Silencing the Snail-Dependent RNA Splice Regulator ESRP1 Drives Malignant Transformation of Human Pulmonary Epithelial Cells.

Epithelial-to-mesenchymal transition (EMT) is organized in cancer cells by a set of key transcription factors, but the significance of this process is still debated, including in non-small cell lung cancer (NSCLC). Here, we report increased expression of the EMT-inducing transcription factor Snail in premalignant pulmonary lesions, relative to histologically normal pulmonary epithelium. In immortalized human pulmonary epithelial cells and isogenic derivatives, we documented Snail-dependent anchorage-independent growth in vitro and primary tumor growth and metastatic behavior in vivo Snail-mediated transformation relied upon silencing of the tumor-suppressive RNA splicing regulatory protein ESRP1. In clinical specimens of NSCLC, ESRP1 loss was documented in Snail-expressing premalignant pulmonary lesions. Mechanistic investigations showed that Snail drives malignant progression in an ALDH+CD44+CD24- pulmonary stem cell subset in which ESRP1 and stemness-repressing microRNAs are inhibited. Collectively, our results show how ESRP1 loss is a critical event in lung carcinogenesis, and they identify new candidate directions for targeted therapy of NSCLC.Significance: This study defines a Snail-ESRP1 cancer axis that is crucial for human lung carcinogenesis, with implications for new intervention strategies and translational opportunities. Cancer Res; 78(8); 1986-99. ©2018 AACR.

A novel molecular pathway for Snail-dependent, SPARC-mediated invasion in non-small cell lung cancer pathogenesis.

Definition of the molecular pathogenesis of lung cancer allows investigators an enhanced understanding of the natural history of the disease, thus fostering development of new prevention strategies. In addition to regulating epithelial-to-mesenchymal transition (EMT), the transcription factor Snail exerts global effects on gene expression. Our recent studies reveal that Snail is upregulated in non-small cell lung cancer (NSCLC), is associated with poor prognosis, and promotes tumor progression in vivo. Herein, we demonstrate that overexpression of Snail leads to the upregulation of secreted protein, acidic and rich in cysteine (SPARC) in models of premalignancy and established disease, as well as in lung carcinoma tissues in situ. Snail overexpression leads to increased SPARC-dependent invasion in vitro, indicating that SPARC may play a role in lung cancer progression. Bioinformatic analysis implicates transforming growth factor beta (TGF-ß), extracellular signal-regulated kinase (ERK)1/2, and miR-29b as potential intermediaries in Snail-mediated upregulation of SPARC. Both the TGF-ß1 ligand and TGF-ß receptor 2 (TGF-ßR2) are upregulated following Snail overexpression. Treatment of human bronchial epithelial cell (HBEC) lines with TGF-ß1 and inhibition of TGF-ß1 mRNA expression modulates SPARC expression. Inhibition of MAP-ERK kinase (MEK) phosphorylation downregulates SPARC. MiR-29b is downregulated in Snail-overexpressing cell lines, whereas overexpression of miR-29b inhibits SPARC expression. In addition, miR-29b is upregulated following ERK inhibition, suggesting a Snail-dependent pathway by which Snail activation of TGF-ß and ERK signaling results in downregulation of miR-29b and subsequent upregulation of SPARC. Our discovery of pathways responsible for Snail-induced SPARC expression contributes to the definition of NSCLC pathogenesis.

Soluble donor-like MHC class I proteins induce CD4(+)CD25(+)CD8(-)FoxP3(+) cells with potential to ameliorate graft chronic injury.

Conventional immunosuppressive therapies failed to prevent allograft chronic rejection. New approaches to modulate recipient immune response are needed. Donor-like MHC class I soluble proteins demonstrated therapeutic potential to suppress chronic rejection. The present study was designed to clarify the ability of MHC class I soluble proteins to induce T regulatory cells with true regulatory potential in a fully allogeneic rat cardiac transplant model. Donor-like MHC class I proteins upregulate small population of splenic CD8(-) negative CD4(+)CD25(+)FoxP3(+) positive cells. CD4(+) splenocytes after MHC therapy suppress lymphocyte proliferation against donor antigens in vitro. ACI recipients of WF hearts treated with CD4(+) cells, induced with donor-like MHC class I proteins (CD4-MHC), demonstrated stable survival of the transplanted organ (MST >120 days; n=17). Histology revealed that grafts of recipients treated with CD4-MHC had 23.6% vessels affected 100 days postgrafting. On the contrary, hearts obtained from long-term surviving hosts treated with CD4(+) cells induced with high-dose CsA (CD4-CsA) had 50-70% of affected vessels. CD4-MHC class I treated transplants were mostly CD3(-) negative, had low level of mast and FoxP3(+) cell infiltration compared to CD4-CsA treated hearts. Intragraft CD4(+) cells were close to mast cells in morphology. The same graft tissues had similar number of CD4(+) positive cells and mast cells suggesting existence of CD4(+) positive mast cells. On the other hand, a negligible number of FoxP3(+) positive cells in the grafts after CD4-MHC treatment supports the idea of CD4(+) positive FoxP3(+) negative mast cells population. We demonstrate that donor-like MHC class I protein therapy induces population of CD4(+)CD25(+)CD8(-)FoxP3(+) cells with potential to ameliorate development of transplant vascular disease and evoke CD4(+) positive FoxP3 negative mast cells in the secondary hosts.