Targeting the rheumatoid arthritis synovial fibroblast via cyclin dependent kinase inhibition: An early phase trial

Introduction: Targeted biologic therapies demonstrate similar efficacies in rheumatoid arthritis despite distinct mechanisms of action. They also exhibit a ceiling effect, with 10% to 20% of patients achieving remission in clinical trials. None of these therapies target synovial fibroblasts, which drive and maintain synovitis. Seliciclib (R-roscovitine) is an orally available cyclin-dependent kinase inhibitor that suppresses fibroblast proliferation, and is efficacious in preclinical arthritis models. We aim to determine the toxicity and preliminary efficacy of seliciclib in combination with biologic therapies, to inform its potential as an adjunctive therapy in rheumatoid arthritis. Methods and analysis: TRAFIC is a non-commercial, multi-center, rolling phase Ib/IIa trial investigating the safety, tolerability, and efficacy of seliciclib in patients with moderate to severe rheumatoid arthritis receiving biologic therapies. All participants receive seliciclib with no control arm. The primary objective of part 1 (phase Ib) is to determine the maximum tolerated dose and safety of seliciclib over 4 weeks of dosing. Part 1 uses a restricted 1-stage Bayesian continual reassessment method based on a target dose-limiting toxicity probability of 35%. Part 2 (phase IIa) assesses the potential efficacy of seliciclib, and is designed as a single arm, single stage early phase trial based on a Fleming-A’Hern design using the maximum tolerated dose recommended from part 1. The primary response outcome after 12 weeks of therapy is a composite of clinical, histological and magnetic resonance imaging scores. Secondary outcomes include adverse events, pharmacodynamic and pharmacokinetic parameters, autoantibodies, and fatigue. Ethics and dissemination: The study has been reviewed and approved by the North East - Tyne & Wear South Research Ethics Committee (reference 14/NE/1075) and the Medicines and Healthcare Products Regulatory Agency (MHRA), United Kingdom. Results will be disseminated through publication in relevant peer-reviewed journals and presentation at national and international conferences. Trials Registration: ISRCTN, ISRCTN36667085. Registered on September 26, 2014; http://www.isrctn.com/ISRCTN36667085 Current protocol version: Protocol version 11.0 (March 21, 2019

The IĸB protein BCL3 controls osteogenesis and bone health.

OBJECTIVE: IĸB protein B-cell lymphoma 3-encoded protein (BCL3) is a regulator of the NF-κB family of transcription factors. NF-κB signalling fundamentally influences the fate of bone-forming osteoblasts and bone-resorbing osteoclasts, but the role of BCL3 in bone biology has not been investigated. The objective of this study was to evaluate BCL3 in skeletal development, maintenance and osteoarthritic pathology. METHODS: To assess the contribution of BCL3 to skeletal homeostasis, neonatal mice (n = 6-14) lacking BCL3 (Bcl3-/- ) and WT controls were characterised for bone phenotype and density. To reveal the contribution to bone phenotype by the osteoblast compartment in Bcl3-/- mice, transcriptomic analysis of early osteogenic differentiation and cellular function (n = 3-7) were assessed. Osteoclast differentiation and function in Bcl3-/- mice (n = 3-5) was assessed. Adult 20-week Bcl3-/- and WT mice bone phenotype, strength and turnover were assessed. A destabilisation of the medial meniscus (DMM) model of osteoarthritic ostephytogenesis was utilised to understand adult bone formation in Bcl3-/- mice (n = 11-13). RESULTS: Evaluation of Bcl3-/- mice revealed congenitally increased bone density, long bone dwarfism, increased bone biomechanical strength and altered bone turnover. Molecular and cellular characterisation of mesenchymal precursors showed that Bcl3-/- cells display an accelerated osteogenic transcriptional profile that leads to enhanced differentiation into osteoblasts with increased functional activity; which could be reversed with a mimetic peptide. In a model of osteoarthritis-induced osteophytogenesis, Bcl3-/- mice exhibit decreased pathological osteophyte formation (P < 0.05). CONCLUSION: Cumulatively, these findings demonstrate that BCL3 controls developmental mineralisation to enable appropriate bone formation, whilst in a pathological setting it contributes to skeletal pathology

Long-term cultivation of colorectal carcinoma cells with anti-cancer drugs induces drug resistance and telomere elongation: an in vitro study

BACKGROUND: The role of telomerase activation in the expression and/or maintenance of drug resistance is not clearly understood. Therefore, we investigated the relationships, among the telomerase activity, telomere length and the expression of multidrug resistance genes in colorectal cancer cell lines cultivated with anti-cancer drugs. METHODS: LoVo and DLD-1 cells were continuously grown in the presence of both CDDP and 5-FU for up to 100 days. Cell proliferation, telomerase activity, telomere length and the expression of multidrug resistance genes were serially monitored as the PDL increased. RESULTS: The expression of multidrug resistance genes tended to increase as the PDL increased. However, an abnormal aneuploid clone was not detected as far as the cells were monitored by a DNA histogram analysis. Tumor cells showing resistance to anti-cancer drugs revealed a higher cell proliferation rate. The telomere length gradually increased with a progressive PDL. The telomerase activity reached a maximum level at 15 PDL in LoVo cells and at 27 PDL in DLD-1 cells. An increase in the mRNA expression of the telomerase components, especially in hTERT and in hTR, was observed at the same PDLs. CONCLUSIONS: These results suggest that a high telomerase activity and an elongation of telomeres both appear to help maintain and/or increase drug resistance in colorectal cancer cells. Cancer cells with long telomeres and a high proliferative activity may thus be able to better survive exposure to anti-cancer drugs. This is presumably due to an increased chromosome stability and a strong expression of both mdr-1 and MRP genes

Airway sizes and proportions in children quantified by a video-bronchoscopic technique

Background: A quantitative understanding of airway sizes and proportions and a reference point for comparisons are important to a bronchoscopist. The aims of this study were to measure large airway areas, and define proportions and predictors of airway size in children. Methods: A validated videobronchoscope technique was used to measure in-vivo airway cross-sectional areas (cricoid, right (RMS) and left (LMS) main stem and major lobar bronchi) of 125 children. Airway proportions were calculated as ratios of airways to cricoid areas and to endotracheal tube (ETT) areas. Mann Whitney U, T-tests, and one-way ANOVA were used for comparisons and standard univariate and backwards, stepwise multivariate regression analyses were used to define airway size predictors. Results: Airways size increased progressively with increasing age but proportions remained constant. The LMS was 21% smaller than the RMS. Gender differences in airways' size were not significant in any age group or airway site. Cricoid area related best to body length (BL): cricoid area (mm2) = 26.782 + 0.254*BL (cm) while the RMS and LMS area related best to weight: RMS area (mm2) = 23.938 + 0.394*Wt (kg) and LMS area (mm2) = 20.055 + 0.263*Wt (kg) respectively. Airways to cricoid ratios were larger than airway to ETT ratios (p=0.0001). Conclusions: The cricoid and large airways progressively increase in size but maintain constant proportional relationships to the cricoid across childhood. The cricoid area correlates with body length while the RMS and LMS are best predicted by weight. These data provide for quantitative comparisons of airway lesions

Mammalian cell entry genes in Streptomyces may provide clues to the evolution of bacterial virulence

Understanding the evolution of virulence is key to appreciating the role specific loci play in pathogenicity. Streptomyces species are generally non-pathogenic soil saprophytes, yet within their genome we can find homologues of virulence loci. One example of this is the mammalian cell entry (mce) locus, which has been characterised in Mycobacterium tuberculosis. To investigate the role in Streptomyces we deleted the mce locus and studied its impact on cell survival, morphology and interaction with other soil organisms. Disruption of the mce cluster resulted in virulence towards amoebae (Acanthamoeba polyphaga) and reduced colonization of plant (Arabidopsis) models, indicating these genes may play an important role in Streptomyces survival in the environment. Our data suggest that loss of mce in Streptomyces spp. may have profound effects on survival in a competitive soil environment, and provides insight in to the evolution and selection of these genes as virulence factors in related pathogenic organisms

Thermoresponsive Micropatterned Substrates for Single Cell Studies

We describe the design of micropatterned surfaces for single cell studies, based on thermoresponsive polymer brushes. We show that brushes made of poly(N-isopropylacrylamide) grafted at high surface density display excellent protein and cell anti-adhesive properties. Such brushes are readily patterned at the micron scale via deep UV photolithography. A proper choice of the adhesive pattern shapes, combined with the temperature-dependent swelling properties of PNIPAM, allow us to use the polymer brush as a microactuator which induces cell detachment when the temperature is reduced below C

Targeting synovial fibroblast proliferation in rheumatoid arthritis (TRAFIC): an open-label, dose-finding, phase 1b trial

Background Current rheumatoid arthritis therapies target immune inflammation and are subject to ceiling effects. Seliciclib is an orally available cyclin-dependent kinase inhibitor that suppresses proliferation of synovial fibroblasts—cells not yet targeted in rheumatoid arthritis. Part 1 of this phase 1b/2a trial aimed to establish the maximum tolerated dose of seliciclib in patients with active rheumatoid arthritis despite ongoing treatment with TNF inhibitors, and to evaluate safety and pharmacokinetics. Methods Phase 1b of the TRAFIC study was a non-randomised, open-label, dose-finding trial done in rheumatology departments in five UK National Health Service hospitals. Eligible patients (aged ≥18 years) fulfilled the 1987 American College of Rheumatology (ACR) or the 2010 ACR–European League Against Rheumatism classification criteria for rheumatoid arthritis and had moderate to severe disease activity (a Disease Activity Score for 28 joints [DAS28] of ≥3·2) despite stable treatment with anti-TNF therapy for at least 3 months before enrolment. Participants were recruited sequentially to a maximum of seven cohorts of three participants each, designated to receive seliciclib 200 mg, 400 mg, 600 mg, 800 mg, or 1000 mg administered in 200 mg oral capsules. Sequential cohorts received doses determined by a restricted, one-stage Bayesian continual reassessment model, which determined the maximum tolerated dose (the primary outcome) based on a target dose-limiting toxicity rate of 35%. Seliciclib maximum concentration (Cmax) and area under the plasma concentration time curve 0–6 h (AUC0–6) were measured. This study is registered with ISRCTN, ISRCTN36667085. Findings Between Oct 8, 2015, and Aug 15, 2017, 37 patients were screened and 15 were enrolled to five cohorts and received seliciclib, after which the trial steering committee and the data monitoring committee determined that the maximum tolerated dose could be defined. In addition to a TNF inhibitor, ten (67%) enrolled patients were taking conventional synthetic disease modifying antirheumatic drugs. The maximum tolerated dose of seliciclib was 400 mg, with an estimated dose-limiting toxicity probability of 0·35 (90% posterior probability interval 0·18–0·52). Two serious adverse events occurred (one acute kidney injury in a patient receiving the 600 mg dose and one drug-induced liver injury in a patient receiving the 400 mg dose), both considered to be related to seliciclib and consistent with its known safety profile. 65 non-serious adverse events occurred during the trial, 50 of which were considered to be treatment related. Most treatment-related adverse events were mild; 20 of the treatment-related non-serious adverse events contributed to dose-limiting toxicities. There were no deaths. Average Cmax and AUC0–6 were two-times higher in participants developing dose-limiting toxicities. Interpretation The maximum tolerated dose of seliciclib has been defined for rheumatoid arthritis refractory to TNF blockade. No unexpected safety concerns were identified to preclude ongoing clinical evaluation in a formal efficacy trial. Funding UK Medical Research Council, Cyclacel, Research into Inflammatory Arthritis Centre (Versus Arthritis), and the National Institute of Health Research Newcastle and Birmingham Biomedical Research Centres and Clinical Research Facilities

A Prospective Study of Aspirin Use and the Risk of Gastrointestinal Bleeding in Men

Data regarding the influence of dose and duration of aspirin use on risk of gastrointestinal bleeding are conflicting.We conducted a prospective cohort study of 32,989 men enrolled in the Health Professionals Follow-up Study (HPFS) in 1994 who provided biennial aspirin data. We estimated relative risk of major gastrointestinal bleeding requiring hospitalization or a blood transfusion.During 14 years of follow-up, 707 men reported an episode of major gastrointestinal bleeding over 377,231 person-years. After adjusting for risk factors, regular aspirin use (≥2 times/week) had a multivariate relative risk (RR) of gastrointestinal bleeding of 1.32 (95% confidence interval [CI], 1.12-1.55) compared to non-regular use. The association was particularly evident for upper gastrointestinal bleeding (multivariate RR, 1.49; 95% CI, 1.16-1.92). Compared to men who denied any aspirin use, multivariate RRs of upper gastrointestinal bleeding were 1.05 (95% CI 0.71-1.52) for men who used 0.5-1.5 standard tablets/week, 1.31 (95% CI 0.88-1.95) for 2-5 aspirin/week, 1.63 (95% CI, 1.15-2.32) for 6-14 aspirin/week and 2.40 (95% CI, 1.10-5.22) for >14 aspirin/week (P(trend)<0.001). The relative risk also appeared to be dose-dependent among short-term users <5 years; P(trend)<.001) and long-term users (≥5 years; P(trend) = 0.015). In contrast, after controlling for dose, increasing duration of use did not appear to be associated with risk (P(trend) = 0.749).Regular aspirin use increases the risk of gastrointestinal bleeding, especially from the upper tract. However, risk of bleeding appears to be more strongly related to dose than to duration of use. Risk of bleeding should be minimized by using the lowest effective dose among short-term and long-term aspirin users

Lifetime environmental tobacco smoke exposure and the risk of chronic obstructive pulmonary disease

BACKGROUND: Exposure to environmental tobacco smoke (ETS), which contains potent respiratory irritants, may lead to chronic airway inflammation and obstruction. Although ETS exposure appears to cause asthma in children and adults, its role in causing COPD has received limited attention in epidemiologic studies. METHODS: Using data from a population-based sample of 2,113 U.S. adults aged 55 to 75 years, we examined the association between lifetime ETS exposure and the risk of developing COPD. Participants were recruited from all 48 contiguous U.S. states by random digit dialing. Lifetime ETS exposure was ascertained by structured telephone interview. We used a standard epidemiologic approach to define COPD based on a self-reported physician diagnosis of chronic bronchitis, emphysema, or COPD. RESULTS: Higher cumulative lifetime home and work exposure were associated with a greater risk of COPD. The highest quartile of lifetime home ETS exposure was associated with a greater risk of COPD, controlling for age, sex, race, personal smoking history, educational attainment, marital status, and occupational exposure to vapors, gas, dusts, or fumes during the longest held job (OR 1.55; 95% CI 1.09 to 2.21). The highest quartile of lifetime workplace ETS exposure was also related to a greater risk of COPD (OR 1.36; 95% CI 1.002 to 1.84). The population attributable fraction was 11% for the highest quartile of home ETS exposure and 7% for work exposure. CONCLUSION: ETS exposure may be an important cause of COPD. Consequently, public policies aimed at preventing public smoking may reduce the burden of COPD-related death and disability, both by reducing direct smoking and ETS exposure

Diversity and dynamics of bacterial communities in early life stages of the Caribbean coral Porites astreoides

In this study, we examine microbial communities of early developmental stages of the coral Porites astreoides by sequence analysis of cloned 16S rRNA genes, terminal restriction fragment length polymorphism (TRFLP), and fluorescence in situ hybridization (FISH) imaging. Bacteria are associated with the ectoderm layer in newly released planula larvae, in 4-day-old planulae, and on the newly forming mesenteries surrounding developing septa in juvenile polyps after settlement. Roseobacter clade-associated (RCA) bacteria and Marinobacter sp. are consistently detected in specimens of P. astreoides spanning three early developmental stages, two locations in the Caribbean and 3 years of collection. Multi-response permutation procedures analysis on the TRFLP results do not support significant variation in the bacterial communities associated with P. astreoides larvae across collection location, collection year or developmental stage. The results are the first evidence of vertical transmission (from parent to offspring) of bacteria in corals. The results also show that at least two groups of bacterial taxa, the RCA bacteria and Marinobacter, are consistently associated with juvenile P. astreoides against a complex background of microbial associations, indicating that some components of the microbial community are long-term associates of the corals and may impact host health and survival