Data from Silencing of Transforming Growth Factor-β1 <i>In situ</i> by RNA Interference for Breast Cancer: Implications for Proliferation and Migration <i>In vitro</i> and Metastasis <i>In vivo</i>

Abstract

&lt;div&gt;Abstract&lt;p&gt;&lt;b&gt;Purpose:&lt;/b&gt; Overexpression of transforming growth factor (TGF)-β has been implicated in promoting immune suppression, tumor angiogenesis, tumor cell migration, and invasion in many cancers, including carcinoma of the breast. Thus, targeted down-regulation of TGF-β1 expression in breast cancer &lt;i&gt;in situ&lt;/i&gt; and determination of its implications would provide new treatment approaches for disease management.&lt;/p&gt;&lt;p&gt;&lt;b&gt;Experimental Design:&lt;/b&gt; Small interfering RNA constructs targeting TGF-β1 were validated and used to develop clonal derivatives of the metastatic breast cancer cell line MDA-MB-435. The cells were used in several &lt;i&gt;in vitro&lt;/i&gt; analyses, including migration, invasion, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide, apoptosis, and signaling assays. A wound-healing assay was used to determine migration of the cells in culture and a Boyden chamber transwell assay was used for invasion. Further, the clones were used in an &lt;i&gt;in vivo&lt;/i&gt; mouse model for the kinetics of tumor growth and gene expression in the primary site and in lungs upon metastasis.&lt;/p&gt;&lt;p&gt;&lt;b&gt;Results:&lt;/b&gt; Inhibition of TGF-β1 expression in MDA-MB-435 cells showed a 35% decrease in migration and a 55% decrease in invasion &lt;i&gt;in vitro&lt;/i&gt;, with a 50% increase in proliferation and no effect on apoptosis. &lt;i&gt;In vivo&lt;/i&gt; analysis indicated a 90% decrease in the number of mice bearing macroscopic lung metastases; however, the primary tumors did not show any difference in the growth kinetics when compared with the parental MDA-MB-435 cells. Analysis of TGF-β signaling pathways in the clonal derivatives showed a decrease in Smad2 activation and an increase in AKT and extracellular signal-regulated kinase activation. Interestingly, analysis of TGF-β receptor expression showed a decrease in both receptor I and II expression in TGF-β1 silenced cells. These results suggest that inhibition of TGF-β1 ligand may act as a negative feedback loop to disrupt the function of all TGF-β isoforms.&lt;/p&gt;&lt;p&gt;&lt;b&gt;Conclusions:&lt;/b&gt; Therapies targeting the TGF-β signaling pathway may be more effective in late-stage disease to prevent organ metastasis but not primary tumor formation and may be combined with other tumor-targeted therapies normally limited by increased circulating TGF-β levels.&lt;/p&gt;&lt;/div&gt;