An endocytosis pathway initiated through neuropilin-1 and regulated by nutrient availability

Abstract

Neuropilins (NRPs) are trans-membrane receptors involved in axon guidance and vascular development. Many growth factors and other signaling molecules bind to NRPs through a C-terminal, basic sequence motif (C-end Rule or CendR motif). Peptides with this motif (CendR peptides) are taken up into cells by endocytosis. Tumor-homing CendR peptides penetrate through tumor tissue and have shown utility in enhancing drug delivery into tumors. Here we show, using RNAi screening and subsequent validation studies, that NRP1-mediated endocytosis of CendR peptides is distinct from known endocytic pathways. Ultrastructurally, CendR endocytosis resembles macropinocytosis, but is mechanistically different. We also show that nutrient-sensing networks such as mTOR signaling regulate CendR endocytosis and subsequent intercellular transport of CendR cargo, both of which are stimulated by nutrient depletion. As CendR is a bulk transport pathway, our results suggest a role for it in nutrient transport; CendR-enhanced drug delivery then makes use of this natural pathway.

Tumor-penetrating peptide enhances transcytosis of silicasome-based chemotherapy for pancreatic cancer

“…It has been demonstrated that binding of the C-terminal end of iRGD to NRP-1 can initiate a bulk transcytosis pathway that involves a novel vesicular transport mechanism (17,19,21). To the best of our knowledge, this transcytosis pathway has never been directly visualized during the transport of a therapeutic nanocarrier to the site of a tumor.…”

“…Binding of the cyclic peptide to the integrins is followed by cleavage and release of the C-terminal end of cleaved iRGD, which mediates the interaction with NRP-1 (this is also known as C-end rule). NRP-1 binding leads to the triggering of a system of vesicles that can assist drug and nanoparticle transport (17,19,21). A schematic demonstrating the mechanism by which iRGD initiates nanoparticle transcytosis is shown in Supplemental Figure 5 (19).…”

“…Another important recent advance has been the discovery of a vascular transcytosis pathway that allows nanocarrier uptake at the tumor site by a mechanism that differs from the EPR effect (17)(18)(19)(20)(21). It has been demonstrated that this pathway can be initiated by the cyclic tumor-penetrating peptide iRGD (CRGDK/RGPD/ EC), which interacts with the neuropilin-1 (NRP-1) receptor (17,19,20).…”

“…It has been demonstrated that this pathway can be initiated by the cyclic tumor-penetrating peptide iRGD (CRGDK/RGPD/ EC), which interacts with the neuropilin-1 (NRP-1) receptor (17,19,20). NRP-1 plays a key role in tumor angiogenesis and can also be engaged by VEGFs that contain an RXXR sequence, also dubbed the C-terminal rule (CendR) motif (18,21). The transcytosis pathway is not only distinct from other endocytic uptake mechanisms (e.g., caveolae, clathrin-coated pits, and macropinocytosis), but it also allows entry of small drug molecules, monoclonal antibodies (e.g., trastuzumab), and nanoparticles (e.g., Abraxane and doxorubicin liposomes) during iRGD coadministration (19,21).…”

“…NRP-1 plays a key role in tumor angiogenesis and can also be engaged by VEGFs that contain an RXXR sequence, also dubbed the C-terminal rule (CendR) motif (18,21). The transcytosis pathway is not only distinct from other endocytic uptake mechanisms (e.g., caveolae, clathrin-coated pits, and macropinocytosis), but it also allows entry of small drug molecules, monoclonal antibodies (e.g., trastuzumab), and nanoparticles (e.g., Abraxane and doxorubicin liposomes) during iRGD coadministration (19,21). Importantly, this effect is not dependent on direct conjugation of the peptide to the drug or carrier, as demonstrated by gemcitabine uptake in some (but not all) experimental PDAC models (22).…”