“Tumor Explants Elucidate a Cascade of Paracrine SHH, WNT, and VEGF Signals Driving Pancreatic Cancer Angiosuppression”

Abstract

The sparse vascularity of Pancreatic Ductal Adenocarcinoma (PDAC) presents a mystery: what prevents this aggressive malignancy from undergoing neoangiogenesis to counteract hypoxia and better support growth? An incidental finding from prior work on paracrine communication between malignant PDAC cells and fibroblasts revealed that inhibition of the Hedgehog (HH) pathway partially relieved angiosuppression, increasing tumor vascularity through unknown mechanisms. Initial efforts to study this phenotype were hindered by difficulties replicating the complex interactions of multiple cell typesin vitro. Here we identify a cascade of paracrine signals between multiple cell types that act sequentially to suppress angiogenesis in PDAC. Malignant epithelial cells promote HH signaling in fibroblasts, leading to inhibition of WNT signaling in fibroblasts and epithelial cells, thereby limiting VEGFR2-dependent activation of endothelial hypersprouting. This cascade was elucidated using human and murine PDAC explant models, which effectively retain the complex cellular interactions of native tumor tissues.

“Tumor-selective treatment of metastatic pancreatic cancer with an engineered, probiotic living drug”

“…Lysates from 4 of the 9 toxin-producing strains (Supp Fig 1A) were effective at reducing the viability of nearly all PDAC lines examined: Theta toxin, Magainin, Heat Stable Enterotoxin (HSEnt), and Hemolysin E (HlyE) (Fig 1A). To more effectively model toxin effects on intact tumor tissue, we carried out a secondary screen using PDAC explant cultures derived from pancreatic tumors in KPC mice ( 3, 17 ). Of the four candidate strains tested, Theta toxin stood out for its ability to induce apoptosis, as measured by cleaved caspase 3 (CC3) staining, compared to a GFP-expressing control strain (Fig 1B, Supp Fig 1B).…”

“…Murine tumors were collected following humane euthanasia and trimmed of healthy pancreas tissue in a sterile petri dish. Tissues were processed into explant slices and cultured as previously described ( 3, 17 ). Explants were treated with murine explant media supplemented with 10% bacterial toxin lysate, replaced daily for up to 3 days.…”

“…Most patients present with advanced or metastatic disease, making systemic chemotherapies a mainstay of treatment ( 2 ). However, the biophysical properties of the PDAC tumor microenvironment can interfere with effective delivery of both cytotoxic and targeted agents ( 3, 4 ). Modern immunotherapy also remains ineffective due to immunosuppressive properties of the microenvironment ( 5 ), highlighting the need for new treatment approaches.…”

Tumor-selective effects of active RAS inhibition in pancreatic ductal adenocarcinoma

“…Explant culture sponges and optimized culture media were prepared as previously described in Hasselluhn and Decker-Farrell et al 26 . Human tissue samples were obtained from de-identified patients undergoing resection surgeries, primarily pancreaticoduodenectomy (Whipple) or distal pancreatectomy, at New York-Presbyterian/Columbia University Irving Medical Center.…”

“…Multiple studies have demonstrated that various stromal cells resident in PDAC tissues can modulate the sensitivity of malignant cells to therapy 25 . To assess the potency of RMC-7977 in the context of an intact, all-human microenvironment, we employed an ex vivo human PDAC explant model 26 , comprising cultured intact slices of freshly resected human PDAC tissue from patients at New York Presbyterian Hospital/Columbia University Irving Medical Center. Immunohistochemistry performed on PDAC explants treated for 24 hours with RMC-7977 showed a concentration-dependent decrease in pERK expression with an accompanying increase in the apoptosis marker, cleaved caspase 3 (CC3), with maximal changes at 100 nM, the highest concentration tested (Fig.…”