Tumor immunotherapy refers to the activation of the immune system in the human body to kill cancer cells, and its unique therapeutic means has been widely noticed in the field of cancer treatment [1]. Immunotherapy is mainly divided into three categories, Chimeric Antigen Receptor T-cell (CAR-T) therapy, immune checkpoint inhibitors (ICIs) therapy, and tumor vaccine method. Among them, the most widely used immune checkpoint inhibitors, represented by the PD-1/PD-L1 inhibitors, are the most clinically effective ones [[2], [3], [4]]. The programmed cell death protein-1 (PD-1) is a member of the CD28 family of immune receptors [5], whose main function is to regulate the activation of the immune cells, such as T-cells, B-cells, tumor-associated macrophages (TAMs) and natural killer (NK) cells [6,7]. The programmed cell death-ligand-1 (PD-L1), expressed on the surface of tumor cells, acts as a ligand for PD-1 with different functions, such as induction of regulatory T-cell survival, development, etc [8]. Its over-expression can deliver misregulatory signals to T-cells, thereby inhibiting the T-cell mediated anti-tumor immune response and contributing to tumor cell escape [9,10]. Therefore, blocking PD-1/PD-L1 interactions is considered a promising therapeutic strategy that could not only rapidly restore the immune system, but trigger tumor-specific T-cell responses [11]. To date, the Food and Drug Administration (FDA) has approved six humanized monoclonal antibodies (mAb), namely three PD-1 inhibitors, nivolumab, pembrolizumab, and cemiplimab, and three PD-L1 inhibitors, atezolizumab, durvalumab, and avelumab [[12], [13], [14]], which have demonstrated significant therapeutic efficacy in a variety of tumors. However, antibody drugs have significant drawbacks, such as long half-life, poor permeability, and high price, which greatly limit their applications [15,16]. Therefore, the design of PD-1/PD-L1 small molecule inhibitors is of great significance.

In recent years, researchers have continued to develop new PD-1/PD-L1 inhibitors. Among them, the most prominent work is the work from BMS (Bristol Myers Squibb). They found a series of inhibitors with biphenyl scaffold as the structural feature against PD-1/PD-L1 [17,18]. In 2017, the Holak group reported the mechanism of the BMS compounds that induce PD-L1 proteins to form homodimers which occupy the interface for binding to PD-1, ultimately blocking its downstream signaling [[19], [20], [21]]. The eutectic structure shows that PD-1/PD-L1 has an extended, flat, and hydrophobic interface, so the design of a suitable drug molecule remains a challenge.

Based on the structural features of the BMS series compounds, new pathway inhibitors are being developed (Fig. 1). In 2019, Gong's group designed a series of triazoline-containing compounds using a docking ring fusion strategy, of which A22 exhibited good biological activity with an IC50 value of 92.3 nM [22]. In 2021, Wang's group used the same strategy in the biphenyl portion of the molecule to develop a novel inhibitor containing the structure of 4-arylindolines, with a representative A30 also showing high biological activity and an IC50 value of 11.2 nM [23]. In the same year, Jiang's group designed a compound with triaryl compounds containing a pyridine ring as the basic backbone, among which, compound 24 not only exhibited good biological activity with an IC50 values of 3.8 nM, but also had good pharmacokinetic characteristics and oral bioavailability in vivo [24]. In 2020, Li's group designed and synthesized a series of triaryl derivatives using linear aliphatic amines as the linker chain, which were able to target PD-1/PD-L1 interactions, of which, 58 had an IC50 value of 12 nM [25]. In 2021, Zhang's group used a unique difluoromethoxy as the linkage chain, replacing the original ether chain, aiming to improve metabolic stability in vivo [26]. Among them, 43 with an IC50 value of 10.2 nM, significantly promoted the activation of CD8+ T-cells at a low dose of 1 mg/kg in a Hepa1-6 gene mouse model. In 2022, a novel oxadiazole-containing inhibitor was discovered also in Jiang's group [27], 17 with an IC50 value of 27.8 nM, inhibited PD-1/PD-L1 interaction and promoted the dimerization, internalization, and degradation of PD-L1.

In this manuscript, we designed a series of novel small molecule inhibitors using triaryl groups as the basic backbone and ester chains as connectors. Finally identified 22, which showed the best biochemical activity in vitro, and subsequent studies on anti-tumor activity in a mouse 4T1 breast cancer model, showed a powerful ability to restore the tumor immune microenvironment, as well as inhibit tumor growth.