Lysosomal Pathways and Autophagy Distinctively Control Endothelial Cell Behavior to Affect Tumor Vasculature

Abstract

Cancer cell-stromal cell crosstalk is orchestrated by a plethora of ligand-receptor interactions generating a tumor microenvironment (TME) which favors tumor growth. The high pro-angiogenic nature of the TME perpetuates the chaotic network of structurally immature, low pericyte-covered vessels characteristic of the tumor vasculature. We previously demonstrated that chloroquine (CQ) -a lysosomotropic agent used as first-generation autophagy blocker in clinical trials- induced tumor vessel normalization and reduced tumor hypoxia. CQ improved both vessel structure and maturation, whereas the conditional knockout of the crucial autophagy gene Atg5 in endothelial cells (ECs) did not, thus highlighting a potential differential role for EC-associated autophagy and the lysosomes in pathological tumor angiogenesis. However, how CQ or ATG5-deficiency in ECs affect angiogenic signals regulating EC-pericyte interface and therefore vessel maturation, remains unknown. Here, we show that in ECs CQ constrained VEGF-A-mediated VEGF receptor (VEGFR)2 phosphorylation, a driver of angiogenic signaling. In the presence of CQ we observed increased expression of the decoy receptor VEGFR1 and of a lower molecular weight form of VEGFR2, suggesting receptor cleavage. Consequently, VEGF-A-driven EC spheroid sprouting was reduced by CQ treatment. Furthermore, CQ significantly affected the transcription and secretion of platelet-derived growth factor (PDGF)-AB/BB (upregulated) and Endothelin-1 (EDN1, downregulated), both modulators of perivascular cell (PC) behavior. In contrast, silencing of ATG5 in ECs had no effect on VEGFR2 to VEGFR1 ratio nor on PDGFB and EDN1 expression. Accordingly, mice harboring B16F10 melanoma tumors treated with CQ, displayed both an increased number of αSMA+ PCs covering tumor vessels and co-expressed PDGF receptor-β, enabling PDGF ligand dependent recruitment. Moreover, upon CQ treatment the tumoral expression of angiopoietin-1 (Angpt1), which retains mural cells, and induces vessel stabilization by binding to the EC-localized cognate receptor (TIE2), was increased thus supporting the vessel normalization function of CQ. These features associated with improved tumor vasculature were not phenocopied by the specific deletion of Atg5 in ECs. In conclusion, this study further unravels endothelial cell autonomous and non-autonomous mechanisms by which CQ “normalizes” the intercellular communication in the tumor vasculature independent of autophagy.

Pulsed-wave Ultrasound Hyperthermia Enhanced Nanodrug Delivery Combined with Chloroquine Exerts Effective Antitumor Response and Postpones Recurrence

“…found that lysosomal inhibition by CQ impaired de novo nucleotide biosynthesis and depleted aspartate in pancreatic ductal adenocarcinoma 24 . Even the aforementioned vessel-normalizing ability of CQ was proposed in a recent study to be related to lysosomal dysfunction 25 .…”

Autophagy modulation altered differentiation capacity of CD146+ cells toward endothelial cells, pericytes, and cardiomyocytes

“…***p < 0.001 groups showed the completion of autophagic response after the exposure to the Met, while the current results showed contradictory P62 changes after autophagy modulation in progenitor cells oriented toward endothelial lineage. One reason for this discrepancy would be that the treatment of cells with lineage-specific factors could alter or blunt the normal function of the autophagic signaling pathway effector [30]. Therefore, one could hypothesize that the treatment of human bone marrow CD146 + cells could alter cellular levels of main autophagic regulators such as Beclin-1, P62, and LC3II/I ratio.…”

Vav1 is Essential for HIF-1α Activation via a Lysosomal VEGFR1-Mediated Degradation Mechanism in Endothelial Cells

“…In this study, we found that Vav1 is transported to lysosomes by another carrier protein, VEGFR1, in endothelial cells. VEGFR1 is a receptor tyrosine kinase molecule that is known to be degraded by lysosomal proteolysis [27]. Vav1 binds at the binding motif of Y-K-E-P to VEGFR1, and knockdown of VEGFR1 inhibits Vav1 degradation, and conversely activation of VEGFR1 increases Vav1 degradation.…”