Lymphovascular tumoral emboli form through budding histogenesis

Florid lymphovascular emboli is the diagnostic signature of inflammatory breast cancer (IBC) and other aggressive metastasizing cancers but the genesis of the florid numbers of emboli observed is not readily explanable. Lymphovascular emboli have been presumed to form as a result of lymphovascular invasion but this event is thought to be rare and therefore would not readily explain the large number of emboli observed in IBC. We carried out both animal and in vitro imaging studies with our human xenograft model of inflammatory breast cancer, MARY‐X, which exhibits florid emboli and in vitro spheroids. We also carried out morphometric studies on the density of the lymphovascular emboli in 25 IBC and 250 non‐IBC cases and laser capture microdissection. Animal and in vitro multicolor imaging studies using anti‐E‐cadherin and anti‐podoplanin showed evidence of self‐budding histogenesis within the lymphovascular spaces with one parent embolus giving rise to daughter emboli. Correspondingly, budding spheroidgenesis was observed in vitro . Density studies of tumoral emboli in the human cases showed their numbers distributed over an exponential rather than linear range. By both RT‐PCR and IHC, emboli compared to their non‐embolic invasive carcinoma areas exhibited five‐ten fold higher stem cell markers including Stellar, H19, Rex‐1, Nestin, CD133 and Aldehyde Dehydrogenase 1 (ALDH1) as well as stem cell transcriptional determinants including OCT4, SOX2 and Nanog, and stem cell signaling pathways, eg., Notch3. These studies, while not addressing the genesis of the initial embolus, show conclusively that emboli beget emboli. This process of self‐budding histogenesis can explain the exponential increases in embolic number seen in human cases.