CD95L Inhibition: a novel approach to combat inflammation driven progression of pancreatic ductal adenocarcinoma
The CD95/CD95L system plays an important role in various biological processes, including cell homeostasis and immunological, neurological and oncological diseases. In addition to its well-known function in apoptotic signaling, evidence has emerged that the CD95/CD95L system is also actively involved in nonapoptotic signaling. However, the role of endogenous CD95 signaling in the progression of PDAC has not yet been studied in detail. In this study, we analyzed the effect of pharmacological CD95L neutralization on the inflammation-driven progression of pancreatic ductal adenocarcinoma (PDAC), one of the deadliest malignancies, which is associated with a high tumor relapse rate even after successful resection of the primary tumor. Previously, it was demonstrated that the activation of CD95 through its cognate ligands enhances tumor cell motility, invasiveness as well as production of proinflammatory cytokines through the activation of NFκB and MAPK signaling. Here we found that inhibition of CD95L using the recombinant fusion protein sCD95Fc reduced cell migration and the expression of proinflammatory cytokines and stem cell markers of PDAC cells. PDAC cells express both CD95 and CD95L, yet blockade of CD95L exerts only marginal effects on the in vitro proliferation of both PDAC cells and immortalized pancreatic stellate cells (RLT-PSCS). In the in vivo experiments, neutralization of CD95L significantly suppressed tumor growth by about 35% in the palliative therapeutic regime. Remarkably, in a PDAC resection model performed with PancTuI-luc cells, sCD95Fc showed a profound impact, reducing tumor growth by 50% in the adjuvant treatment group. It also reduced the number of liver and spleen metastases by 50% compared to the control group. In both cases sCD95Fc downregulated the proliferation index in the experimental tumors, as assessed by Ki67 staining. Moreover, immunohistochemical analyses of CD95L, CD95 and sCD95Fc in treated vs. untreated experimental tumors revealed (i) expression of the ligand, both weakly in the human tumor cell compartment and strongly in the murine stroma in naïve tumors, (ii) reduced expression and/or blockade of CD95L in the fibroblastoid and endothelial cells in treated tumors and (iii) a major localization of sCD95Fc in the tumor microenvironment. Using human sequence-specific qRT-PCR we further observed downregulation of inflammatory cytokines such as IL6 and IL8, particularly in the locally recurrent tumors upon treatment and also a decrease in the stem cell marker CD133. Taken together, the data suggest that blocking CD95/CD95L signaling might be a promising therapeutic strategy to repress the tumor-promoting effects in PDAC via modulation of the proinflammatory signals derived either from tumor cells or – even more pronounced - through factors secreted from the tumor microenvironment (PSCs and PCSCs) in a paracrine manner. The tolerability and the safety profile of sCD95Fc were examined in both mice and humans. sCD95Fc was generally well tolerated without any signs of toxicity. In a phase II clinical trial, sCD95Fc has shown therapeutic efficacy for the treatment of recurrent glioblastoma. Thus, our findings are further evidence in support of the clinical application of CD95L inhibitors as an emerging strategy for improving the therapeutic outcome in PDAC patients.