SF-9-2 is a PD-L1/PD-1 binding inhibitor with an IC50 of 24.9 nM. It suppresses epithelial-mesenchymal transition, migration, invasion, and proliferation of SK-N-SH cells, while also inducing apoptosis and causing cell cycle arrest. SF-9-2 blocks PD-L1-induced growth of SK-N-SH cells via the MAPK signaling pathway. It restores GSK-3β activity and enhances PD-L1 degradation through the ubiquitin-proteasome pathway. In the SK-N-SH NOG mouse model, SF-9-2 inhibits tumor growth without notable toxicity. Additionally, SF-9-2 acts as an immune checkpoint inhibitor, blocking PD-L1 to restore T cell function and is applicable to neuroblastoma research.

SF-9-2 exhibits significant inhibitory activity against SK-N-SH (IC 50 = 5.9 μM) and SK-N-AS cells (IC 50 = 8.67 μM) in a concentration range of 0.5-16 μM over 48 hours, but shows less inhibition of SH-SY5Y and SK-N-BE (2) cells and low cytotoxicity to normal MRC-5 cells (IC 50 = 12.15 μM). At concentrations of 1-6 μM for 24 hours, SF-9-2 suppresses the proliferation, migration, invasion, and epithelial-mesenchymal transition (EMT) of SK-N-SH cells, and induces mitochondrial-dependent apoptosis and cell cycle arrest. It also inhibits the MAPK pathway and downregulates PD-L1 levels in SK-N-SH cells by targeting the ERK signaling pathway, at concentrations of 2.5-8 μM for 24-48 hours. SF-9-2 induces GSK-3β-mediated PD-L1 internalization and proteasomal degradation in SK-N-SH cells within a range of 2.5-5 μM for 24 hours. With concentrations from 2.25-200 nM, SF-9-2 reduces fluorescence signals in 293T cells overexpressing PD-1 and Fc-PD-L1 proteins, effectively blocking the PD-1/PD-L1 immune checkpoint. It enhances IFN-γ secretion levels in PBMCs at concentrations of 1-16 μM over 48 hours, with no significant toxicity at 1, 2, and 4 μM. Furthermore, SF-9-2 exhibits direct tumor-killing activity and T-cell-mediated cytotoxicity against SK-N-SH cells at concentrations of 1-8 μM for 48 hours.

SF-9-2 (20-40 mg/kg, administered via intraperitoneal injection, once daily for 21 days) inhibits the growth of SK-N-SH tumors in the NOG mouse model.