The PPARγ Agonist Rosiglitazone Is Antifibrotic for Scleroderma Lung Fibroblasts: Mechanisms of Action and Differential Racial Effects

We present novel data demonstrating that the expression of PPARγ is reduced in lung fibroblasts from black SSc-ILD patients as compared to white patients. Activating PPARγ with the agonist rosiglitazone increased the expression of MMP-1 and inhibited collagen type I in lung fibroblasts isolated from white, but not black, SSc-ILD patients. Blocking the c-Met receptor abolishes rosiglitazone's effects on collagen and MMP-1 in lung fibroblasts isolated from white SSc-ILD patients, while augmenting the expression of the c-Met receptor in fibroblasts from black SSc-ILD patients replicates the effects of rosiglitazone seen in whites. We conclude that PPARγ agonists warrant consideration as potential antifibrotic drugs in patients with SSc-ILD. Differential therapeutic effects might be anticipated especially relative to racial differences and the functional expression of the c-Met receptor

Novel lung imaging biomarkers and skin gene expression subsetting in dasatinib treatment of systemic sclerosis-associated interstitial lung disease.

BackgroundThere are no effective treatments or validated clinical response markers in systemic sclerosis (SSc). We assessed imaging biomarkers and performed gene expression profiling in a single-arm open-label clinical trial of tyrosine kinase inhibitor dasatinib in patients with SSc-associated interstitial lung disease (SSc-ILD).MethodsPrimary objectives were safety and pharmacokinetics. Secondary outcomes included clinical assessments, quantitative high-resolution computed tomography (HRCT) of the chest, serum biomarker assays and skin biopsy-based gene expression subset assignments. Clinical response was defined as decrease of >5 or >20% from baseline in the modified Rodnan Skin Score (MRSS). Pulmonary function was assessed at baseline and day 169.ResultsDasatinib was well-tolerated in 31 patients receiving drug for a median of nine months. No significant changes in clinical assessments or serum biomarkers were seen at six months. By quantitative HRCT, 65% of patients showed no progression of lung fibrosis, and 39% showed no progression of total ILD. Among 12 subjects with available baseline and post-treatment skin biopsies, three were improvers and nine were non-improvers. Improvers mapped to the fibroproliferative or normal-like subsets, while seven out of nine non-improvers were in the inflammatory subset (p = 0.0455). Improvers showed stability in forced vital capacity (FVC) and diffusing capacity for carbon monoxide (DLCO), while both measures showed a decline in non-improvers (p = 0.1289 and p = 0.0195, respectively). Inflammatory gene expression subset was associated with higher baseline HRCT score (p = 0.0556). Non-improvers showed significant increase in lung fibrosis (p = 0.0313).ConclusionsIn patients with SSc-ILD dasatinib treatment was associated with acceptable safety profile but no significant clinical efficacy. Patients in the inflammatory gene expression subset showed increase in skin fibrosis, decreasing pulmonary function and worsening lung fibrosis during the study. These findings suggest that target tissue-specific gene expression analyses can help match patients and therapeutic interventions in heterogeneous diseases such as SSc, and quantitative HRCT is useful for assessing clinical outcomes.Trial registrationClinicaltrials.gov NCT00764309

Establishment of an indirect ELISA for detection of the novel antifibrotic peptide M10

<div><p>Objective</p><p>M10 is a ten amino acid peptide generated from the intracellular cytoplasmic tail of the hepatocyte growth factor (HGF) receptor c-Met following cleavage by caspase-3. Recently we reported that M10 interacts with Smad2 and demonstrates antifibrotic properties in vitro and in vivo and can be advanced into a novel antifibrotic remedy. The current study was undertaken to develop an immunoassay to measure M10 concentration in biological specimens.</p><p>Experimental design</p><p>An Indirect Enzyme-Linked Immunosorbent Assay (ELISA) for detection of M10 in biological fluids was developed using pharmaceutical grade synthetic M10 as a calibrator and commercially available anti-c-Met C12 antibody.</p><p>Results</p><p>M10 ELISA specifically detected in plasma M10, but not a scrambled peptide, following a single intraperitoneal administration of M10 (1mg/kg) to mice. The detection limit was 9.6 ng/ml, and the measuring limit was between 15 ng/ml and 200 ng/ml. The recovery limits of M10 were between 80% and 120%; intra-assay coefficient of variation was between 5.3% and 6.3%; inter-assay coefficient of variation was between 5.0% and 8.0% over the buffer concentration tested in the range from 15 ng /ml to 250 ng /ml. The peak of M10 concentration following a single intraperitoneal injection (1mg/kg) was achieved within 6 hours and declined to minimal levels by 48 hours. The experimentally obtained half-life for M10 was comparable to the theoretically predicted half-life for M10.</p><p>Conclusions</p><p>We have established a highly sensitive ELISA to detect the antifibrotic peptide M10 in plasma samples, which should prove to be a novel tool to study the pharmacokinetics and efficacy of M10 in the treatment of fibroproliferative disorders.</p></div

Effects of primary antibody concentrations on sensitivity in M10 ELISA.

<p>A series of primary antibody dilutions (1:500, 1:600, 1:700, 1:800, 1:900 and 1:1000) in PBS were tested against 1:10,000 (0.2 μg /ml) concentration of detection antibody. Each data point represents the mean value of the corresponding triplicates.</p

The pharmacokinetic parameters of M10.

<p>The pharmacokinetic parameters of M10.</p