Glycine N‐Methylation in NGR‐Tagged Nanocarriers Prevents Isoaspartate Formation and Integrin Binding without Impairing CD13 Recognition and Tumor Homing

Abstract

NGR (asparagine-glycine-arginine) is a tumor vasculature-homing peptide motif widely used for the functionalization of drugs, nanomaterials, and imaging compounds for cancer treatment and diagnosis. Unfortunately, this motif has a strong propensity to undergo rapid deamidation. This reaction, which converts NGR into isoDGR, is associated with receptor switching from CD13 to integrins, with potentially important manufacturing and pharmacological and toxicological implications. It is found that glycine N-methylation of NGR-tagged nanocarriers completely prevents asparagine deamidation without impairing CD13 recognition. Studies in animal models have shown that the methylated NGR motif can be exploited for delivering radiolabeled compounds and nanocarriers, such as tumor necrosis factor-α-bearing nanogold and liposomal doxorubicin, to tumors with improved selectivity. These findings suggest that this NGR derivative is a stable and efficient tumor-homing ligand that can be used for delivering functional nanomaterials to tumor vasculature.

Overcoming Biological Barriers in Neuroblastoma Therapy: The Vascular Targeting Approach with Liposomal Drug Nanocarriers

“…It seems, therefore, that glycine N ‐methylation does not impair the tumor‐homing properties of the NGR peptide and increases its tumor selectivity, possibly by preventing isoDGR formation. These studies have also shown that this peptide can be used for the development of drug nanocarriers that are more stable and selective than those obtained with conventional NGR peptides, including peptide‐tagged liposomal doxorubicin and gold nanoparticles . Thus, this novel NGR derivative could represent a valid alternative to conventional NGR peptides for liposome functionalization.…”

“…Remarkably, this change prevents the deamidation reaction without impairing CD13 binding (see Figure ). Studies performed in mice‐bearing solid tumors have demonstrated that this NGR‐derivative is more selective for tumor tissues than the corresponding isoDGR and NGR peptides . In particular, although these studies showed similar accumulation of radiolabeled methylated NGR and the corresponding NGR and isoDGR peptides (cyclicCGNGRG and CisoDGRG) in tumors (fourfold, compared to blood and other tissues), in the case of the NGR and isoDGR peptides a certain degree of accumulation was observed also in other organs, such as stomach, spleen, liver, intestine, and femur.…”

“…This reaction could happen during the preparation and storage of NGR‐drug conjugates, and, potentially, also in vivo, after drug injection to animals or to patients. Indeed, using a peptide–antibody conjugate it has been recently shown that the conversion of NGR into isoDGR can significantly occur in the blood after systemic administration of the conjugate to mice . This could be a critical problem in the development, manufacturing, and storage of liposomal drugs.…”

“…Furthermore, considering the important receptor switching associated with the transition of NGR to isoDGR, the poor stability of NGR may have pharmacological and toxicological implications. This problem has been overcome in the head‐to‐tail cyclized peptide (CGNGRG) by replacing the glycine residue of NGR with N ‐methylglycine (sarcosine) . Remarkably, this change prevents the deamidation reaction without impairing CD13 binding (see Figure ).…”

Boosting Interleukin‐12 Antitumor Activity and Synergism with Immunotherapy by Targeted Delivery with isoDGR‐Tagged Nanogold

“…We have previously shown that peptides containing isoAsp‐Gly‐Arg (isoDGR), a motif that recognizes the αvβ3 integrin overexpressed in tumor vessels and on different tumor cell types, can be exploited as ligands for targeted delivery of various drugs and nanoparticles to tumors 16b,17. In particular, we have identified a head‐to‐tail‐cyclized hexapeptide, CGisoDGRG (called Iso1), which recognizes αvβ3 with high selectivity and affinity, binds tumor vessels, and works as an efficient ligand for delivering fluorescent nanoparticles or radioactive compounds to tumors . The Iso1 peptide, after chemical conjugation to human serum albumin, has been exploited for tagging gold nanoparticles loaded with tumor necrosis factor‐alpha (TNF), a cytokine endowed with antitumor activity .…”