Viktor Martyanov

We identify the cell cycle-regulated mRNA transcripts genome-wide in the osteosarcoma-derived U2OS cell line. This results in 2140 transcripts mapping to 1871 unique cell cycle-regulated genes that show periodic oscillations across multiple synchronous cell cycles. We identify genomic loci bound by the G2/M transcription factor FOXM1 by chromatin immunoprecipitation followed by high-throughput sequencing (ChIP-seq) and associate these with cell cycle-regulated genes. FOXM1 is bound to cell cycle-regulated genes with peak expression in both S phase and G2/M phases. We show that ChIP-seq genomic loci are responsive to FOXM1 using a real-time luciferase assay in live cells, showing that FOXM1 strongly activates promoters of G2/M phase genes and weakly activates those induced in S phase. Analysis of ChIP-seq data from a panel of cell cycle transcription factors (E2F1, E2F4, E2F6, and GABPA) from the Encyclopedia of DNA Elements and ChIP-seq data for the DREAM complex finds that a set of core cell cycle genes regulated in both U2OS and HeLa cells are bound by multiple cell cycle transcription factors. These data identify the cell cycle-regulated genes in a second cancer-derived cell line and provide a comprehensive picture of the transcriptional regulatory systems controlling periodic gene expression in the human cell division cycle.

Systemic sclerosis (SSc) is a rare systemic autoimmune disease characterized by skin and organ fibrosis. The pathogenesis of SSc and its progression are poorly understood. The SSc intrinsic gene expression subsets (inflammatory, fibroproliferative, normal-like, and limited) are observed in multiple clinical cohorts of patients with SSc. Analysis of longitudinal skin biopsies suggests that a patient's subset assignment is stable over 6-12 months. Genetically, SSc is multi-factorial with many genetic risk loci for SSc generally and for specific clinical manifestations. Here we identify the genes consistently associated with the intrinsic subsets across three independent cohorts, show the relationship between these genes using a gene-gene interaction network, and place the genetic risk loci in the context of the intrinsic subsets. To identify gene expression modules common to three independent datasets from three different clinical centers, we developed a consensus clustering procedure based on mutual information of partitions, an information theory concept, and performed a meta-analysis of these genome-wide gene expression datasets. We created a gene-gene interaction network of the conserved molecular features across the intrinsic subsets and analyzed their connections with SSc-associated genetic polymorphisms. The network is composed of distinct, but interconnected, components related to interferon activation, M2 macrophages, adaptive immunity, extracellular matrix remodeling, and cell proliferation. The network shows extensive connections between the inflammatory- and fibroproliferative-specific genes. The network also shows connections between these subset-specific genes and 30 SSc-associated polymorphic genes including STAT4, BLK, IRF7, NOTCH4, PLAUR, CSK, IRAK1, and several human leukocyte antigen (HLA) genes. Our analyses suggest that the gene expression changes underlying the SSc subsets may be long-lived, but mechanistically interconnected and related to a patients underlying genetic risk.

OBJECTIVE: To assess the safety and efficacy of treatment with belimumab in patients with early diffuse cutaneous systemic sclerosis (dcSSc) treated with background mycophenolate mofetil (MMF). METHODS: In this 52-week, investigator-initiated, single-center, double-blind, placebo-controlled, pilot study, 20 patients with dcSSc recently started on MMF were randomized 1:1 to additionally receive belimumab at 10 mg/kg intravenously or placebo. We assessed safety, efficacy, and differential gene expression. RESULTS: In the belimumab group, the median modified Rodnan skin thickness score (MRSS) decreased from 27 (interquartile range [IQR] 26.5, 31) to 18 (IQR 11, 23) (P = 0.039). In the placebo group, the median MRSS decreased from 28 (IQR 22, 28) to 21 (IQR 14, 25) (P = 0.023). The median change in MRSS was -10 (IQR -13, -9) in the belimumab group and -3.0 (IQR -15, -1) in the placebo group (P = 0.411). There were no significant differences between the groups in the number of adverse events (AEs). A significant decrease in expression of B cell signaling and profibrotic genes and pathways was observed in patients with improved MRSS in the belimumab group but not in the placebo group. CONCLUSION: Patients in both treatment groups experienced significant improvements in MRSS. The median difference was greater in the belimumab group but did not achieve statistical significance in this small pilot study. AEs were similar between the groups. Changes in gene expression were consistent with mechanism of action and showed that clinical response to treatment with belimumab is associated with a significant decrease in profibrotic genes and pathways. Additional studies are needed to determine the role of belimumab in the treatment of dcSSc.

INTRODUCTION: Systemic sclerosis is an autoimmune disease characterized by inflammation and fibrosis of the skin and internal organs. We sought to assess the clinical and molecular effects associated with response to intravenous abatacept in patients with diffuse cutaneous systemic. METHODS: Adult diffuse cutaneous systemic sclerosis patients were randomized in a 2:1 double-blinded fashion to receive abatacept or placebo over 24 weeks. Primary outcomes were safety and the change in modified Rodnan Skin Score (mRSS) at week 24 compared with baseline. Improvers were defined as patients with a decrease in mRSS of ≥30% post-treatment compared to baseline. Skin biopsies were obtained for differential gene expression and pathway enrichment analyses and intrinsic gene expression subset assignment. RESULTS: Ten subjects were randomized to abatacept (n = 7) or placebo (n = 3). Disease duration from first non-Raynaud's symptom was significantly longer (8.8 ± 3.8 years vs. 2.4 ± 1.6 years, p = 0.004) and median mRSS was higher (30 vs. 22, p = 0.05) in the placebo compared to abatacept group. Adverse events were similar in the two groups. Five out of seven patients (71%) randomized to abatacept and one out of three patients (33%) randomized to placebo experienced ≥30% improvement in skin score. Subjects receiving abatacept showed a trend toward improvement in mRSS at week 24 (-8.6 ± 7.5, p = 0.0625) while those in the placebo group did not (-2.3 ± 15, p = 0.75). After adjusting for disease duration, mRSS significantly improved in the abatacept compared with the placebo group (abatacept vs. placebo mRSS decrease estimate -9.8, 95% confidence interval -16.7 to -3.0, p = 0.0114). In the abatacept group, the patients in the inflammatory intrinsic subset showed a trend toward greater improvement in skin score at 24 weeks compared with the patients in the normal-like intrinsic subset (-13.5 ± 3.1 vs. -4.5 ± 6.4, p = 0.067). Abatacept resulted in decreased CD28 co-stimulatory gene expression in improvers consistent with its mechanism of action. Improvers mapped to the inflammatory intrinsic subset and showed decreased gene expression in inflammatory pathways, while non-improver and placebos showed stable or reverse gene expression over 24 weeks. CONCLUSIONS: Clinical improvement following abatacept therapy was associated with modulation of inflammatory pathways in skin. TRIAL REGISTRATION: ClinicalTrials.gov NCT00442611. Registered 1 March 2007.