Possible involvement of ΔNp63 downregulation in the invasion and metastasis of oral squamous cell carcinoma via induction of a mesenchymal phenotype

Abstract

Epithelial-to-mesenchymal transition (EMT), an essential developmental program, is involved in tumor progression. ΔNp63, a homolog of p53, is associated with the EMT program, but the detailed mechanism remains to be elucidated. In this study, we investigated the role of ΔNp63 in EMT during progression of oral squamous cell carcinoma (OSCC). Five OSCC cell lines and specimens from 78 patients with OSCC were used. The expressions of ΔNp63, p63α, p63β and epithelial markers (cytokeratins 5 and 14) was detected in the OSCC cells, but not in SQUU-B cells (high metastatic potential). E-cadherin was expressed in all OSCC cells. Mesenchymal markers were strongly expressed in the SQUU-B cells. Knockdown of endogenous ΔNp63 in HSC-2 cells induced morphological changes to the spindle shape, decreased the expression of epithelial markers, increased the expression of mesenchymal markers, increased migration and reduced proliferation. By contrast, SQUU-B cells overexpressing ΔNp63β showed changed their morphology from stromal cell-like to epithelial cells. However, E-cadherin expression was not affected by ΔNp63 knockdown or overexpression. Immunohistochemical staining revealed that cancer cells expressing vimentin were found at the invasive front in the OSCC specimens. The intensity of ΔNp63 expression was also decreased in these cells. Interestingly, the vimentin positivity or decreased intensity of ΔNp63 was positively associated with metastases and poor prognosis in the OSCC patients. These results indicated that ΔNp63 downregulation in cancer cells induces a mesenchymal phenotype that is related to tumor progression of OSCC.

Tumor‐suppressive roles of ΔNp63β‐miR‐205 axis in epithelial–mesenchymal transition of oral squamous cell carcinoma via targeting ZEB1 and ZEB2

“…In our previous study, we showed that down‐regulated vimentin and down‐regulated E‐cadherin expression was found in the oral cancer cells at the invasive front. Interestingly, the vimentin positive rate or the presence of decreased intensity of ΔNp63 was positively associated with the frequencies of metastases and poor prognosis in the OSCC patients (Goto et al, ). These results indicated that ΔNp63 down‐regulation in cancer cells is associated with mesenchymal phenotype in tumor progression of OSCC.…”

“…SQUU‐B cells show histologically invasive growth and have high metastatic potential (86.7% incidence of cervical lymph node metastasis), whereas SQUU‐A cells demonstrate expansive growth and have low metastatic ability (Morifuji, Taniguchi, Sakai, Nakabeppu, & Ohishi, ). SQUU‐BO cells were generated by transfection with ΔNp63β expression vector, as previously described (Goto et al, ). All cell lines were maintained in a humidified atmosphere of 5% CO 2 at 37 °C, and cultured in Dulbecco's modified Eagle's medium (DMEM)/F‐12 (Life Technologies, Carlsbad, CA) supplemented with 10% fetal bovine serum (FBS) and 100 units/ml penicillin/streptomycin.…”

“…Conversely, ΔNp63 exerts dominant‐negative activities against TAp63 and p53, and is thus considered an oncoprotein (Higashikawa et al, ). Our previous data demonstrated that down‐regulation of ΔNp63 accompanied with EMT and that re‐expression of ΔNp63β by stable cDNA transfection in OSCC cell lines reverted the EMT phenotype (Goto et al, ). However, the detail mechanisms of ΔNp63β‐mediated EMT remain to be unclear.…”

“…Other zinc finger transcription factors, including zinc finger E‐box binding homeobox (ZEB) 1, Slug and Twist have also been reported to repress E‐cadherin expression and induce EMT (Adhikary et al, ; Sarrio et al, ; Yang et al, ). Previously, we have also studied on the association of p63 gene, a homolog of the p53 tumor suppressor gene, with EMT in OSCC cells (Goto et al, ; Matsubara et al, ). p63 has two different promoter domains that generate two protein isoforms, TAp63 and ΔNp63 (Levine, Tomasini, McKeon, Mak, & Melino, ).…”

“…As samples for miRNA microarray, we used two clones established from an OSCC cell line SQUU‐B, which lacks ΔNp63 expression and displays EMT phenotypes. One is SQUU‐BO cell overexpressing ΔNp63β, and another is SQUU‐BC cell transfected with empty vector (Goto et al, ). By comparing expression profiles of the two OSCC clones, we focused on miR‐205, which is involved in epithelial differentiation (Ryan, Oliveira‐Fernandes, & Lavker, ), because miR‐205 expression was remarkably increased in SQUU‐BO cells.…”