Abstract Background Pancreatic ductal adenocarcinoma (PDAC) is a highly metastatic malignancy with limited treatment options. Metastatic dissemination is the principal cause of mortality in PDAC, yet the cellular mechanism by which PDAC tumor cells enter the bloodstream remains unknown. A portal of intravasation is the Tumor Microenvironment of Metastasis (TMEM) doorways. The TMEM doorway is composed of a tumor cell, a Tie2 + macrophage, and endothelial cell, in direct contact triggering a brief, localized vascular opening that permits intravasation. Methods We performed time-lapse intravital microscopy in PDAC mouse models to visualize serum extravasation and tumor cell intravasation. TMEM doorway density was quantified using immunohistochemistry of resected human PDAC specimes. TMEM doorway activity was quantified by aligned immunohistochemistry/immunofluorescence of tumor specimens. Mechanistic underpinnings of intravasation were tested using an in vitro intravasation transendothelial migration (iTEM) assay. Tie2 signaling was inhibited in vivo with the Tie2 inhibitor rebastinib (~ 0.44 mg/day in chow for 3 weeks). A macrophage-specific Tie2 conditional knockout mouse was generated to evaluate macrophage Tie2-mediated PDAC dissemination. The therapeutic impact of Tie2 blockade was evaluated in an orthotopic perioperative PDAC model incorporating distal pancreatectomy and perioperative FOLFIRINOX plus rebastinib. Statistical analyses used included Mann–Whitney and Kruskal–Wallis tests for clinicopathologic comparisons, one-way ANOVA with Tukey’s post hoc test for iTEM, Student’s t-test for two-group comparisons, and Kaplan–Meier survival analysis with log-rank testing. Results In vivo imaging revealed transient, localized vascular openings spatially linked to TMEM doorways. PDAC tumor cell intravasation was observed at TMEM doorways. TMEM doorways were detectable in human PDAC tissues; higher TMEM density was associated with aggressive pathologic factors and was reduced after neoadjuvant therapy. Tie2 inhibition selectively impaired macrophage-driven vascular opening and reduced TMEM doorway activity, diminished tumor cell transendothelial migration, and lowered disseminated tumor cell burden in vivo. In therapeutic studies, Tie2 inhibition combined with FOLFIRINOX improved survival compared with FOLFIRINOX alone. Conclusions Intravasation and dissemination is TMEM doorway mediated in PDAC. TMEM doorway function is mediated by Tie2 signaling. Inhibition of Tie2 pharmacologically and genetically decreases TMEM doorway function and PDAC dissemination. Tie2 inhibition may have therapeutic potential combined with chemotherapy or emerging therapies for PDAC.
Zambalde et al. (Sat,) studied this question.