Abstract Triple-negative breast cancer (TNBC) is an aggressive subtype lacking molecular targets and resistant to conventional therapy. Small GTPases such as RAS, RHO, and RAC regulate proliferation and cytoskeletal organization through prenylation and subsequent methylation-demethylation cycles. The methylation of prenylated cysteine residues by prenylated protein methyltransferase (PPMTase) and demethylation by polyisoprenylated methylated protein methyl esterase (PMPMEase) maintains GTPase turnover, activation and membrane localization. Overexpression of PMPMEase in cancers accelerates this cycle, disrupting the balance between methylated and demethylated states and sustaining aberrant MAPK and PI3K/AKT signaling that drives malignant growth. To selectively disrupt this process, polyisoprenylated phosphonyl ester inhibitors (PPEIs) were designed as irreversible, mechanism-based inhibitors that exploits PMPMEase’s substrate recognition. Each PPEI incorporates a farnesyl tail that mimics the prenylated cysteine terminus of native substrates, promoting high-affinity binding and covalent modification of the catalytic serine. PPEIs were synthesized from farnesylated cysteine precursors and characterized by NMR and ion-trap mass spectrometry. Treatment of MDA-MB-231 and MDA-MB-468 cells with PPEIs produced dose-dependent reductions in viability and actin cytoskeletal integrity. NSL-AB-01 exhibited EC50 values of 4.1 µM (MDA-MB-231) and 3.3 µM (MDA-MB-468), while NSL-AJ-01 showed higher potency (3.2 µM and 2.7 µM, respectively). Actin-staining analysis revealed ∼90% reduction in mean cell area, indicating impaired Rho-family GTPase signaling. Western blotting showed decreased p-AKT (-25% in MDA-MB-231; -50% in MDA-MB-468) and increased p-ERK (≈2.8-fold in MDA-MB-231; +70% in MDA-MB-468). To determine whether PPEI-induced cytotoxicity was associated with oxidative stress, intracellular ROS generation was quantified using DCFDA fluorescence. NSL-AJ-01 and NSL-AB-01 induced strong, concentration-dependent ROS production in TNBC cells. NSL-AJ-01 increased ROS levels approximately 25-fold in MDA-MB-231 and 24-fold in MDA-MB-468 at 3 µM (p 0.001). NSL-AB-01 induced even greater oxidative responses, causing an approximately 32-fold increase of ROS in MDA-MB-231 and 30-fold increase in MDA-MB-468 at 3 µM (p 0.0001). Together, these findings demonstrate that PPEIs induce oxidative damage to breast cancer cells, suppress pro-survival AKT signaling, and activate ERK-mediated stress pathways, driving cytoskeletal disassembly and reduced TNBC cell viability. Citation Format: Joshua Kofi Ablordeppey, Kweku Ofosu-Asante, Amarender Burra, Jahnissi Odoom, Desmond Kwakye, Nazarius S. Lamango. Synthesis, characterization, and biological evaluation of polyisoprenylated phosphonyl ester inhibitors on triple-negative breast cancer abstract. In: Proceedings of the American Association for Cancer Research Annual Meeting 2026; Part 1 (Regular Abstracts); 2026 Apr 17-22; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2026;86(7 Suppl):Abstract nr 988.
Ablordeppey et al. (Fri,) studied this question.