Abstract 4468: Modified 5-FU immunoliposome targeting EGFR-expressing pancreatic tumors: <i>In vitro</i> and <i>in vivo</i> evaluation

Abstract Purpose: Epidermal growth factor receptor (EGFR) is vital in tumor growth, invasion, and metastasis. Many pancreatic tumors express EGFR, making it important in targeted therapy. XYZ-I-73, a novel 5-Fluorouracil (5-FU) analog formed from conjugating lauroyl chloride to 5-fluorocytosine from our previous study, has shown cytotoxicity against pancreatic cancer (PCa) cells. The study aimed to develop an immunoliposome by conjugating pegylated XYZ-I-73 liposomal nanoparticles to EGFR monoclonal antibody and determining in vitro and in vivo anticancer effects in PCa. Methods: EGFR expression in PCa cells was determined via Western blot analysis. XYZ-I-73 liposomes were prepared from dipalmitoyl phosphatidylcholine (DPPC),1,2-Distearoyl-sn-glycero-3-phosphoethanolamine-Polyethylene glycol (DSPE-PEG 2000), and Labrasol as surfactant using a thin-film hydration method. We conjugated EGFR monoclonal antibody to XYZ-I-73 PEGylated liposomes (XYZ-I-73LnP) to produce an antibody-conjugated PEGylated liposome (Ab-XYZ-I-73LnP). Immunoliposomes conjugation was confirmed and characterized for size, entrapment, and stability. The in vitro cytotoxicity and apoptotic effects were determined in MIA PaCa-2 cells and PANC-1 cells. Pharmacokinetics and biodistribution studies were conducted on KPC mice. Results: Characterization of XYZ-I-73LnP showed a mean particle size of 105±2.1nm, PDI (0.24), zeta potential of -34.3±0.9mV, and entrapment efficiency (EE) of 84.3±3.8%. Ab-XYZ-I-73LnP mean particle size was 143nm±2.3, PDI (0.37), and zeta potential -46.2±1.3mV. The coupling efficiency and conjugation of the antibody to the liposomes were confirmed by Bicinchoninic Acid Assay (BCA) and Fourier Transform Infra-Red (FTIR). The in vitro cytotoxicity of Ab-XYZ-I-73LnP treated MIA PaCa-2 cells (IC50 (2D) = 2.5±0.9μM and IC50 (3D) = 8.1±1.1μM) was significantly higher than 5-FU-treated cells (IC50 (2D) = 7.4±1.3μM; IC50 (3D) = 26.7±1.1 μM). Ab-XYZ-I-73LnP inhibition in PANC-1 culture was significantly higher (IC50 (2D) = 2.9±1.1 μM; IC50 (3D) = 11.6±1.9 μM) than 5-FU-treated culture (IC50 (2D) = 26.7±1.1 μM; IC50 (3D) = 37.1±0.9 μM). Ab-XYZ-I-73LnP showed a remarkable apoptotic effect as concentration increased (12.5, 25, and 50μM) in MIA PaCa-2 and PANC-1 3D spheroids compared to 5-FU. MIA PaCa-2-treated Ab-XYZ-I-73LnP showed a significantly reduced expression of EGFR than 5FU. The pharmacokinetic profile of Ab-XYZ-I-73LnP showed a longer half-life than 5-FU (t1/2 = 1.62±0.03h vs 0.49±0.01h, p&amp;lt; 0.001). The area under the curve (AUC) of Ab-XYZ-I-73LnP was 2.5-fold higher than 5-FU. Ab-XYZ-I-73LnP demonstrated a significantly higher tumor accumulation than 5-FU. Conclusion: The study demonstrated that modified 5-FU immunoliposome nanoparticles may enhance the therapeutic activity of 5-FU in treating PCa. Citation Format: Esther Frimpong, Raviteja Bulusu, Joy Okoro, Xue Zhu, Renee Reams, Bo Han, Saunjoo Yoon, Edward Agyare. Modified 5-FU immunoliposome targeting EGFR-expressing pancreatic tumors: In vitro and in vivo evaluation [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 4468.