Abstract A017: FAP-Activated Prodrugs Target Cancer Stem Cells in Esophageal Adenocarcinoma
Esophageal adenocarcinoma (EAC) is a rare and lethal malignancy that evolves through a stepwise mucosal progression — from an intestinal metaplasia known as “Barrett's esophagus (BE)” to low-grade dysplasia (LGD), high-grade dysplasia (HGD), and finally invasive carcinoma — each stage harboring rare clonogenic stem cells (approximately 1:1,000 cells) that drive disease evolution, therapeutic resistance, and recurrence. These lesion-specific cancer stem cells (CSCs) display intralesional genomic heterogeneity and enormous proliferative potential, yet remain inadequately targeted by current systemic regimens, underscoring the unmet need in this rare malignancy. We previously established that TP101 — a synthetic lethal combination of the bivalent squaramide-linked IAP antagonist 7532N and ponatinib — selectively eradicates BE, LGD, HGD, and EAC stem cells at low-nanomolar potency while sparing normal esophageal stem cells. Mechanistically, 7532N drives rapid degradation of cIAP1/2 and, together with ponatinib, triggers selective caspase-3 activation in malignant cells; in vivo, this combination produced near-complete regression of EAC xenografts and depletion of FAP-positive cancer-associated fibroblasts (CAFs) (Xian et al., 2025). However, ponatinib's dose-limiting vascular, cardiac, and hepatic toxicities constrain its systemic use in solid tumors. To address this, we engineered FAP-activated prodrugs of ponatinib that exploit the selective, high-level expression of fibroblast activation protein alpha (FAP) in EAC stromal CAFs to focus drug activation to the tumor microenvironment. FAP-dependent cleavage of prodrugs was confirmed in biochemical and cell-based assays, with minimal activation under FAP-negative conditions. In KCL-22 (ABL-activated CML line) xenograft models, the ponatinib prodrugs achieved antitumor efficacy comparable to parental ponatinib, while prodrug-treated animals exhibited minimal body-weight change versus marked weight loss (greater than 10%) observed with ponatinib at equivalent efficacious doses — indicating a meaningfully improved systemic tolerability profile. Bystander killing of adjacent malignant tissue including CSCs, pericytes, and endothelial cells was confirmed as the mechanistic basis for tumor eradication beyond the FAP-expressing stromal compartment. Broad anti-tumor activity of the synthetic lethal was further demonstrated across FAP-high solid tumor models including stomach, lung, and pancreatic cancers. These data demonstrate that FAP-activated ponatinib prodrugs achieve selective, TME-compartmentalized delivery with substantially reduced systemic exposure and, when combined with the IAP antagonist 7532N as the TP101 regimen, provide a precision, stem cell-selective synthetic lethal strategy for this rare and aggressive malignancy. Frank McKeon, Wa Xian, Ruiwen Zhang, Jaffer Ajani, Christopher Crum, Souneek Chakraborty, Yen-hsiang Huang, Guanglin Zhang, Guangyan Zhou, Longyue Liu, Melina Khorrami, Huiqin Wang, Thomas Benton. FAP-Activated Prodrugs Target Cancer Stem Cells in Esophageal Adenocarcinoma [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Breaking Barriers in the Fight against Rare Cancers; 2026 Jul 18-20; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2026;86(14_Suppl):Abstract nr A017.