Combined proteasome and histone deacetylase inhibition attenuates epithelial–mesenchymal transition through E-cadherin in esophageal cancer cells

Objective

Metastasis is thought to be governed partially by induction of epithelial–mesenchymal transition. Combination of proteasome and histone deacetylase inhibitors has shown significant promise, but no studies have investigated this in esophageal cancer. This study investigated effects of vorinostat (histone deacetylase inhibitor) and bortezomib (proteasome inhibitor) on esophageal cancer epithelial–mesenchymal transition.

Methods

Three-dimensional tumor spheroids mimicking tumor architecture were created with esophageal squamous and adenocarcinoma cancer cells. Cells were treated with tumor necrosis factor α (to simulate proinflammatory tumor milieu) and transforming growth factor β (cytokine critical for induction of epithelial–mesenchymal transition). Tumor models were then treated with vorinostat, bortezomib, or both. Cytotoxic assays assessed cell death. Messenger RNA and protein expressions of metastasis suppressor genes were assessed. After treatment, Boyden chamber invasion assays were performed.

Results

Combined therapy resulted in 3.7-fold decrease in adenocarcinoma cell invasion (P = .002) and 2.8-fold decrease in squamous cell invasion (P = .003). Three-dimensional invasion assays demonstrated significant decrease in epithelial–mesenchymal transition after combined therapy. Quantitative reverse transcriptase polymerase chain reaction and Western blot analyses revealed robust rescue of E-cadherin transcription and protein expression after combined therapy. Importantly, inhibition of the E-cadherin gene resulted in abolition of the salutary benefits of combined therapy, highlighting the importance of this metastasis suppressor gene in the epithelial–mesenchymal transition process.

Conclusions

Combined vorinostat and bortezomib therapy significantly decreased esophageal cancer epithelial–mesenchymal transition. This combined therapeutic effect on esophageal cancer epithelial–mesenchymal transition was associated with upregulation of E-cadherin protein expression.

CTSNet classification: 7

Abbreviations and Acronyms: ARHGDIB, Rho guanosine diphosphate dissociation inhibitor β gene, BRMS1, breast cancer metastasis suppressor 1 gene, CDH1, gene for E-cadherin, EMT, epithelial–mesenchymal transition, HDAC, histone deacetylase, GAPDH, gene for glyceraldehyde-3-phosphate dehydrogenase, KISS1, KiSS-1 metastasis-suppressor gene, NME1, protein (NM23A) expressed in nonmetastatic cells 1, PCR, polymerase chain reaction, TGF-β, transforming growth factor β, TNF-α, tumor necrosis factor α