title: Transcriptional inhibition of type I collagen gene expression in scleroderma fibroblasts by the antineoplastic drug ecteinascidin 743.
abstract: We previously showed that COL1A1 expression is up-regulated at the transcriptional level in systemic sclerosis (SSc) fibroblasts and that the CCAAT-binding factor (CBF) is involved in this increased expression. Ecteinascidin 743 (ET-743) is a chemotherapeutic agent that binds with sequence specificity to the minor groove of DNA and inhibits CBF-mediated transcriptional activation of numerous genes. Therefore, we examined the effects of ET-743 on the increased COL1A1 expression in SSc fibroblasts. The drug caused a potent and dose-dependent inhibition of type I collagen biosynthesis, which reached 70-90% at 700 pM without affecting cell viability. The same drug concentration caused 60-80% reduction in COL1A1 mRNA levels. The stability of the corresponding transcripts was not affected. In vitro nuclear transcription assays demonstrated a 54% down-regulation of COL1A1 transcription. Transient transfections with COL1A1 promoter constructs containing the specific CBF binding sequence into SSc cells previously treated with 700 pM ET-743 failed to show an effect on COL1A1 promoter activity. Furthermore, ET-743 did not affect the binding of CBF or Sp1 transcription factors to their cognate COL1A1 elements. However, treatment with 700 pM ET-743 of stably transfected NIH 3T3 cells expressing a human type II procollagen gene under the control of the human COL1A1 promoter caused a greater than 50% reduction in the production of type II procollagen and a similar decrease in the corresponding type II procollagen transcripts. These results indicate that ET-743 is a potent inhibitor of COL1A1 transcription. However, this effect cannot be explained by a direct effect on CBF binding to the COL1A1 promoter. Although the exact mechanisms responsible for the transcriptional inhibition of COL1A1 by ET-743 are not apparent, our observations suggest that the drug may be an effective agent to decrease collagen overproduction in SSc and other fibrotic diseases.
authors.affiliation: Department of Medicine, Division of Rheumatology, Thomas Jefferson University, Philadelphia, Pennsylvania 19107, USA.
authors.forename: Natalia
authors.initials: N
authors.lastname: Louneva
authors.forename: Biagio
authors.initials: B
authors.lastname: Saitta
authors.forename: David J
authors.initials: DJ
authors.lastname: Herrick
authors.forename: Sergio A
authors.initials: SA
authors.lastname: Jimenez
mesh_terms: D016475:3T3 Cells; D000818:Animals; D018906:Antineoplastic Agents, Alkylating / Q000494:pharmacology*; D001483:Base Sequence; D002460:Cell Line; D024042:Collagen Type I / Q000235:genetics*; D000090602:Collagen Type I, alpha 1 Chain; D024043:Collagen Type II / Q000235:genetics; D004247:DNA / Q000235:genetics; D004149:Dioxoles / Q000494:pharmacology*; D005347:Fibroblasts / Q000187:drug effects / Q000378:metabolism; D015870:Gene Expression / Q000187:drug effects; D006801:Humans; D007546:Isoquinolines / Q000494:pharmacology*; D051379:Mice; D011401:Promoter Regions, Genetic; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D045743:Scleroderma, Diffuse / Q000188:drug therapy* / Q000235:genetics*; D044005:Tetrahydroisoquinolines; D000077606:Trabectedin; D014162:Transfection
chemical_list: D018906:Antineoplastic Agents, Alkylating; D024042:Collagen Type I; D000090602:Collagen Type I, alpha 1 Chain; D024043:Collagen Type II; D004149:Dioxoles; D007546:Isoquinolines; D012333:RNA, Messenger; D044005:Tetrahydroisoquinolines; D004247:DNA; D000077606:Trabectedin