In moving epithelia, stable cell–cell junctions maintained by E-cadherin are critical in mediating multicellular coordination and polarity, mechanotransduction and movement [
6,
22]. Here, we could discriminate at least two types of collective invasion using cytokeratin as an epithelial identifier together with AJ markers and vimentin as EMT and stromal marker: (i) E-cadherin positive and vimentin-negative compact strands in IDC and (ii) thinner and arguably more loose files in ILC, which retain cell–cell interactions but lack both E-cadherin and vimentin. In IDC, all lesions and, per lesion, the vast majority of cells retain E-cadherin expression, consistent with collective epithelial invasion [
8]. The invasive patterns in ILC, although predominantly collective, show higher probability of cell individualization than in IDC. This suggests that both collective and single-cell invasion occur in parallel in ILC and may provide a broader range of metastatic phenotypes. In ILC, CD44 was used to visualize regions of overlapping membrane staining in juxtaposed cells. The pattern of CD44-positive cell–cell interactions in ILC is consistent with an experimental ILC mouse model after somatic inactivation of the E-cadherin gene, where invading cells retain multicellular organization instead of disseminating individually [
23]. Whether CD44 mediates cell–cell adhesion during collective invasion of ILC or rather acts as functionally inert marker for cell–cell interactions remains to be clarified. Both CD44 and hyaluronic acid are expressed in breast cancer cells, thus CD44 could potentially engage to hyaluronic acid present at the counterpart membrane of the contacted cell [
24]. In T cells, CD44 supports cell–cell adhesion, via hyaluronic acid, to the endothelium and initiates transendothelial migration [
19,
25], and cell–cell interactions may additionally be supported via engagement of a heparan and chondroitin sulfate proteoglycan homologue of CD44 between adjacent cells [
18]. Beyond CD44, other adhesion systems, including immunoglobulin family members (e.g. LCAM, ALCAM, L1-CAM, NCAM) may support cell–cell interactions in the absence of E-cadherin in ILC [
26]. Cell–cell junctions in ILC are likely sufficiently stable to mediate cell–cell binding when tissue density is high and extracellular confinements force cells together (“cell-jamming”) [
27].
The molecular variability of collective patterns suggests that local tissue penetration represents a continuum from quiescent epithelium to multicellular epithelial invasion [
28], recapitulating variants of collective invasion during tissue morphogenesis and regeneration [
5]. Likely, such variety of collective invasion patterns is relevant for other epithelial neoplasms, including colorectal and pancreatic cancer [
10].