Nonshrinkable Thermosensitive Hydrogels Combined with Bispecific Anti-PSMA/CD3 T-Cell Engager for Effective Against Tumors in Mice Model

Pu-Sheng Wei,1,* Po-Yu Chou,1,* Hao-Yi Hsu,2 Michael Chen,3 Yi-Jou Chen,3 Tung-Han Tsai,1 Bang-Yu Wen,4 Ming-Thau Sheu,1 Kuo-Hsiang Chuang,3,5– 10 Hong-Liang Lin2,5 1School of Pharmacy, College of Pharmacy, Taipei Medical University, Taipei, Taiwan; 2School of Pharmacy, College of Pharmacy, Kaohsiung Medical University, Kaohsiung, Taiwan; 3Graduate Institute of Pharmacognosy, Taipei Medical University, Taipei, Taiwan; 4Department of Biotechnology and Pharmaceutical Technology, Yuanpei University of Medical Technology, Hsinchu, Taiwan; 5Drug Development and Value Creation Research Center, Kaohsiung Medical University, Kaohsiung, Taiwan; 6Program in Clinical Genomics and Proteomics, Taipei Medical University, Taipei, Taiwan; 7Ph.D. Program in Clinical Drug Development of Herbal Medicine, Taipei Medical University, Taipei, Taiwan; 8Traditional Herbal Medicine Research Center, Taipei Medical University Hospital, Taipei, Taiwan; 9Ph.D Program in Biotechnology Research and Development, Taipei Medical University, Taipei, Taiwan; 10The Ph.D. Program of Translational Medicine, Taipei Medical University, Taipei, Taiwan*These authors contributed equally to this workCorrespondence: Kuo-Hsiang Chuang; Hong-Liang Lin, Email [email protected]; [email protected]: CD3-based Bispecific T-cell engagers (BiTEs) are effective for solid tumors due to their tumor specificity and tissue penetration, but they face challenges like short half-lives and narrow therapeutic windows. Innovative delivery systems, like thermosensitive hydrogels, show the potential to enhance stability, sustained release, and therapeutic efficacy.Methods: We developed PEGylated PLGA (PEG–PLGA) thermosensitive hydrogels with a nonshrinkable property (nsTPPgels) for effective controlled release and loaded them with bispecific anti-prostate surface membrane antigen (PSMA) Fab/anti-CD3 scFv T-cell engager (BiPTE) to form in situ drug deposits with a sustained-release profile after subcutaneous injection. Each group of hydrogels was first tested for differences in properties through rheological and in vitro drug release profiles. Meanwhile, in vivo pharmacokinetics, anti-tumor efficacy studies, and T-cell tracking studies were conducted to analyze the advantages of nsTPPgels included D2gel and DTgels.Results: The cytotoxicity of BiPTE against PSMA-overexpressing tumor cells and the drug release functionality of nsTPPgels were validated in vitro. Rheological studies showed that both D2gel and DTgels remained in solution below 27 °C for easy injection and solidified at physiological temperatures to form localized depots for sustained BiPTE release. All nsTPPgels demonstrated a 5-day in vitro sustained release, prolonged elimination half-life, steady plasma BiPTE levels, and extended mean residence time. In an LNCaP-xenograft mouse model, tumor growth inhibition rates for BiPTE/DTgel-2, BiPTE/DTgel-2S, and BiPTE/D2gel were 74.3%, 96.1%, and 113.1%, respectively, compared to 35.6% for intravenous and 46% for subcutaneous BiPTE administration. Furthermore, all nsTPPgels effectively achieved T-cell recruitment to lymph nodes and tumor sites in tracking studies.Conclusion: In conclusion, we developed relatively convenient injectable thermosensitive D2gel with a desirable gelation temperature window, which have the potential to be used for antibody drug delivery in several biomedical applications. Keywords: Bispecific T-cell engager, thermosensitive hydrogel, nonshrinkable, in situ drug deposits, anti-PSMA, T-cell recruitment