The therapeutic blockade of immune checkpoint has emerged as an effective treatment option for a broad range of
tumors. However, the objective
tumor response is still limited to a small number of cases and
tumor types. The full utility of
monoclonal antibody (mAb)-based treatment is hindered by several inherent limitations. Thus, there is an urgent requirement to explore alternative modalities targeting the same pathways. In the present study, two
amide analogues of
brefelamide, TPFS-201 and TPFS-202, were identified as small molecular
immune checkpoint inhibitors, as they downregulated PD-L1 expression in
tumor cells. PD-L1 was suppressed in
cancer cells treated with TPFD compounds at both
mRNA and
protein levels, as detected by reverse transcription quantitative PCR and flow cytometric analysis, respectively. Reporter assays using a PD-L1 promoter
luciferase construct confirmed the transcriptional inhibition of PD-L1 by TPFS compunds. TPFS compound-mediated PD-L1 downregulation in
cancer cells consequently restored T cell activity, as identified by the reduction of apoptosis and an increase in
interleukin-2 promoter activity in Jurkat T cells, which were co-cultured with TPFS compound-treated A549 cells. TPFS compound-mediated PD-L1 inhibition was partially abolished by the disruption of the putative transcriptional co-activator with PDZ (TAZ)/
TEA domain (TEAD)-binding motif in the PD-L1 promoter. The inhibitory effect of TPFS compounds on PD-L1 was markedly inhibited in mouse cell lines, which is consistent with previous research demonstrating that PD-L1 regulation by TAZ is not conserved in mice due to distinct promoter sequences flanking the TAZ/TEAD-binding motif. Together, the data of the current study indicated the potential utility of the
brefelamide amide analogues as small
molecule immune checkpoint inhibitors, thereby providing therapeutic alternatives, which could be used as monotherapy or in combination with mAbs-based treatment.