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. In this study, we synthesized an iRGD conjugate containing monomethyl auristatin F (MMAF), a highly toxic antimitotic agent, and characterized its dual function as a tumor-specific cytotoxic agent and co-injected drug delivery scaffold.
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. 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. These challenges are particularly exacerbated in desmoplastic tumors such as pancreatic ductal adenocarcinoma (PDAC).
Materials and Methods:
Tumor-bearing mouse models are widely recognized as a valuable system that closely mimics the human tumor microenvironment. We used tumor mice because complex in vivo kinetics such as tumor-specific homing, anti-tumor efficacy, and off-target side effects of drugs cannot be precisely studied without an in vivo model despite the recent advancement in in vitro systems. All animal experiments were performed according to procedures approved by the Institutional Animal Care and Use Committee (IACUC) at Columbia University (Protocol number: AC-AABU3706).
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. 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.
