 semaines, 89,5 % (IC 95 % : 83,6 ; 93,9) ont atteint le PASI 75, 78,4 % (71,1 ; 84,7) le PASI 90 et 52,9 % (44,7 ; 61,1) le PASI 100. Durant les 96 semaines de l’extension, 93,4 % des patients traites ont presentes des EI. Les EI les plus frequents (≥ 10 % des patients) ont inclus rhinopharyngites (25 %), infections des voies respiratoires superieures (18 %), arthralgies (13 %). Des EI de grade ≥ 3 ont ete rapportes chez 22 patients (12,2 %), des EI graves (EIG) chez 12 patients (6,6 %). Les EI relies au traitement ont ete rapportes chez 26,5 % des patients, dont 5 (2,8 %) avec un EI de grade ≥ 3 et 3 (1,7 %) avec un EIG. Les EIG rapportes entre les semaines 48 et 96 ont inclus cholecystite aigue, constipation (consideree comme reliee au traitement), polynephrite et tumeur parathyroide benigne ( n = 1 pour chaque). Un EIG de type adenocarcinome oesophagien a ete rapporte, ainsi qu’une rupture d’anevrisme aortique fatale, tous deux avant la semaine 48. Discussion Lors de toutes les evaluations faites entre la semaine 8 et presque 2 ans apres le debut du traitement par brodalumab, la majorite des patients a atteint le PASI 100. Les EI les plus frequents ont compris des affections communes (par ex. rhinopharyngite, infection des voies respiratoires superieures), principalement de grade ≤ 2. Conclusion Le rapport benefice/risque a long terme du brodalumab justifie de poursuivre les etudes chez les patients atteints de psoriasis en plaque modere a severe.

Structural features of the single and mixed micellar systems of sodium dodecyl sulfate (SDS) and dodecyltrimethylammonium bromide (DTAB) were characterized using the fluorescence probe 6-propionyl-2-(dimethylamino)naphthalene (Prodan). In particular, our investigations capitalized on the spectral sensitivity of Prodan to its environment as well as the extensive solubility of Prodan in solvents of varying polarity and/or hydrophobicity to effectively use a three-mode factor analysis technique to resolve the coincident emission from Prodan in multiple microenvironments of single and mixed micelle systems. Our investigations reveal parameters of Prodan fluorescence that reflect the relative polarities of the surfaces of SDS and DTAB micellar cores, the permeability of the SDS micelle interface, and the heterogeneity of SDS−DTAB mixed micellar systems. In particular, we observe a strong affinity of Prodan for both SDS and DTAB micelles at the water−surfactant interface with the emission λmax of Prodan consist...

Few clinical trials have evaluated the combination of topical corticosteroids plus systemic therapies for psoriasis.We sought to evaluate efficacy and safety of etanercept plus topical clobetasol propionate (CP) foam versus etanercept monotherapy for treatment of moderate to severe plaque psoriasis.Adults with Psoriasis Area and Severity Index (PASI) score greater than or equal to 10 and psoriasis-affected body surface area greater than or equal to 10% were randomized to etanercept with CP as needed to clear (2 up-to-2-week courses, weeks 11-12 and 23-24) or etanercept alone (each arm at 50 mg twice weekly for 12 weeks followed by 50 mg once weekly for 12 weeks).A total of 592 patients enrolled (295 etanercept + CP arm; 297 etanercept arm). At week 12, significant differences were observed for response of 75% improvement in PASI score (primary end point, 65.2% vs 48.3% in the etanercept + CP vs etanercept arms, respectively; P.001), response of 90% improvement in PASI score (29.7% vs 19.4%; P = .009), percentage PASI score improvement (76.5% vs 68.2%; P.001), static physician global assessment of clear/almost clear (63.1% vs 47.3%; P.001), and patient satisfaction with treatment (P = .006). Response of 75% improvement in PASI score and static physician global assessment of clear/almost clear were not significantly different between arms at week 24. Patient satisfaction with treatment (P = .001) and percentage improvement in PASI score (P = .031) were also greater in the etanercept + CP arm compared with etanercept only at week 24. Comparable numbers of adverse events occurred in each arm.No placebo for CP foam was provided in the etanercept arm.Addition of CP to etanercept yielded increased efficacy compared with etanercept alone at week 12 without an increase in treatment-related adverse events.

Psoriasis is a chronic inflammatory disease with a prevalence of approximately 1–5% in the general population.1 The majority of diagnosed patients have plaque psoriasis, and about 20% have moderate to severe disease.2 Because of the chronic nature of psoriasis, patients typically need long-term treatment, even though traditional systemic therapies are often associated with cumulative, dose-related toxicities.3 Thus, the current focus for treatment of patients with moderate to severe psoriasis is intermittent or combination therapy to overcome loss of efficacy over time and the cumulative toxicities of traditional agents.4–7 Etanercept is a fully human, soluble tumour necrosis factor (TNF)–receptor fusion protein that is approved to treat moderate to severe plaque psoriasis at a dosage of 50 mg twice weekly for up to 3 months, followed by 50 mg once weekly.8,9 Etanercept has been shown to improve disease symptoms and health-related quality of life while maintaining an acceptable safety and tolerability profile in patients with moderate to severe plaque psoriasis.10–16 Methotrexate is a systemic therapy that has demonstrated clinical efficacy in patients with psoriasis. In a randomized controlled trial, methotrexate was significantly more effective at 16 weeks than placebo (36·5% vs. 18·9%; P < 0·05) but less effective than adalimumab (79·6%; P < 0·001) in reducing the Psoriasis Area and Severity Index (PASI) by 75% (PASI 75) in patients with moderate to severe plaque psoriasis.17 Similarly, a recent randomized, controlled study reported that infliximab was significantly more effective than methotrexate at 16 weeks in patients with moderate to severe psoriasis in terms of PASI 75.18 Methotrexate has also been found to be effective at inducing and maintaining remission in patients with moderate to severe psoriasis in open-label and retrospective studies and in a small randomized comparative trial.19,20 Combination therapy with methotrexate plus other systemic treatments (e.g. ciclosporin, acitretin) or phototherapy is an approach that is used to treat patients with psoriasis.4,5 Despite its efficacy and acceptable tolerability in patients with rheumatoid arthritis (RA),21–25 the combination of etanercept plus methotrexate has not been widely investigated in psoriasis. One small single-centre case series reported that the combination could be effective in patients with insufficient response to etanercept monotherapy.26 More recently, a small study compared etanercept plus either continuous or tapered/discontinued methotrexate in 59 patients with moderate to severe psoriasis.27 Disease symptoms were greatly improved at 24 weeks in the combination arm and adverse events (AEs) were similar in both groups.27 The objective of this study was to assess the efficacy and safety of the addition of methotrexate to etanercept compared with etanercept monotherapy in patients with moderate to severe plaque psoriasis using a randomized, double-blind, placebo-controlled, multicentre design.

Background Brodalumab (anti–interleukin-17-receptor antibody) was effective in treating moderate to severe psoriasis in a 12-week, dose-ranging, placebo-controlled trial. Objective We sought to evaluate efficacy and safety of long-term brodalumab treatment. Methods In this interim analysis at week 120 of an open-label extension study, patients received brodalumab 210 mg every 2 weeks. Protocol amendments reduced the dose (140 mg) in patients weighing 100 kg or less and subsequently increased the dose (210 mg) in patients with inadequate responses. Efficacy was measured by static physician global assessment and 75% or greater, 90% or greater, or 100% improvement in Psoriasis Area and Severity Index score (PASI-75, PASI-90, and PASI-100, respectively). Results Of 181 patients, 144 completed week 120. Static physician global assessment scores of clear/almost clear and clear were achieved by 90% and 63% of patients, respectively, at week 12 and by 72% and 51% at week 120. The PASI-75, PASI-90, and PASI-100 response rates at week 12 (95%/85%/63%) were sustained through week 120 (86%/70%/51%). Most commonly reported adverse events were nasopharyngitis (26.5%), upper respiratory tract infection (19.9%), arthralgia (16.0%), and back pain (11.0%). Four patients had grade-2 absolute neutrophil count. Limitations There was no control group in this open-label extension. Conclusion Brodalumab demonstrated sustained clinical response and an acceptable safety profile through 120 weeks in patients with moderate to severe psoriasis.

The balance between effective tumor necrosis factor (TNF) blockade to control aggressive autoimmune disease states and adequate remaining TNF activity to confer immunoprotection against infections such as tuberculosis is an important and complex issue. An increased scientific understanding of how each of the TNF antagonist agents affects the complex interactions of the inflammation cascade and apoptosis, and whether the effects are modulatory or destructive, is needed. The data presented in this supplement highlight the need for further research into these key areas, and illustrate our current understanding of the mode of action of TNF blockers as only the tip of the iceberg.