Chemically Modified Peptides Targeting the PDZ Domain of GIPC as a Therapeutic Approach for Cancer

Abstract

GIPC (GAIP-interacting protein, C terminus) represents a new target class for the discovery of chemotherapeutics. While many of the current generation of anticancer agents function by directly binding to intracellular kinases or cell surface receptors, the disruption of cytosolic protein-protein interactions mediated by non-enzymatic domains is an underdeveloped avenue for inhibiting cancer growth. One such example is the PDZ domain of GIPC. Previously we developed a molecular probe, the cell permeable octapeptide CR1023 (N-myristoyl-PSQSSSEA), which diminished proliferation of pancreatic cancer cells. We have expanded upon that discovery using a chemical modification approach, and here report a series of cell permeable, side chain-modified lipopeptides that target the GIPC PDZ domain in vitro and in vivo. These peptides exhibit significant activity against pancreatic and breast cancers, both in vitro and in animal models. CR1166 (N-myristoyl-PSQSK(εN-4-bromobenzoyl)SK(εN-4-bromobenzoyl)A), bearing two halogenated aromatic units on alternate side chains, was found to be the most active compound, with pronounced down-regulation of EGFR/1GF-1R expression. We hypothesize that these organic acid-modified residues extend the productive reach of the peptide beyond the canonical binding pocket, which defines the limit of accessibility for the native proteinogenic sequences that the PDZ domain has evolved to recognize. Cell permeability is achieved with N-terminal lipidation using myristate, rather than a larger CPP (cell-penetrating peptide) sequence. This, in conjunction with optimization of targeting through side chain modification, has yielded an approach that will allow the discovery and development of next-generation cellular probes for GIPC PDZ as well as other PDZ domains.

Synectin promotes fibrogenesis by regulating PDGFR isoforms through distinct mechanisms

“…Although synectin is a known binding partner of NRP-1 (9), the effects of synectin on fibrosis may be broader, given that synectin -unlike NRP-1 -regulates both PDG-FR isoforms. While potential human therapies targeting both molecules are evolving, NRP-1 targeting has focused on a neutralizing antibody that can target the extracellular domain of the protein (6), while synectin targeting has focused on small molecules that can be internalized by target cells and act as a neutralizing peptide (42). Given the expression of synectin in multiple liver cell types, it follows that synectin may regulate diverse functions in liver beyond fibrosis; therapies may require HSC selectivity, and caution would be needed for possible off-target effects.…”

Inhibition of N-Methyl-d-aspartate-induced Retinal Neuronal Death by Polyarginine Peptides Is Linked to the Attenuation of Stress-induced Hyperpolarization of the Inner Mitochondrial Membrane Potential

“…Study-Peptides were synthesized using N-(9-fluorenyl)methoxycarbonyl (Fmoc)-solid phase peptide synthesis protocols employing microwave protocols (27). Some were manually prepared using PS3 automated peptide synthesizer (Ranin Instrument Co., Inc.) at room temperature.…”

Inhibition of Endoglin–GIPC Interaction Inhibits Pancreatic Cancer Cell Growth

“…In order to promote cell permeability, we adopted and expanded upon two strategies we had employed previously (35). For the first, we synthesized derivatives in which the amino terminus of the peptide was acylated with natural fatty acids of increasing carbon length (lauric, myristic, palmitic and stearic), as well as simple acetate, to serve as a form of control for the effect of lipidation (Table 1).…”

“…The second influence from our earlier work (35) inspired us to chemically modify the side chain in each of two different positions within the endoglin peptide sequence. Sequentially substituting Lys for the endogenous Met and Ser residues at the “−1” and “−3” positions, respectively, we acylated the side chain amine with benzoate.…”

“…Therefore, it is quite understandable that disruption of this GIPC PDZ-specific interaction with selective inhibitors should inhibit protein-protein interactions within cellular signaling pathways that are required for cancer growth. Previous studies from our group had utilized peptide-based inhibitors for this purpose (30, 34, 35). …”