Tumor penetrating peptides for improved drug delivery

Abstract

In vivo screening of phage libraries in tumor-bearing mice has been used to identify peptides that direct phage homing to a tumor. The power of in vivo phage screening is illustrated by the recent discovery of peptides with unique tumor-penetrating properties. These peptides activate an endocytic transport pathway related to but distinct from macropinocytosis. They do so through a complex process that involves binding to a primary, tumor-specific receptor, followed by a proteolytic cleavage, and binding to a second receptor. The second receptor, neuropilin-1 (or neuropilin-2) activates the transport pathway. This trans-tissue pathway, dubbed the C-end Rule (CendR) pathway, mediates the extravasation transport through extravascular tumor tissue of payloads ranging from small molecule drugs to nanoparticles. The CendR technology provides a solution to a major problem in tumor therapy, poor penetration of drugs into tumors. Targeted delivery with tumor-penetrating peptides has been shown to specifically increase the accumulation of drugs, antibodies and nanotherapeutics in experimental tumors in vivo, and in human tumors ex vivo. Remarkably the payload does not have to be coupled to the peptide; the peptide activates a bulk transport system that sweeps along a drug present in the blood. Treatment studies in mice have shown improved anti-tumor efficacy and less damage to normal tissues with drugs ranging from traditional chemotherapeutics to antibodies, and to nanoparticle drugs.

Keywords: Cell-penetrating peptides; CendR pathway; Endocytosis; Integrins; Nanomedicine; Neuropilins; Synaphic targeting; Tumor vasculature.

Fig. 1

Schematic representation of the CendR trans-tissue transport pathway. A tumor-penetrating peptide with a cryptic CendR motif penetrates tumor tissue in a 3-step process: (i) The peptide binds to a primary receptor on tumor endothelium. In iRGD, the RGD motif recognizes the αvβ3/αvβ5 integrins; the primary receptor for the LyP-1 family of peptides is p32/gC1qR. (ii) The peptide is then cleaved by proteases to expose the cryptic CendR element, R/KXXR/K, at the C-terminus; and, (iii) the CendR element mediates binding to neuropilin-1 (NRP-1), inducing vascular and tissue permeability. See the text for details. Note that CendR effect enhances the tissue penetration of molecules (depicted here as a black dots) that are co-administered with the peptide [25], as well as of cargo coupled to the peptide [10]. The inset shows an electron microscopic image of a CendR endocytic vesicle that is budding from the cell surface into the cytoplasm and contains CendR peptide-coated gold nanoparticles (dark dots) (see [24]).

Fig. 2

Nanoparticles functionalized with the p32-binding compound #4008 home to p32-positive breast tumors. OrthotopicMCF10Ca1A breast tumors from mice injected with iron oxide NPs (nanoworms; NWs) coated with FAM-#4008 [23], FAM-LyP-1, or FAM-labeled inert control peptide. The NWs were allowed to circulate for 5 h, the mice were perfused through the heart with PBS, and tumors were collected and sectioned, Confocal images of the tumors are shown (green, NWs; red, the receptor, p32; blue, nuclei). Scale bar = 100 µm.

Fig. 3

Nanosystem treatment of glioblastomas in mouse models. Mice bearing lentiviral oncogene-induced (left) or transplantable (right) brain tumors were intravenously treated with a targeted nanosystem schematically depicted in the figure (see text for description of the system). Kaplan–Meier survival plots are shown. CGKRK is the homing peptide, and _D(KLAKLAK)₂ is the pro-apoptotic peptide. The lentiviral induced tumors were highly responsive to the nanosystem (CGKRK- _D(KLAKLAK)₂-NWs), with complete eradication of the tumors in almost all mice. The growth of the more aggressive transplantable tumors was delayed, and this effect was further enhanced when the iRGD peptide was co-administered with the nanosystem. Modified from [ Agemy, et al., 2011] with permission.