A cytotoxic peptide-drug conjugate for tumor-specific delivery of co-injected molecules
Norio Miyamura, Chisato M. Yamazaki, Yasuaki Anami, Kyoji Tsuchikama, Kazuki N. Sugahara
Abstract
An ideal cancer therapy enhances anti-tumor effects while minimizing side effects. iRGD, a non-cytotoxic peptide that activates a tumor-specific molecular transport machinery, promotes the penetration of co-injected drugs into tumor tissues. Clinical trials have demonstrated its potential as a tumor-specific delivery scaffold and potentiator of anti-cancer agents.
Introduction
Cancer therapy remains challenged by suboptimal anti-cancer effects and significant off-target toxicity. One major cause is poor drug penetration into tumors. Solid tumors often have high interstitial fluid pressure, preventing systemic drugs from extravasating and spreading within the tumor tissue [1,2]. As a result, only 1–3% of the injected dose enters the tumor leaving a major proportion of the dose available for off-target toxicity [3]. These challenges are particularly exacerbated in desmoplastic tumors such as pancreatic ductal adenocarcinoma (PDAC).
Materials and method
Synthesis of iRGD-conjugates
All information regarding synthesis procedures and characterization data for these compounds are provided in the Supporting Information.
Results
Design of iRGD-drug conjugates
Since iRGD contains multiple reactive side chains that are essential for its function (e.g., lysine, aspartic acid), covalent linking of an anticancer drug to iRGD must avoid altering these critical moieties. To this end, we designed iRGD drug conjugates and synthetic schemes based on the orthogonal alkyne–azide click reaction, as illustrated in Fig 1 and SI. The conjugates consist of the tumor-penetrating iRGD peptide (cyclic -CRGDKGPDC-), a glutamic acid–valine–citrulline (EVCit) tripeptide linker [15–17], and monomethyl auristatin F (MMAF, Fig 1A and 1B).
Discussion
The aim of this study was to characterize iRGD-MMAF and its capacity as a tumor-specific drug delivery scaffold. We chemically conjugated an iRGD peptide to a highly cytotoxic molecule MMAF using an EVC linker. The EVC linker facilitates effective in vivo drug delivery by providing stability in the circulation and high susceptibility to cathepsin B within tumors [17]. The iRGD-MMAF conjugate showed αv integrin-dependent internalization and cytotoxicity in cultured cells indicating proper iRGD functions.
Acknowledgments
We thank Yoko Odagiri for assisting the experiments
Citation: Miyamura N, Yamazaki CM, Anami Y, Tsuchikama K, Sugahara KN (2025) A cytotoxic peptide-drug conjugate for tumor-specific delivery of co-injected molecules. PLoS One 20(9): e0331564. https://doi.org/10.1371/journal.pone.0331564
Editor: Mohamed Abdelkarim, Faculty of Medicine of Tunis, TUNISIA
Received: May 28, 2025; Accepted: August 18, 2025; Published: September 2, 2025
Copyright: © 2025 Miyamura et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
Data Availability: All relevant data are within the manuscript and its Supporting Information files.
Funding: This work was supported by grants R01CA167174 (K.N.S.) from the National Cancer Institute of NIH, the Translational Research Grant from the Pancreatic Cancer Action Network (K.N.S.), Idea Award with Special Focus from the Department of Defense (K.N.S.). “The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.”
Competing interests: “K.N.S. is a co-founder of Cend Therapeutics, Inc (now Lisata Therapeutics, Inc), and has ownership interest in the company. K.N.S. is an inventor on a patent application relevant to this article (WO2009126349). C.M.Y., Y.A. and K.T. are named inventors on the patent applications (WO2018218004A1, US11629122B2, EP3630189A4 and WO2023122587A3) relating to the linker technologies described in this article. K.T. is a co-founder of and holds equity in CrossBridge Bio. This does not alter our adherence to PLOS ONE policies on sharing data and materials. N.M. has no competing interests to declare.” WO2009126349: Methods and compositions related to internalizing RGD peptides WO2018218004A1: Linkers for antibody drug conjugates US11629122B2: Linkers for antibody drug conjugates EP3630189A4: Linker for antibody medicinal conjugates WO2023122587A3: Tripeptide linkers and methods of use thereof.
