Comparative nonclinical assessments of the biosimilar PF-06410293 and originator adalimumab.

Adalimumab, a recombinant fully human monoclonal antibody targeting tumor necrosis factor (TNF), is approved in the United States and Europe to treat various inflammatory and autoimmune indications. Biosimilars are approved biologics highly similar, but not identical, to approved biotherapeutics. To support clinical development of PF-06410293, an adalimumab biosimilar, nonclinical studies evaluated the structural, functional, toxicologic, and toxicokinetic similarity to originator adalimumab sourced from the United States (adalimumab-US) and European Union (adalimumab-EU). Structural similarity was assessed by peptide mapping. Biologic activity was measured via inhibition of TNF-induced apoptosis and Fc-based functionality assessments. In vivo nonclinical similarity was evaluated in a toxicity study in cynomolgus monkeys administered subcutaneous PF-06410293 or adalimumab-EU (0 or 157 mg/kg/week). Peptide mapping demonstrated PF-06410293, adalimumab-US, and adalimumab-EU had identical amino acid sequences. Comparative functional and binding assessments were similar. Effects of PF-06410293 and adalimumab-EU were similar and limited to pharmacologically mediated decreased cellularity of lymphoid follicles and germinal centers in spleen. Toxicokinetics were similar; maximum plasma concentration and area-under-the-concentration-time curve ratio of PF-06410293:adalimumab-EU ranged from 1.0 to 1.2. These studies supported PF-06410293 entry into clinical development. Many regulatory agencies now only request nonclinical in vivo testing if there is residual uncertainty regarding biosimilarity after in vitro analytical studies.

Selective inhibition of BTK prevents murine lupus and antibody-mediated glomerulonephritis.

Autoantibody production and immune complex deposition within the kidney promote renal disease in patients with lupus nephritis. Thus, therapeutics that inhibit these pathways may be efficacious in the treatment of systemic lupus erythematosus. Bruton's tyrosine kinase (BTK) is a critical signaling component of both BCR and FcR signaling. We sought to assess the efficacy of inhibiting BTK in the development of lupus-like disease, and in this article describe (R)-5-amino-1-(1-cyanopiperidin-3-yl)-3-(4-[2,4-difluorophenoxy]phenyl)-1H-pyrazole-4-carboxamide (PF-06250112), a novel highly selective and potent BTK inhibitor. We demonstrate in vitro that PF-06250112 inhibits both BCR-mediated signaling and proliferation, as well as FcR-mediated activation. To assess the therapeutic impact of BTK inhibition, we treated aged NZBxW_F1 mice with PF-06250112 and demonstrate that PF-06250112 significantly limits the spontaneous accumulation of splenic germinal center B cells and plasma cells. Correspondingly, anti-dsDNA and autoantibody levels were reduced in a dose-dependent manner. Moreover, administration of PF-06250112 prevented the development of proteinuria and improved glomerular pathology scores in all treatment groups. Strikingly, this therapeutic effect could occur with only a modest reduction observed in anti-dsDNA titers, implying a critical role for BTK signaling in disease pathogenesis beyond inhibition of autoantibody production. We subsequently demonstrate that PF-06250112 prevents proteinuria in an FcR-dependent, Ab-mediated model of glomerulonephritis. Importantly, these results highlight that BTK inhibition potently limits the development of glomerulonephritis by impacting both cell- and effector molecule-mediated pathways. These data provide support for evaluating the efficacy of BTK inhibition in systemic lupus erythematosus patients.

Quantitative analysis of target coverage and germinal center response by a CXCL13 neutralizing antibody in a T-dependent mouse immunization model.

PURPOSE: Study the impact of CXCL13 neutralization on germinal center (GC) response in vivo, and build quantitative relationship between target coverage and pharmacological effects at the target tissue. METHODS: An anti-CXCL13 neutralizing monoclonal antibody was dosed in vivo in a T-dependent mouse immunization (TDI) model. A quantitative site-of-action (SoA) model was developed to integrate antibody PK and total CXCL13 levels in serum and spleen towards estimating target coverage as a function of dose. To aid in the SoA model development, a radio-labeled study using [I(125)] CXCL13 was conducted in mice. Model estimated target coverage was linked to germinal center response using a sigmoidal inhibitory effect model. RESULTS: In vivo studies demonstrated that CXCL13 inhibition led to an architectural change in B-cell follicles, dislocation of GCs and a significant reduction in the GC absolute numbers per square area (GC/mm(2)). The SoA modeling analysis indicated that ~79% coverage in spleen was required to achieve 50% suppression of GC/mm(2). The 3 mg/kg dose with 52% spleen coverage resulted in no PD suppression, whereas 30 mg/kg with 93% coverage achieved close to maximum PD suppression, highlighting the steepness of PD response. CONCLUSIONS: This study showcases an application of SoA modeling towards a quantitative understanding of CXCL13 pharmacology.

The integrated effects of leader-member exchange social comparison on job performance and OCB in the Chinese context.

Although it has been long recognized that leader-member exchange social comparison (LMXSC) has critical implications for employee productivity, little attention has been given to systematically exploring the effects of LMXSC on employee performance in a specific cultural context. Integrating social exchange theory with social comparison theory, we examine a dual process model to explain how and when LMXSC affects employee performance outcomes in the Chinese context. Results based on multiphase, multisource data from China revealed that the mediating roles of employees' perceived obligation toward the leader and self-esteem are examined simultaneously in the relationship between LMXSC and job performance and organizational citizenship behavior (OCB). Additionally, guanxi strengthens the connection between LMXSC and perceived obligation, while Zhongyong thinking erodes the connection between LMXSC and self-esteem. Taken together, these findings enhance our understanding of LMXSC in China.

Creative Performance and Conflict through the Lens of Humble Leadership: Testing a Moderated Mediation Model.

This study developed and tested a moderated mediation model by examining the relationships between humble leadership (HL), emotional intelligence, employee conflict (EC), and creative performance (CP), using resource-based theory as the theoretical foundation. We conducted a cross-sectional survey of 322 employees and their immediate supervisors (n = 53) from the telecom sector in Pakistan. The data was analyzed using AMOS 21 and SPSS 26. The results demonstrate that HL has a positive effect on creative performance and a negative relationship with employee conflict. Furthermore, employee conflict has a negative impact on CP and mediates the impact of HL on CP. Moreover, a leader's emotional intelligence moderates the negative relationship between HL and EC. Finally, this study reveals that EI moderates the indirect effects of HL on CP. The conclusions and implications are discussed at the end of this paper.

Thrombospondin-I is a critical modulator in non-alcoholic steatohepatitis (NASH).

Non-alcoholic fatty liver disease (NAFLD) is a progressive liver disease characterized by dysregulated lipid metabolism and chronic inflammation ultimately resulting in fibrosis. Untreated, NAFLD may progress to non-alcoholic steatohepatitis (NASH), cirrhosis and death. However, currently there are no FDA approved therapies that treat NAFLD/NASH. Thrombospondin-I (TSP-1) is a large glycoprotein in the extracellular matrix that regulates numerous cellular pathways including transforming growth factor beta 1 (TGF-ß1) activation, angiogenesis, inflammation and cellular adhesion. Increased expression of TSP-1 has been reported in various liver diseases; however, its role in NAFLD/NASH is not well understood. We first examined TSP-1 modulation in hepatic stellate cell activation, a critical initiating step in hepatic fibrosis. Knockdown or inhibition of TSP-1 attenuated HSC activation measured by alpha smooth muscle actin (α-SMA) and Collagen I expression. To investigate the impact of TSP-1 modulation in context of NAFLD/NASH, we examined the effect of TSP-1 deficiency in the choline deficient L-amino acid defined high fat diet (CDAHFD) model of NASH in mice by assessing total body and liver weight, serum liver enzyme levels, serum lipid levels, liver steatosis, liver fibrosis and liver gene expression in wild type (WT) and TSP-1 null mice. CDAHFD fed mice, regardless of genotype, developed phenotypes of NASH, including significant increase in liver weight and liver enzymes, steatosis and fibrosis. However, in comparison to WT, CDAHFD-fed TSP-1 deficient mice were protected against numerous NASH phenotypes. TSP-1 null mice exhibited a decrease in serum lipid levels, inflammation markers and hepatic fibrosis. RNA-seq based transcriptomic profiles from the liver of CDAHFD fed mice determined that both WT and TSP-1 null mice exhibited similar gene expression signatures following CDAHFD, similar to biophysical and histological assessment comparison. Comparison of transcriptomic profiles based on genotype suggested that peroxisome proliferator activated receptor alpha (PPARα) pathway and amino acid metabolism pathways are differentially expressed in TSP-1 null mice. Activation of PPARα pathway was supported by observed decrease in serum lipid levels. Our findings provide important insights into the role of TSP-1 in context of NAFLD/NASH and TSP-1 may be a target of interest to develop anti-fibrotic therapeutics for NAFLD/NASH.

CXCL10/CXCR3 axis is associated with disease activity and the development of mucocutaneous lesions in patients with Behçet's disease.

The objective of this study was to investigate CXC chemokines and its receptor in patients with Behcet's disease (BD) and their associations with disease activity. Blood samples were collected from 109 BD patients and 36 age- and sex-matched healthy controls (HCs). Twenty-two follow-up blood samples were collected in BD patients. Serum CXC chemokines (CXCL1, CXCL8, CXCL9, CXCL10, CXCL12, CXCL13 and CXCL16) and cell surface marker expression (CD3, CD4 and CXCR3) in peripheral blood mononuclear cells (PBMCs) were assayed. Clinical features including disease activity were evaluated at the time of blood collection. CXCR3 expression in skin and intestinal lesions from BD patients and HCs was assessed via immunohistochemistry. Serum CXCL10 levels were correlated with disease activity in terms of Behçet's Disease Current Activity Form (BDCAF) (p < 0.001). In follow-up BD patients, changes in serum CXCL10 levels tended to be correlated with those of BDCAF. The percentage of CXCR3 expression in CD3-positive cells in PBMCs was inversely correlated with serum CXCL10 levels in BD patients (p = 0.022). By immunohistochemistry, the number of CXCR3-positive mononuclear cells was higher in skin and intestinal lesions of BD patients than in those of HCs. These results suggest that the CXCL10/CXCR3 axis may contribute to the pathogenesis of BD.

Soluble Fn14 Is Detected and Elevated in Mouse and Human Kidney Disease.

The cytokine TWEAK and its cognate receptor Fn14 are members of the TNF/TNFR superfamily and are upregulated in tissue injury to mediate local tissue responses including inflammation and tissue remodeling. We found that in various models of kidney disease, Fn14 expression (mRNA and protein) is upregulated in the kidney. These models include: lupus nephritis mouse models (Nephrotoxic serum Transfer Nephritis and MRL.Faslpr/lpr), acute kidney injury models (Ischemia reperfusion injury and Folic acid injury), and a ZSF-1 diabetic nephropathy rat model. Fn14 expression levels correlate with disease severity as measured by disease histology. We have also shown for the first time the detection of soluble Fn14 (sFn14) in the urine and serum of mice. Importantly, we found the sFn14 levels are markedly increased in the diseased mice and are correlated with disease biomarkers including proteinuria and MCP-1. We have also detected sFn14 in human plasma and urine. Moreover, sFn14 levels, in urine are significantly increased in DN patients and correlated with proteinuria and MCP-1 levels. Thus our data not only confirm the up-regulation of Fn14/TWEAK pathway in kidney diseases, but also suggest a novel mechanism for its regulation by the generation of sFn14. The correlation of sFn14 levels and disease severity suggest that sFn14 may serve as a potential biomarker for both acute and chronic kidney diseases.

Loss of coiled-coil domain containing 80 negatively modulates glucose homeostasis in diet-induced obese mice.

Coiled-coil domain containing 80 (Ccdc80) is a secreted protein highly enriched in mouse and human white adipose tissue (WAT) that plays an important role during adipocyte differentiation in vitro. To investigate the physiological function of Ccdc80 in energy and glucose homeostasis, we generated mice in which the gene encoding Ccdc80 was disrupted. Mice lacking Ccdc80 showed increased sensitivity to diet-induced hyperglycemia and glucose intolerance while displaying reduced glucose-stimulated insulin secretion in vivo. Gene expression analysis by microarray revealed that only 10 transcripts were simultaneously altered in pancreas, skeletal muscle, and WAT from Ccdc80(-/-) mice, including some components of the circadian clock. Expression of the core clock member Arntl/Bmal1 was reduced whereas that of the oscillating transcription factors Dbp and Tef was increased in all tissues examined. Furthermore, knockdown of Ccdc80 in 3T3-L1 cells led to an increase of Dbp mRNA levels during adipocyte differentiation, suggesting that Ccdc80 might be involved in the regulation of this gene in a cell-autonomous manner. Importantly, transcriptional alterations in Ccdc80(-/-) mice were associated with changes in feeding behavior, increased caloric intake, decreased energy expenditure, and obesity. Taken together, our results suggest that Ccdc80 is a novel modulator of glucose and energy homeostasis during diet-induced obesity.

Disruption of G protein-coupled receptor 39 impairs insulin secretion in vivo.

GPR39 is a G protein-coupled receptor expressed in liver, gastrointestinal tract, adipose tissue, and pancreas. We have recently shown that young GPR39(-/-) mice have normal body weight, food intake, and fasting glucose and insulin levels. In this study, we examined the role of GPR39 in aging and diet-induced obese mice. Body weight and food intake were similar in wild-type and GPR39(-/-) mice as they aged from 12 to 52 wk or when fed a low-fat/high-sucrose or high-fat/high-sucrose diet. Fifty-two-week-old GPR39(-/-) mice showed a trend toward decreased insulin levels after oral glucose challenge. When fed either a low-fat/high-sucrose or high-fat/high-sucrose diet, GPR39(-/-) mice had increased fed glucose levels and showed decreased serum insulin levels during an oral glucose tolerance test in the face of unchanged insulin tolerance. Pancreas morphology and glucose-stimulated insulin secretion in isolated islets from wild-type and GPR39(-/-) mice were comparable, suggesting that GPR39 is not required for pancreas development or ex vivo insulin secretion. Small interfering RNA-mediated knockdown of GPR39 in clonal NIT-1 beta-cells revealed that GPR39 regulates the expression of insulin receptor substrate-2 and pancreatic and duodenal homeobox-1 in a cell-autonomous manner; insulin receptor substrate-2 mRNA was also significantly decreased in the pancreas of GPR39(-/-) mice. Taken together, our data indicate that GPR39 is required for the increased insulin secretion in vivo under conditions of increased demand, i.e. on development of age-dependent and diet-induced insulin resistance. Thus, GPR39 agonists may have potential for the treatment of type 2 diabetes.