Abstract A125: Development of TROP2-targeted cytotoxic peptides and drug conjugates for treatment of triple negative breast cancer

Abstract Trophoblast cell-surface antigen 2(TROP2) is a transmembrane glycoprotein overexpressed in various epithelial malignancies, including cholangiocarcinoma and triple-negative breast cancer (TNBC). Its elevated expression is associated with tumor aggressiveness, metastasis, and poor clinical outcomes. While TROP2-targeting antibody-drug conjugates (ADCs) such as sacituzumab govitecan have shown clinical efficacy, peptide-drug conjugates (PDCs) offer advantages including enhanced tumor penetration, reduced immunogenicity, and synthetic versatility. In this study, we have exploited a cyclic form of TROP2-binding peptide linked with a pro-apoptotic D-form klaklakklaklak (kla) peptide or monomethyl auristatin E (MMAE) via a fatty acid linker. A CX7C peptide library displayed on T7 phage was screened against TROP2-expressing HEK 293T cells, leading to identification of a candidate peptide (named TROP2pep). Internalization was assessed in TROP2-high (HCC 1806) and TROP2-low (MDA-MB-231) TNBC cells using pH-sensitive dye labeled peptide. Peptide binding under non-internalizing conditions (4°C) was visualized with FITC-labeled peptides. TROP2pep showed higher binding to recombinant TROP2 protein than to BSA and to Trop2-high HCC1806 cells than to TROP2-low MDA-MB-231 cells. Based on this lead peptide, alanine scanning was performed to generate and optimized mutant cyclic peptide with improved internalization and selectivity. The mutant demonstrated higher binding and more efficient internalization in HCC1806 cells with minimal signal in MDA-MB-231 and BEAS-2B normal cells. Conjugation with the pro-apoptotic kla peptide induced selective cytotoxicity toward HCC1806 (IC50=110.7 ± 2.7 μM), with > 18-fold reduced activity in MDA-MB-231. The MMAE-fatty acid-conjugated PDC based on the mutant showed potent cytotoxicity in HCC1806 (IC50=25.6 ± 5.4 nM) and negligible toxicity in TROP2-low or normal cells, highlighting its therapeutic selectivity. We developed a novel TROP2-targeted PDCs that selectively bind to and internalize into TROP2-overexpressing TNBC cells, inducing potent cytotoxic effects in vitro. Given the clinical relevance of TROP2 in aggressive tumors, this platform holds promise as a next-generation targeted therapy. Ongoing studies include in vivo validation, organoid evaluation, and application to different types of TROP2-high tumors through multidisciplinary collaboration. Citation Format: Hyeongyu Park, Uttapol Permpoon, Hyunji Kim, Hyejin Ahn, Sang Jeon Chung, Byungheon Lee. Development of TROP2-targeted cytotoxic peptides and drug conjugates for treatment of triple negative breast cancer [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference on Molecular Targets and Cancer Therapeutics; 2025 Oct 22-26; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2025;24(10 Suppl):Abstract nr A125.