A new model system identifies <em>EGFR/HER2</em> and <em>HER2/HER3</em> heterodimers as potent inducers of oesophageal epithelial cell invasion.

Oesophageal squamous cell carcinomas and oesophageal adenocarcinomas display distinct patterns of ErbB expression and dimers. The functional effects of specific ErbB homo- or heterodimers on oesophageal (cancer) cell behaviour, particularly invasion of early carcinogenesis remains unknown. Here, a new cellular model system for controlled activation of EGFR or HER2 and EGFR/HER2 or HER2/HER3 homo- and heterodimers was studied in non-neoplastic squamous oesophageal epithelial Het-1A cells. EGFR, HER2 and HER3 intracellular domains (ICDs) were fused to dimerization domains (DmrA / DmrA and DmrC), and transduced into Het-1A cells lacking ErbB expression. Dimerization of EGFR, HER2 or EGFR/HER2, HER2/HER3 ICDs was induced by synthetic ligands (A/A or A/C dimerizers). This was accompanied by phosphorylation of the respective EGFR, HER2 and HER3 ICDs and activation of distinct down-stream signalling pathways, such as PLC&gamma;1, Akt, STAT and Src family kinases. Phenotypically, ErbB homo- and heterodimers caused cell rounding and non-apoptotic blebbing in EGFR/HER2 and HER2/HER3 heterodimer cells. In a Transwell assay, cell migration velocity was elevated in HER2-dimer as compared to empty vector cells. In addition, HER2-dimer cells showed in increased cell invasion, reaching significance for induced HER2/HER3 heterodimers (p=0.015). Importantly, in three-dimensional organotypic cultures, empty vector cells grew as a superficial cell layer, resembling oesophageal squamous epithelium. In contrast, induced HER2-dimer cells (HER2 homodimers) were highly invasive into the matrix and formed cell clusters. This was associated with partial loss of CK7 (when HER2 homodimers were modelled) and p63 (when EGFR/HER2 heterodimers were modelled), which suggests a change or loss of squamous cell differentiation. Controlled activation of specific EGFR, HER2 and HER3 homo- and heterodimers caused oesophageal squamous epithelial cell migration and/or invasion, especially in a three dimensional microenvironment, thereby functionally identifying ErbB homo- and heterodimers as important drivers of oesophageal carcinogenesis