ABSTRACT
Smooth muscle cells (SMC) in blood vessels are normally growth quiescent and transcriptionally inactive. Our objective was to understand promoter usage and dynamics in SMC acutely exposed to a prototypic growth factor or pro-inflammatory cytokine. Using cap analysis gene expression (FANTOM5 project) we report differences in promoter dynamics for immediate-early genes (IEG) and other genes when SMC are exposed to fibroblast growth factor-2 or interleukin-1ß. Of the 1871 promoters responding to FGF2 or IL-1ß considerably more responded to FGF2 (68.4%) than IL-1ß (18.5%) and 13.2% responded to both. Expression clustering reveals sets of genes induced, repressed or unchanged. Among IEG responding rapidly to FGF2 or IL-1ß were FOS, FOSB and EGR-1, which mediates human SMC migration. Motif activity response analysis (MARA) indicates most transcription factor binding motifs in response to FGF2 were associated with a sharp induction at 1 h, whereas in response to IL-1ß, most motifs were associated with a biphasic change peaking generally later. MARA revealed motifs for FOS_FOS{B,L1}_JUN{B,D} and EGR-1..3 in the cluster peaking 1 h after FGF2 exposure whereas these motifs were in clusters peaking 1 h or later in response to IL-1ß. Our findings interrogating CAGE data demonstrate important differences in promoter usage and dynamics in SMC exposed to FGF2 or IL-1ß.
Subject(s)
Fibroblast Growth Factor 2/pharmacology , Gene Expression Regulation/drug effects , Genes, Immediate-Early , Interleukin-1beta/pharmacology , Myocytes, Smooth Muscle/drug effects , Promoter Regions, Genetic , Cell Movement/drug effects , Cell Proliferation/drug effects , Culture Media, Serum-Free/chemistry , Culture Media, Serum-Free/pharmacology , Early Growth Response Protein 1/genetics , Early Growth Response Protein 1/metabolism , Gene Expression Profiling , Humans , Myocytes, Smooth Muscle/cytology , Myocytes, Smooth Muscle/metabolism , Nucleotide Motifs , Primary Cell Culture , Protein Binding , Proto-Oncogene Proteins c-fos/genetics , Proto-Oncogene Proteins c-fos/metabolism , Signal TransductionABSTRACT
Worldwide, one in three cancers is skin-related, with increasing incidence in many populations. Here, we demonstrate the capacity of a DNAzyme-targeting c-jun mRNA, Dz13, to inhibit growth of two common skin cancer types-basal cell and squamous cell carcinomas-in a therapeutic setting with established tumors. Dz13 inhibited tumor growth in both immunodeficient and immunocompetent syngeneic mice and reduced lung nodule formation in a model of metastasis. In addition, Dz13 suppressed neovascularization in tumor-bearing mice and zebrafish and increased apoptosis of tumor cells. Dz13 inhibition of tumor growth, which required an intact catalytic domain, was due in part to the induction of tumor immunity. In a series of good laboratory practice-compliant toxicology studies in cynomolgus monkeys, minipigs, and rodents, the DNAzyme was found to be safe and well tolerated. It also did not interfere in more than 70 physiologically relevant in vitro bioassays, suggesting a reduced propensity for off-target effects. If these findings hold true in clinical trials, Dz13 may provide a safe, effective therapy for human skin cancer.
