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

Purpose: 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) F_ab/anti-CD3_scF_v 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 pharmacoki-netics, anti-tumor efficacy studies, and T-cell tracking studies were conducted to analyze the advantages of nsTPPgels included D₂ gel 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 D₂ gel with a desirable gelation temperature window, which have the potential to be used for antibody drug delivery in several biomedical applications.