RESUMO
A series of novel 2-arylmethoxy-4-(2-fluoromethyl-biphenyl-3-ylmethoxy) benzylamine derivatives was designed, synthesized, and evaluated for their antitumor effects as PD-1/PD-L1 inhibitors both in vitro and in vivo. Firstly, the ability of these compounds to block the PD-1/PD-L1 immune checkpoint was assessed using the homogeneous time-resolved fluorescence (HTRF) assay. Two of the compounds can strongly block the PD-1/PD-L1 interaction, with IC50 values of less than 10 nM, notably, compound HD10 exhibited significant clinical potential by inhibiting the PD-1/PD-L1 interaction with an IC50 value of 3.1 nM. Further microscale thermophoresis (MST) analysis demonstrated that HD10 had strong interaction with PD-L1 protein. Co-crystal structure (2.7 Å) analysis of HD10 in complex with the PD-L1 protein revealed a strong affinity between the compound and the target PD-L1 dimer. This provides a solid theoretical basis for further in vitro and in vivo studies. Next, a typical cell-based experiment demonstrated that HD10 could remarkably prevent the interaction of hPD-1 293 T cells from human recombinant PD-L1 protein, effectively restoring T cell function, and promoting IFN-γ secretion in a dose-dependent manner. Moreover, HD10 was effective in suppressing tumor growth (TGI = 57.31 %) in a PD-1/PD-L1 humanized mouse model without obvious toxicity. Flow cytometry, qPCR, and immunohistochemistry data suggested that HD10 inhibits tumor growth by activating the immune system in vivo. Based on these results, it seems likely that HD10 is a promising clinical candidate that should be further investigated.
Assuntos
Antineoplásicos , Antígeno B7-H1 , Benzilaminas , Desenho de Fármacos , Ensaios de Seleção de Medicamentos Antitumorais , Receptor de Morte Celular Programada 1 , Humanos , Antineoplásicos/farmacologia , Antineoplásicos/síntese química , Antineoplásicos/química , Receptor de Morte Celular Programada 1/antagonistas & inibidores , Receptor de Morte Celular Programada 1/metabolismo , Antígeno B7-H1/antagonistas & inibidores , Antígeno B7-H1/metabolismo , Animais , Camundongos , Relação Estrutura-Atividade , Benzilaminas/farmacologia , Benzilaminas/química , Benzilaminas/síntese química , Estrutura Molecular , Relação Dose-Resposta a Droga , Proliferação de Células/efeitos dos fármacos , Inibidores de Checkpoint Imunológico/farmacologia , Inibidores de Checkpoint Imunológico/síntese química , Inibidores de Checkpoint Imunológico/química , Linhagem Celular Tumoral , Feminino , Modelos MolecularesRESUMO
The PD-1/PD-L1 pathway is considered as one of the most promising immune checkpoints in tumour immunotherapy. However, researchers are faced with the inherent limitations of antibodies, driving them to pursue PD-L1 small molecule inhibitors. Virtual screening followed by experimental validation is a proven approach to discover active compounds. In this study, we employed multistage virtual screening methods to screen multiple compound databases to predict new PD-1/PD-L1 ligands. 35 compounds were proposed by combined analysis of fitness scores, interaction pattern and MM-GBSA binding affinities. Enzymatic assay confirmed that 10 out of 35 ligands were potential PD-L1 inhibitors, with inhibitory rate higher than 50% at the concentration of 30 µM. Among them, ZDS20 was identified as the most effective inhibitor with low micromolar activity (IC50 = 3.27 µM). Altogether, ZDS20 carrying novel scaffold was identified and could serve as a lead for the development of new classes of PD-L1 inhibitors.
Assuntos
Antígeno B7-H1 , Relação Dose-Resposta a Droga , Avaliação Pré-Clínica de Medicamentos , Receptor de Morte Celular Programada 1 , Bibliotecas de Moléculas Pequenas , Antígeno B7-H1/antagonistas & inibidores , Antígeno B7-H1/metabolismo , Receptor de Morte Celular Programada 1/antagonistas & inibidores , Receptor de Morte Celular Programada 1/metabolismo , Humanos , Relação Estrutura-Atividade , Bibliotecas de Moléculas Pequenas/farmacologia , Bibliotecas de Moléculas Pequenas/química , Bibliotecas de Moléculas Pequenas/síntese química , Estrutura Molecular , Inibidores de Checkpoint Imunológico/farmacologia , Inibidores de Checkpoint Imunológico/síntese química , Inibidores de Checkpoint Imunológico/química , LigantesRESUMO
The immune checkpoint blockade represents a pivotal strategy for tumor immunotherapy. At present, various programmed cell death-1 (PD-1)/programmed cell death-ligand 1 (PD-L1) monoclonal antibodies have been successfully applied to tumor treatment. Additionally, numerous small molecule inhibitors of the PD-1/PD-L1 interaction have also been developed, with some advancing into clinical trials. Here, a novel PD-L1 proteolysis-targeting chimera (PROTAC) library was designed and synthesized utilizing the PD-L1 inhibitor BMS202 and the E3 ligand PG as foundational components. Among these, we identified a highly potent molecule PA8 for PD-L1 degradation in 4T1 cells (DC50 = 0.609 µM). Significantly, compound PA8 potentially inhibits 4T1 cell growth both in vitro and in vivo. Further mechanistic studies revealed that PA8 effectively promoted the immune activation of model mice. Thus, these results suggest that PA8 could be a novel strategy for cancer immunotherapy in the 4T1 tumor model. Although PA8 exhibits weaker degradation activity in some human cancer cells, it still provides a certain basis for further research on PD-L1 PROTAC.
Assuntos
Antineoplásicos , Antígeno B7-H1 , Neoplasias da Mama , Proteólise , Proteólise/efeitos dos fármacos , Animais , Antígeno B7-H1/metabolismo , Antígeno B7-H1/antagonistas & inibidores , Humanos , Camundongos , Feminino , Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/patologia , Neoplasias da Mama/metabolismo , Antineoplásicos/farmacologia , Antineoplásicos/química , Antineoplásicos/síntese química , Antineoplásicos/uso terapêutico , Linhagem Celular Tumoral , Camundongos Endogâmicos BALB C , Proliferação de Células/efeitos dos fármacos , Descoberta de Drogas , Inibidores de Checkpoint Imunológico/farmacologia , Inibidores de Checkpoint Imunológico/uso terapêutico , Inibidores de Checkpoint Imunológico/química , Inibidores de Checkpoint Imunológico/síntese química , Acetamidas , PiridinasRESUMO
Based on the close relationship between programmed death protein ligand 1 (PD-L1) and epidermal growth factor receptor (EGFR) in glioblastoma (GBM), we designed and synthesized a series of small molecules as potential dual inhibitors of EGFR and PD-L1. Among them, compound EP26 exhibited the highest inhibitory activity against EGFR (IC50 = 37.5 nM) and PD-1/PD-L1 interaction (IC50 = 1.77 µM). In addition, EP26 displayed superior in vitro antiproliferative activities and in vitro immunomodulatory effects by promoting U87MG cell death in a U87MG/Jurkat cell coculture model. Furthermore, EP26 possessed favorable pharmacokinetic properties (F = 22%) and inhibited tumor growth (TGI = 92.0%) in a GBM mouse model more effectively than Gefitinib (77.2%) and NP19 (82.8%). Moreover, EP26 increased CD4+ cells and CD8+ cells in tumor microenvironment. Collectively, these results suggest that EP26 represents the first small-molecule-based PD-L1/EGFR dual inhibitor deserving further investigation as an immunomodulating agent for cancer treatment.
Assuntos
Antineoplásicos , Antígeno B7-H1 , Receptores ErbB , Glioblastoma , Animais , Camundongos , Antineoplásicos/farmacologia , Antineoplásicos/química , Antineoplásicos/uso terapêutico , Antineoplásicos/síntese química , Antígeno B7-H1/antagonistas & inibidores , Antígeno B7-H1/metabolismo , Neoplasias Encefálicas/tratamento farmacológico , Neoplasias Encefálicas/patologia , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Descoberta de Drogas , Receptores ErbB/antagonistas & inibidores , Receptores ErbB/metabolismo , Glioblastoma/tratamento farmacológico , Glioblastoma/patologia , Inibidores de Checkpoint Imunológico/farmacologia , Inibidores de Checkpoint Imunológico/química , Inibidores de Checkpoint Imunológico/síntese química , Inibidores de Checkpoint Imunológico/uso terapêutico , Inibidores de Checkpoint Imunológico/farmacocinética , Imunoterapia/métodos , Bibliotecas de Moléculas Pequenas/química , Bibliotecas de Moléculas Pequenas/farmacologia , Bibliotecas de Moléculas Pequenas/síntese química , Relação Estrutura-AtividadeRESUMO
In this work, we report 14 novel quinazoline derivatives as immune checkpoint inhibitors, IDO1 and PD-L1. The antitumor screening of synthesized compounds on ovarian cancer cells indicated that compound V-d and V-l showed the most activity with IC50 values of about 5 µM. Intriguingly, compound V-d emerges as a stand out, triggering cell death through caspase-dependent and caspase-independent manners. More importantly, V-d presents its ability to hinder tumor sphere formation and re-sensitized cisplatin-resistant A2780 cells to cisplatin treatment. These findings suggest that compound V-d emerges as a promising lead candidate for the future development of immuno anticancer agents.
Assuntos
Antineoplásicos , Desenho de Fármacos , Ensaios de Seleção de Medicamentos Antitumorais , Inibidores de Checkpoint Imunológico , Quinazolinas , Humanos , Quinazolinas/farmacologia , Quinazolinas/química , Quinazolinas/síntese química , Relação Estrutura-Atividade , Inibidores de Checkpoint Imunológico/farmacologia , Inibidores de Checkpoint Imunológico/síntese química , Inibidores de Checkpoint Imunológico/química , Antineoplásicos/farmacologia , Antineoplásicos/síntese química , Antineoplásicos/química , Estrutura Molecular , Linhagem Celular Tumoral , Relação Dose-Resposta a Droga , Proliferação de Células/efeitos dos fármacos , Indolamina-Pirrol 2,3,-Dioxigenase/antagonistas & inibidores , Indolamina-Pirrol 2,3,-Dioxigenase/metabolismo , Antígeno B7-H1/antagonistas & inibidores , Antígeno B7-H1/metabolismoRESUMO
In this work, a series of bifunctional PD-L1/CD73 (cluster of differentiation 73) small-molecule inhibitors were designed and synthesized. Among them, CC-5 showed the strongest PD-L1 inhibitory effects with an IC50 of 6 nM and potent anti-CD73 activity with an IC50 of 0.773 µM. The high PD-L1/CD73 inhibitory activity of CC-5 was further confirmed by SPR assays with KD of 182 nM for human PD-L1 and 101 nM for CD73, respectively. Importantly, CC-5 significantly suppressed tumor growth in a CT26 and B16-F10 tumor model with TGI of 64.3% and 39.6%, respectively. Immunohistochemical (IHC) and flow cytometry analysis of tumor-infiltrating lymphocytes (TILs) indicated that CC-5 exerted anticancer effects via activating the tumor immune microenvironment. Collectively, CC-5 represents the first dual PD-L1/CD73 inhibitor worthy of further research as a bifunctional immunotherapeutic agent.
Assuntos
5'-Nucleotidase , Antígeno B7-H1 , Imunoterapia , 5'-Nucleotidase/antagonistas & inibidores , 5'-Nucleotidase/metabolismo , Humanos , Antígeno B7-H1/antagonistas & inibidores , Antígeno B7-H1/metabolismo , Animais , Camundongos , Imunoterapia/métodos , Linhagem Celular Tumoral , Microambiente Tumoral/efeitos dos fármacos , Bibliotecas de Moléculas Pequenas/química , Bibliotecas de Moléculas Pequenas/farmacologia , Bibliotecas de Moléculas Pequenas/síntese química , Descoberta de Drogas , Antineoplásicos/farmacologia , Antineoplásicos/química , Antineoplásicos/síntese química , Antineoplásicos/uso terapêutico , Proteínas Ligadas por GPI/antagonistas & inibidores , Proteínas Ligadas por GPI/metabolismo , Camundongos Endogâmicos C57BL , Proliferação de Células/efeitos dos fármacos , Relação Estrutura-Atividade , Camundongos Endogâmicos BALB C , Inibidores de Checkpoint Imunológico/farmacologia , Inibidores de Checkpoint Imunológico/química , Inibidores de Checkpoint Imunológico/uso terapêutico , Inibidores de Checkpoint Imunológico/síntese químicaRESUMO
PD-1/PD-L1 pathway blockade is a promising immunotherapy for the treatment of cancer. In this manuscript, a series of triaryl compounds containing ester chains were designed and synthesized based on the pharmacophore studies of the lead BMS-1. After several SAR iterations, 22 showed the best biochemical activity binding to hPD-L1 with an IC50 of 1.21 nM in HTRF assay, and a KD value of 5.068 nM in SPR analysis. Cell-based experiments showed that 22 effectively promoted A549 cell death by restoring T-cell immune function. 22 showed significant in vivo antitumor activity in a 4T1 mouse model without obvious toxicity, with a TGI rate of 67.8 % (20 mg/kg, ip). Immunohistochemistry data indicated that 22 activates the immune activity in tumors. These results suggest that 22 is a promising compound for further development of PD-1/PD-L1 inhibitor for cancer therapy.
Assuntos
Antineoplásicos , Antígeno B7-H1 , Ésteres , Receptor de Morte Celular Programada 1 , Humanos , Animais , Antígeno B7-H1/antagonistas & inibidores , Antígeno B7-H1/metabolismo , Receptor de Morte Celular Programada 1/antagonistas & inibidores , Receptor de Morte Celular Programada 1/metabolismo , Camundongos , Relação Estrutura-Atividade , Antineoplásicos/farmacologia , Antineoplásicos/química , Antineoplásicos/síntese química , Estrutura Molecular , Ésteres/química , Ésteres/farmacologia , Ésteres/síntese química , Ensaios de Seleção de Medicamentos Antitumorais , Relação Dose-Resposta a Droga , Proliferação de Células/efeitos dos fármacos , Camundongos Endogâmicos BALB C , Feminino , Neoplasias Experimentais/tratamento farmacológico , Neoplasias Experimentais/patologia , Inibidores de Checkpoint Imunológico/farmacologia , Inibidores de Checkpoint Imunológico/química , Inibidores de Checkpoint Imunológico/síntese químicaRESUMO
In the context of peptide drug development, glycosylation plays a pivotal role. Accordingly, L-type peptides were synthesized predicated upon the PD-1/PD-L1 blocker DPPA-1. Subsequent glycosylation resulted in the production of two distinct glycopeptides, D-glu-LPPA-1 and D-gal-LPPA-1, by using D-glucose (D-glu) and D-galactose (D-gal), respectively, during glycosylation. Both glycopeptides significantly inhibited the interaction between PD-1 and PD-L1, and the measured half maximal inhibitory concentrations (IC50s) were 75.5 µM and 101.9 µM for D-glu-LPPA-1 and D-gal-LPPA-1, respectively. Furthermore, D-gal-LPPA-1 displayed a pronounced ability to restore T-cell functionality. In an MC38 tumor-bearing mouse model, D-gal-LPPA-1 demonstrated a significant inhibitory effect. Notably, D-gal-LPPA-1 substantially augmented the abundance and functionality of CD8+ T cells in the tumor microenvironment. Additionally, in the lymph nodes and spleens, D-gal-LPPA-1 significantly increased the proportion of CD8+ T cells secreting interferon-gamma (IFN-γ). These strong findings position D-gal-LPPA-1 as a potent enhancer of the antitumor immune response in MC38 tumor-bearing mice, underscoring its potential as a formidable PD-1/PD-L1 blocking agent.