IMO-8400, a toll-like receptor 7, 8, and 9 antagonist, demonstrates clinical activity in a phase 2a, randomized, placebo-controlled trial in patients with moderate-to-severe plaque psoriasis.

BACKGROUND: Aberrant toll-like receptors (TLRs) 7, 8, and 9 activation by self-nucleic acids is implicated in immune-mediated inflammatory diseases (IMIDs) such as psoriasis. In preclinical IMID models, blocking TLR-activation reduced disease severity. IMO-8400 is a first-in-class, oligonucleotide-based antagonist of TLRs 7, 8, and 9. We evaluated the short-term safety and proof-of-concept for efficacy of IMO-8400 in a first-in-patient phase 2 trial. METHODS: Forty-six psoriasis patients were randomly assigned to IMO-8400 in four dose levels or placebo for 12weeks. Post-treatment follow-up was seven weeks. Primary outcome was incidence of adverse events. Secondary, exploratory outcomes included changes in psoriasis area and severity index (PASI). RESULTS: IMO-8400 across all dose levels did not cause any serious or severe adverse events. The most common treatment-related adverse events were dose-dependent injection-site reactions. All IMO-8400 groups showed clinical improvement, but a clear dose-response relationship and statistically significant differences with placebo were not observed (P=0.26). Eleven (38%) of 29 subjects on IMO-8400 achieved ≥50% PASI-reduction, compared to 1 (11%) of 9 subjects on placebo. Five (17%) and 2 (7%) IMO-8400-treated subjects achieved PASI-75 and PASI-90, respectively, compared to none on placebo. CONCLUSIONS: Short-term IMO-8400-treatment was well tolerated and reduced psoriasis severity. These findings warrant further investigation of endosomal TLR-antagonism as a therapeutic approach in psoriasis and other TLR-mediated IMIDs. TRIAL REGISTRATION: EudraCT 2013-000164-28 and Clinicaltrials.govNCT01899729.

Design, synthesis and biological evaluation of novel antagonist compounds of Toll-like receptors 7, 8 and 9.

Oligonucleotides containing an immune-stimulatory motif and an immune-regulatory motif act as antagonists of Toll-like receptor (TLR)7 and TLR9. In the present study, we designed and synthesized oligonucleotide-based antagonists of TLR7, 8 and 9 containing a 7-deaza-dG or arabino-G modification in the immune-stimulatory motif and 2'-O-methylribonucleotides as the immune-regulatory motif. We evaluated the biological properties of these novel synthetic oligoribonucleotides as antagonists of TLRs 7, 8 and 9 in murine and human cell-based assays and in vivo in mice and non-human primates. In HEK293, mouse and human cell-based assays, the antagonist compounds inhibited signaling pathways and production of a broad range of cytokines, including tumour necrosis factor alpha (TNF-α), interleukin (IL)-12, IL-6, interferon (IFN)-α, IL-1ß and interferon gamma-induced protein (IP)-10, mediated by TLR7, 8 and 9. In vivo in mice, the antagonist compounds inhibited TLR7- and TLR9-mediated cytokine induction in a dose- and time-dependent fashion. Peripheral blood mononuclear cells (PBMCs) obtained from antagonist compound-treated monkeys secreted lower levels of TLR7-, 8- and 9-mediated cytokines than did PBMCs taken before antagonist administration. The antagonist compounds described herein provide novel agents for the potential treatment of autoimmune and inflammatory diseases.

Design of synthetic oligoribonucleotide-based agonists of Toll-like receptor 3 and their immune response profiles in vitro and in vivo.

Double-stranded RNA of viral origin and enzymatically synthesized poly I:C act as agonists of TLR3 and induce immune responses. We have designed and synthesized double-stranded synthetic oligoribonucleotides (dsORNs) which act as agonists of TLR3. Each strand of dsORN contains two distinct segments, namely an alignment segment composed of a heteronucleotide sequence and an oligo inosine (I) or an oligo cytidine (C) segment. We report here the results of studies of dsORNs containing varying lengths and compositions of alignment and oligo I/oligo C segments. dsORNs of 50-mer length with a 15-mer alignment segment and a 35-mer oligo I/oligo C segment form stable duplexes under physiological conditions and induce TLR3-mediated immune responses. dsORNs activated the IRF3 signaling pathway in J774 cells, induced production of cytokines, including IFN-ß, IFN-α, IP-10, IL-12 and IL-6, in murine and human cell-based assays and also induced multiple cytokines following systemic administration in mice and non-human primates.

A Structured Benefit-Risk Assessment Operating Model for Investigational Medicinal Products in the Pharmaceutical Industry.

Robust and transparent formal benefit-risk (BR) analyses for medicinal products represent a means to better understand the appropriate use of medicinal products, and to maximize their value to prescribers and patients. Despite regulatory and social imperatives to conduct structured BR (sBR) assessments, and the availability of a plethora of methodological tools, there exists large variability in the uptake and execution of sBR assessments among pharmaceutical companies. As such, in this paper we present an sBR assessment framework developed and implemented within a large global pharmaceutical company that aims to guide the systematic assessment of BR across the continuum of drug development activities, from first-time-in-human studies through to regulatory submission. We define and emphasize the concepts of Key Clinical Benefits and Key Safety Risks as the foundation for BR analysis. Furthermore, we define and foundationally employ the concepts of sBR and a Core Company BR position as the key elements for our BR framework. We outline 3 simple stages for how to perform the fundamentals of an sBR analysis, along with an emphasis on the weighting of Key Clinical Benefits and Key Safety Risks, and a focus on any surrounding uncertainties. Additionally, we clarify existing definitions to differentiate descriptive, semi-quantitative, and fully quantitative BR methodologies. By presenting our framework, we wish to stimulate productive conversation between industry peers and health authorities regarding best practice in the BR field. This paper may also help facilitate the pragmatic implementation of sBR methodologies for organizations without an established framework for such assessments.

Predictors of in-hospital surgical site infections in surgically managed acetabular fractures: A nationwide analysis.

Background: A surgical site infection (SSI) rate of 4%-8% has been reported in patients who undergo open reduction and internal fixation (ORIF) for acetabular fractures. Studies have identified risk factors for SSI, but none have performed a nationwide analysis of SSI in surgically managed acetabular fracture patients. Methods: The National Inpatient Sample (NIS) database was queried for patients who underwent ORIF for acetabular fractures from 2016 to 2019. Analysis was performed on all patients who underwent ORIF for acetabular fractures, as well as those who only underwent ORIF for isolated acetabular fractures. Clinical characteristics, hospital course, discharge disposition, and hospitalization costs were compared between groups. Multivariate regression analysis was conducted to assess predictors of SSI. Results: 41,725 patients undergoing acetabular fracture repair were identified, of which 490 (1.2%) developed SSI during hospitalization. Age (45.90 vs 49.90, p < 0.001) and Injury Severity Scale (5.99 vs 8.30, p < 0.001) were increased in patients who developed SSI. History of hypertension (HTN) (OR = 2.343, 95% CI 1.96-2.80, p < 0.001), longer hospital length of stay (30.27 days vs 10.00 days, p < 0.001) and total charges ($469,005 vs $193,032, p < 0.001) were associated with SSI. Lower rates of routine discharge were seen in SSI patients (OR = 0.333, 95% CI 0.260-0.426, p < 0.001). Higher rates of inpatient death were associated with SSI (OR = 2.210, 95% CI 1.172-4.17, p = 0.019). Multiple procedures in addition to acetabular fracture repair, iliac artery embolization, substance abuse, later time to internal fixation and HTN were predictive of SSI (p < 0.001). Conclusions: Severity of injury, time to fixation, and factors associated with compromised cardiovascular integrity were predictors of SSI. Identifying patients at risk for SSI should lead to clinical maneuvers that may optimize outcome.

Free hemoglobin induction of pulmonary vascular disease: evidence for an inflammatory mechanism.

Cell-free hemoglobin (Hb) exposure may be a pathogenic mediator in the development of pulmonary arterial hypertension (PAH), and when combined with chronic hypoxia the potential for exacerbation of PAH and vascular remodeling is likely more pronounced. We hypothesized that Hb may contribute to hypoxia-driven PAH collectively as a prooxidant, inflammatory, and nitric oxide (NO) scavenger. Using programmable micropump technology, we exposed male Sprague-Dawley rats housed under room air or hypoxia to 12 or 30 mg per day Hb for 3, 5, and 7 wk. Blood pressure, cardiac output, right ventricular hypertrophy, and indexes of pulmonary vascular remodeling were evaluated. Additionally, markers of oxidative stress, NO bioavailability and inflammation were determined. Hb increased pulmonary arterial (PA) pressure, pulmonary vessel wall stiffening, and right heart hypertrophy with temporal and dose dependence in both room air and hypoxic cohorts. Hb induced a modest increase in plasma oxidative stress markers (malondialdehyde and 4-hydroxynonenal), no change in NO bioavailability, and increased lung ICAM protein expression. Treatment with the antioxidant Tempol attenuated Hb-induced pulmonary arterial wall thickening, but not PA pressures or ICAM expression. Chronic exposure to low plasma Hb concentrations (range = 3-10 µM) lasting up to 7 wk in rodents induces pulmonary vascular disease via inflammation and to a lesser extent by Hb-mediated oxidation. Tempol demonstrated a modest effect on the attenuation of Hb-induced pulmonary vascular disease. NO bioavailability was found to be of minimal importance in this model.

Lipoxin A4 and benzo-lipoxin A4 attenuate experimental renal fibrosis.

Unresolved inflammation underlies the development of fibrosis and organ failure. Here, we investigate the potential of the proresolving eicosanoid lipoxinA4 (LXA4) and its synthetic analog benzo-LXA4 to prophylactically modulate fibrotic and inflammatory responses in a model of early renal fibrosis, unilateral ureteric obstruction (UUO). Male Wistar rats (Animalia, Chordata, Rattus norvegicus) were injected intravenously with vehicle (0.1% ethanol), LXA4 (45 µg/250-g rat), or benzo-LXA4 (15 µg/250-g rat) 15 min prior to surgery and sacrificed 3 d postligation. Renal gene and protein expression, collagen deposition, macrophage infiltration, and apoptosis were analyzed using manipulated kidneys from sham operations as control. Lipoxins (LXs) attenuated collagen deposition and renal apoptosis (P<0.05) and shifted the inflammatory milieu toward resolution, inhibiting TNF-α and IFN-γ expression, while stimulating proresolving IL-10. LXs attenuated UUO-induced activation of MAP kinases, Akt, and Smads (P<0.05) in injured kidneys. We explored whether the underlying mechanism reflected LX-induced modulation of fibroblast activation. Using cultured rat renal NRK-49F fibroblasts, we report that LXA4 (1 nM) inhibits TGF-ß1 (10 ng/ml)-induced activation of Smad2 and MAP-kinases (P<0.05), and furthermore, LXA4 reduced TGF-ß1-stimulated PAI-1 luciferase activation (P<0.05) relative to vehicle-stimulated cells. We propose that LXs may represent a potentially useful and novel therapeutic strategy for consideration in the context of renal fibrosis.

Discovery of potent and specific CXCR3 antagonists.

The optimization of a series of 8-aza-quinazolinone analogs for antagonist activity against the CXCR3 receptor is reported. Compounds were optimized to avoid the formation of active metabolites and time-dependent-inhibitors of CYP3A4. In addition, antagonists showed potent against CXCR3 activity in whole blood and optimized to avoid activity in the chromosomal aberration assay. Compound 25 was identified as having the optimal balance of CXCR3 activity and pharmacokinetic properties across multiple pre-clinical species, which are reported herein.

A Structured Methodology to Assess Safety Signal Strength and Inform Causality Assessment.

Causality assessment of safety signals observed with medicinal products is a foundational element of pharmacovigilance and regulatory practice, typically performed by a global introspection process. We have developed a novel, structured methodological framework to support the global introspection process for safety signal causality assessment. This Signal Assessment Guide (SAGe) tool was developed by AstraZeneca and is used internally, both to assess safety signal strength and to inform causality decisions related to safety signals. The term 'safety signal' refers to information arising from one or multiple sources, which suggests a new potentially causal association, or a new aspect of a known association, between an intervention and an adverse event. The key concept underlying the SAGe tool is that safety signal data can be reliably sorted into one of three categories: aggregate safety data, plausibility data, and case-level data. When applying the tool, an evidence grade score (Levels A, B, C, and D) is transparently assigned to the available data in each category. This information can then be summarised and presented for formal decision making regarding causality for safety signals. By using a transparent method to categorise the grade of evidence for causal association, with an option to additionally derive a quantitative strength of safety signal score, the SAGe tool can support the global introspection process for causality decisions, contributing to the quality of safety information for medicinal products provided to healthcare professionals and patients. Our anecdotal experience of using the SAGe tool at AstraZeneca is that it has resulted in more efficient and robust conversations regarding the strength of safety signals and the causality question. Wider use of the SAGe tool may bring increased levels of transparency and consistency to the evaluation of safety signals.

Systemic gene delivery protects the photoreceptors in the retinal degeneration slow mouse.

The retinal degeneration slow (rds/rds) mouse was used to test photoreceptor protection by systemic gene delivery of non-erythropoietic forms of erythropoietin (EPO). Two Epo mutants were generated and packaged into recombinant adeno-associated virus (rAAV) serotype 2/5, controls included rAAV2/5.Epo and rAAV2/5.enhanced green fluorescent protein (eGFP). Mice were injected in the quadriceps at postnatal day seven and analyses were performed at postnatal day 90. Hematocrit, serum EPO levels, and outer nuclear layer (ONL) thickness were quantified. Hematocrit and serum EPO levels in rAAV2/5.eGFP, rAAV2/5.Epo, and rAAV2/5.EpoR103E treated mice were: 46%, 8 mU/ml; 63%, 117 mU/ml; and 52%, 332 mU/ml, respectively. The ONL from rds/rds mice treated with the Epo vectors were approximately twice as thick as the negative controls. This demonstrates that the photoreceptors can be protected without performing an intraocular injection and without increasing the hematocrit to unsafe levels. Intramuscular delivery of rAAV.EpoR103E is an attractive treatment for retinal degenerative diseases.

Reducing maternal mortality: a review of progress and evidence-based strategies to achieve millennium development goal 5.

Maternal mortality represents a major global health challenge. Millennium Development Goal 5 (MDG 5) set a range of targets pertaining to maternal mortality and universal access to reproductive health care. While the realization of these targets seems unlikely, cost-effective population-level approaches in combination with evidence-based interventions targeting the acute management of the major causes of maternal mortality present the potential for considerable progress as the 2015 deadline approaches.

Practical considerations for creating a strategic and proactive clinical safety and pharmacovigilance organization for the future.

BACKGROUND: In 2012, Patient Safety (PS) in AstraZeneca was facing a situation with multiple challenges, scientifically and structurally. OBJECTIVE: To meet these and support AstraZeneca's ambition to return to growth after years of patent expiry, we undertook a project to fundamentally revisit ways of working to create an organisation set up to provide strategic safety in support of drug project decision-making. METHOD: In this paper, we describe the challenges we faced, the project to deliver changes to respond to them, and the methodology used. The project had two main components: creating a new operating model and simplifying the procedural framework. RESULTS: It was delivered in a focused effort by internal PS resources with cross-functional input. The framework simplification resulted in a 71% reduction in procedural documents and a survey of PS staff revealed an increase in satisfaction of 10%-20% across all scores. CONCLUSIONS: With >3 years of observation time, this project has provided AstraZeneca with a PS organisation able to provide strategic safety, supporting successful portfolio delivery, while ensuring patient safety and maintaining compliance with global pharmacovigilance regulations. It has driven efficiency and set the foundation for continued organisational evolution to meet future business needs in an everchanging environment.

Knowledge of disease, diagnosis, adherence and impact of research in an Irish cohort of patients with inflammatory arthritis.

Background: Patient engagement with clinicians results in shared decision making and increased adherence to medication. However, in order for strong patient: clinician partnerships to be achieved, communication barriers need to be identified. Therefore, the aim of this study was to examine the level of understanding of inflammatory arthritis patients and the need for strong patient-partnership in research. Methods: An online anonymous survey was distributed to patients living with inflammatory arthritis which addressed questions about diagnosis, routine tests, medications and how they work, medication adherence, disease flare, heredity, pregnancy, and patient involvement in research. Results: There were 1,873 respondents, 1416 of which had inflammatory arthritis (IA)- rheumatoid arthritis (RA) (65.8%) and psoriatic arthritis (PsA) (34.2%). They were predominantly female (RA 86%, PsA 85 %), aged 55±13 and 50±12 years. Less than 35% of patients had an understanding of diagnostic tests, what was measured and the implication for disease, with 75.5% also concerned about heredity. There was a high level of understanding of how specific medications treat inflammatory arthritis (72.9%). Adherence was also very high (>87%), with the main reasons for stopping medication without the advice of their clinician,  'feeling better' and 'side effects' however  a significant proportion of patients (69.9%) reported a disease-flare following cessation of medication. Patients (31%) were also concerned that inflammatory arthritis reduced their chances of getting pregnant, with only 8% believing arthritis medications were safe to take during pregnancy. Finally, only 9% of patients had ever been asked to participate in a research study. Conclusions: This study demonstrates a need for the development of stronger patient-partnerships with clinicians and researchers in relation to patient education and engagement with research, to create a platform where patients can have meaningful input and involvement in future research studies.

Optimization of a series of quinazolinone-derived antagonists of CXCR3.

The evaluation of the CXCR3 antagonist AMG 487 in clinic trials was complicated due to the formation of an active metabolite. In this Letter, we will discuss the further optimization of the quinazolinone series that led to the discovery of compounds devoid of the formation of the active metabolite that was seen with AMG 487. In addition, these compounds also feature increased potency and good pharmacokinetic properties. We will also discuss the efficacy of the lead compound 34 in a mouse model of cellular recruitment induced by bleomycin.

Discovery and optimization of CRTH2 and DP dual antagonists.

A series of phenylacetic acid derivatives was discovered as CRTH2 antagonists. Modification of the series led to compounds that are also antagonists of DP. Since activation of CRTH2 and DP are believed to play key roles in mediating responses of asthma and other immune diseases, this series was optimized to increase the dual antagonistic activities and improve pharmacokinetic properties. These efforts led to selection of AMG 009 as a clinical candidate.

Impact of secondary structure of toll-like receptor 9 agonists on interferon alpha induction.

Oligodeoxynucleotides containing a CpG motif and double- or multistranded structure-forming sequences act as agonists of Toll-like receptor 9 (TLR9) and induce high levels of interferon alpha (IFN-alpha) in addition to other Th1-type cytokines. In the present study, we evaluated three highly effective IFN-alpha-inducing agonists of TLR9 to determine the type of duplex structures formed and the agonist's ability to induce immune responses, including IFN-alpha induction, in human cell-based assays and in vivo in mice and nonhuman primates. Thermal melting studies showed that two of the agonists evaluated had a single melting transition with similar hyperchromicity in both heating and cooling cycles, suggesting the formation of intermolecular duplexes. A third agonist showed a biphasic melting transition in the heating cycle and a monophasic melting transition with lower hyperchromicity during the cooling cycle, suggesting the formation of both intra- and intermolecular duplexes. All three agonists induced the production of Th1-type cytokines and chemokines, including high levels of IFN-alpha, in human peripheral blood mononuclear cell and plasmacytoid dendritic cell cultures. Subcutaneous administration of the two intermolecular duplex-forming agonists, but not the intramolecular duplex-forming agonist, induced cytokine secretion in mice. In nonhuman primates, the two agonists that formed intermolecular duplexes induced IFN-alpha and IP-10 secretion. On the contrary, the agonist that formed an intramolecular duplex induced only low levels of cytokines in nonhuman primates, suggesting that this type of structure formation is less immunostimulatory in vivo than the other structure. Taken together, the present results suggest that oligonucleotide-based agonists of TLR9 that form intermolecular duplexes induce potent immune responses in vivo.

Techniques and outcomes of total upper and lower eyelid reconstruction.

OBJECTIVES: To describe techniques used for reconstruction of the eyelids following total loss of the upper and lower eyelids and to describe visual and functional outcomes. METHODS: Multicenter, retrospective, interventional case series of all of the patients requiring unilateral reconstruction of both the upper and lower eyelids. RESULTS: Six cases were identified, 1 following trauma and 5 following tumor excision (4 with basal cell carcinoma and 1 with melanoma). The median age was 69 years (range, 18-90 years). Primary repair using preserved tissue was carried out in the case of traumatic avulsion. Following tumor excision, bilamellar repair was performed using composite grafts for the posterior lamella and skin-muscle flaps for the anterior lamella. Graft necrosis occurred in 3 cases (50%). In all of the cases, the reconstructed eyelids were stiff and immobile. Lagophthalmos (6 cases [100%]), ptosis (3 cases [50%]), lower eyelid retraction (3 cases [50%]), and ectropion (2 cases [33%]) were common. Useful vision was retained in all of the cases. CONCLUSIONS: Total eyelid defects are rare and often unanticipated. Adequate corneal protection can be achieved using lamellar repair principles and local tissues; however, poor vascularity demands careful planning, with vascularized flaps favored over free grafts. Reconstructed eyelids have poor function in the setting of total upper and lower eyelid loss, and revision surgery is often required to improve eyelid structure and function.

Carotid extramedial thickness is associated with local arterial stiffness in children.

OBJECTIVES: Experimental evidence suggests that structural changes to the arterial adventitia may be a key vascular determinant of early arterial stiffening, although this has not been directly studied. Accordingly, we hypothesized that in young children, in whom this relationship would not be altered by atheroma, carotid extramedial thickness (EMT), a measure that incorporates the thickness of the arterial adventitia, perivascular tissues and the internal jugular venous wall, would be associated with localized arterial stiffness of the same arterial region. METHODS: We studied 248 healthy prepubescent children (aged 8 years). Carotid diameter and carotid EMT were measured by high-resolution ultrasound. Carotid blood pressure was derived from brachial blood pressure and carotid tonometry. Three measures of localized arterial stiffness (ß stiffness index, distensibility coefficient and incremental modulus of elasticity) were calculated for the common carotid artery. Results were adjusted for heart rate and DBP, two important hemodynamic determinants of arterial stiffness. RESULTS: Carotid EMT was associated with all three measures of arterial stiffness (ß stiffness index: standardized ß = 0.121, P = 0.03; distensibility coefficient: standardized ß = -0.121, P = 0.05; incremental modulus of elasticity: standardized ß = 0.140, P = 0.02). These associations remained significant after adjustment for potential confounders such as sex, height, waist circumference, BMI and body surface area. CONCLUSION: Carotid EMT is associated with the stiffness of the same arterial segment in children, suggesting that the arterial adventitia may be involved in early changes in arterial stiffness during childhood.

Phenotypic assessment of galanin overexpressing and galanin receptor R1 knockout mice in the tail suspension test for depression-related behavior.

RATIONALE: Galanin and its receptors exert inhibitory neuromodulatory control over brain monoamines. Rat studies revealed that galanin expression is upregulated by exposure to stressors and that galanin manipulations modify neuroendocrine and behavioral responses to stress, leading to the hypothesis that galanin mediates depression-related behaviors. METHODS: In the present study, we examined the role of galanin in modulating antidepressant-related behavior in galanin overexpressing transgenic (GAL-tg) mice and galanin receptor R1 knockout (GAL-R1 KO) mice, using the tail suspension test (TST). Quantitative autoradiography for 5-HT(1A)-R and serotonin transporter binding density tested for changes in these two major regulatory components of the 5-HT system in galanin mutant mice. RESULTS: Baseline TST behavior was normal in GAL-tg and GAL-R1 KO mice, and intracerebroventricular administration of galanin failed to alter TST behavior in normal C57BL/6J mice. The TST anti-immobility effects of acute treatment with the serotonin reuptake inhibitor, fluoxetine (0-30 mg/kg), and the norepinephrine reuptake inhibitor, desipramine (0-30 mg/kg), were unaltered in galanin mutant mice. Hippocampal 5-HT(1A)-R density was significantly elevated in GAL-tg and GAL-R1 KO mice, while hippocampal 5-HTT density was reduced in GAL-R1 KO mice, relative to controls. CONCLUSION: Neither pharmacological nor molecular genetic manipulations of galanin altered depression-related profiles in the TST. Possible functional alterations in hippocampal 5-HT neurotransmission may have contributed to these negative results. These preliminary findings provide evidence against the hypothesis that galanin plays a central role in mouse depression-related behaviors. It remains possible that galanin modulates depression-related responses in other experimental paradigms and species.