Study on the Anti-tumor Mechanism of Immunological Checkpoint Inhibitor PD-1

,2,3*

1. College of Life Science & Technology, Heilongjiang Bayi Agricultural University, Daqing 163319, China; 2. College of Food Science & Technology, Heilongjiang Bayi Agricultural University, Daqing 163319, China; 3. National Coarse Cereals Engineering Research Center, Daqing 163319, China

Abstract In recent years, the research of programmed death factor-1 (PD-1) and its ligand (PD-L1) has made a great breakthrough in tumor therapy, and it is expected to change the clinical treatment manner of anti-tumor. In this paper, the role and mechanism of PD-1 inhibitors in anti-tumor are reviewed, thereby promoting their clinical application in anti-tumor immunotherapy.

Key words PD-1, PD-L1, Anti-tumor, Immunotherapy

1 Introduction

Programmed death factor-1 (PD-1) is co-inhibitory molecule on the surface of immune cells, and it negatively regulates the immune response. The combination between PD-1 and its ligand PD-L1 (B7-H1 and CD274),PD-L2 (B7-H2 and CD273) and corresponding antibodies could inhibit immune microenvironment around tumor tissue, down regulate activity of specific T cells, and promote tumor cells to escape immune surveillance. Different from radiotherapy and chemotherapy, which directly kill tumor cells, PD-1 inhibitors activate anti-tumor immune response by blocking the access of immunological checkpoint in tumor microenvironment, further inhibiting growth of tumor cells. Via clinical statistics of PD-1 inhibitors, it is found that PD-1 inhibitors are only suitable for cancer patients whose PD-L1 expression is more than 50%, and the curative effect is different in patients. For therapy limitations of PD-1 inhibitors, researchers turn their attention to combination therapy, hoping to improve the effective treatment rates of patients. In this paper, the anti-tumor effects and mechanisms of mono-therapy and combination therapy by PD-1 inhibitors in recent years are reviewed, which could provide theoretical basis for clinical immunotherapy of anti-tumor.

2 Structure of PD-1

PD-1 is also called CD279, is mainly expressed on immune related cells, such as CD4+T cells, B cells and natural killer cells (NK cells), and is a kind of co-inhibitory receptor on cell surface. It suppresses immune response during carcinogenesis, and promotes the tumor cells to escape the immune response of effector T cells. PD-1 in mature form contains extracellular domain (147 aa), hydrophobic transmembrane region (27 aa), cytoplasmic region (94 aa), tail of its cytoplasmic segment has immunoreceptor tyrosine-based inhibitory motif (ITIM) and immunoreceptor tyrosine-based switch motif (ITSM). It is found that ITIM and ITSM are important structural bases of PD-1 exerting immunosuppressive function[1].

3 Expression of PD-1

In normal conditions of the body, expression of PD-1 is induced by T cell activation, which starts feedback inhibition, prevents over activation of effector T cells to damage to the body. In body of cancer patients, PD-1 combines with PD-L1 on surface of tumor cells, which could induce phosphorylation of N-terminal ITIM of PD-1 in cytoplasm, thereby collecting the activated protein tyrosine phosphatase (SHP-2) to C-terminal ITSM, and the activated PD-1 mediates cycle arrest of immune cells, declines production of cytokines, and down regulates metabolism of immune cells. The researches show that positive rate of PD-1 has significantly positive correction with Foxp3+Tregs infiltration in renal cell carcinoma, lymph node metastasis, FIGO staging of ovarian cancer infiltrating lymphocytes (TILs). Additionally, over-expression of PD-L1 in ovarian cancer, meningioma, and melanoma directly promotes tumor growth and metastasis[2]. Therefore, the down regulation of PD-1/PD-L1 combination could obviously inhibit growth and invasion of tumor cells.

4 Anti-tumor effects of PD-1 inhibitors

4.1 PD-1 inhibitors against tumor growthTumor immunotherapy is the current research hotspot, and successful development and clinical application of PD-1 inhibitors has raised the tumor immunotherapy to a new height. The research by Lavakumaretal.[3]showed that PD-1 inhibitors blocked release of NF-κB-dependent cytokines, which provided new ideas for clinical treatment of tumor by PD-1 inhibitors. The research by Javieretal.[4]showed that PD-1 inhibitors could up regulate PI3K/AKT/mTOR signaling pathway, activate T cells, and promote proliferation and differentiation of T cells, further inhibiting recurrence and metastasis of tumor cells. The research by Zhaoetal.[5]showed that PD-1 inhibitors could activate dendritic cells (DCs) to submit tumor associated antigens to T cells, and significantly inhibit tumor growth. In tumor microenvironment, PD-1/PD-L1 inhibitors could block accesses of T lymphocyte immunoglobulin mucin-3 (TIM-3) and PD-1/PD-L1, relieve immunosuppressive function, and promote release of interferon regulatory factors (IRFs). The research by Tangetal.[6]found that T cells inosculating antigen receptor (PD-1-ACR) could conduct targeted aggressive tumor area, and then cytokine secretion, activate and prolong survival time of effector T cells and memory T cells. At present, PD-1 inhibitors have been applied in treating lung cancer, metastatic melanoma and digestive tract tumor. The research showed that it could achieve the effect of anti-tumor by adding PD-1 inhibitors (Nivolumab and Pembrolizumab) or PD-L1 inhibitors (Atezolizumab)[7]. Nivolumab is the first approved PD-1 inhibitor, has better anti-tumor activity in treatment of squamous cell carcinoma and carcinoma nonsquamous cell, and increases overall survival rate of tumor patients. Compared with conventional chemotherapy drugs, it has lower toxicity[8]. Approved by FDA, Pembrolizumab was officially applied in clinical treatment of melanoma in 2014. Clinical results displayed that median survival time of patients in Pembrolizumab treatment group was obviously longer than that in chemotherapy group[9]. Atezolizumab is the first approved PD-L1 inhibitor, and has significant treatment effect for urinary epithelial cell carcinoma[10].

4.2 PD-1 inhibitors coordinating with stimulus signals against tumor growthAccording to the experimental data issued in 2018, the effective anticancer rate of PD-1 inhibitors alone was not ideal. How to delay the survival of patients is still a hot issue. Co-therapy by PD-1 inhibitors and stimulus signals is more effective than monomer therapy. The research by Jiangetal.[11]found that PD-1 antibody coordinating with endostatin could increase the concentration of interferon-γ (INF-γ) in plasma, significantly decline expression of IL-17 and human vascular endothelial growth factor (VEGF), and effectively slow down the growth of tumor in mice. Huangetal.[12]revealed that double antibody blockers (LAG3 and PD-1) could significantly enhance function of effector T cells. Additionally, the research by Shietal.[13]found that PD-1 inhibitors associating with (S)-(-)-n-[2-(3-Hydroxy-1H-indol-3-yl) methyl]acetamide significantly inhibited MDSCs, activated CD8+T cells, and prolonged survival time of tumor bearing mice. The research by Zeng Yalinetal.[14]found that PD-1 inhibitors coordinating with cytochalasin B (CB) could up regulate GPR37/JNK/PPAR-γ pathway, significantly decline the expression of AMP-dependent transcription factor (ATF6), protein kinase (P-ERK), inositol-requiring kinase 1α (IRE1α), P-JNK and PPAR-γ, and promote phagocytosis of macrophages. PD-1 inhibitors coordinating with cytotoxic T-lymphocyte associated antigen-4 (CTLA-4) could restore activity and proliferation of T cells, improve microenvironment in tumor, effectively promote apoptosis of H22 mice tumor cells, and inhibit growth and proliferation of mice hepatoma H22 cells[15]. Additionally, the therapy of irreversible electroporation ablation coordinating with PD-1 inhibitors could inhibit the formation, growth and metastasis of tumor, and significantly prolong survival time of mice[16]. Above results show that combined treatment of PD-1 inhibitors could effectively reduce tumor’s volume and mass and delay the survival of experimental mice.

4.3 PD-1 inhibitors coordinating with immunotoxic cells against tumor growthIn cancer therapy, by coordinating PD-1 inhibitors with killer cells such as CIK cells, NK cells and CTLs cells, it could stimulate effector T cells to produce strong immune response, which could not only regulate tumor cells’ microenvironment but also increase immune function of host cells[17]. Compared with chemotherapy alone, it could significantly enhance the killing effect of CIK cells on A520 lung cancer cell line and prolong survival of patients by coordinating CIK cells with PD-1 inhibitors[18]. But when combined with PD-1 inhibitorsinvivo, the concentration and time of the drug still needed further clinical research, to get a safe and more effective treatment effect. PD-1 inhibitors could significantly up regulate activity of NK cellsinvitro. In MM mice, PD-1 inhibitors coordinating with NK cells could significantly enhance the formation of targeted immune complex between NK cells and tumor cells when compared with PD-1 inhibitor monomer, obviously inhibit the growth of tumor RPMI8226 cells, and improve the therapeutic effect of NK cellsinvivo[19]. The research by Zhaoetal.[20]found that PD-1 inhibitors coordinating with CTLs cells could obviously improve the secretion of tumor necrosis factor-α (TNF-α) and IFN-γ, enhance cytotoxic effect of CTLs on tumor cells, up regulate the activity of caspase, and induce apoptosis of tumor cells. PD-1 combining with CTLs cells had good anti-tumor effectinvivo, could effectively inhibit growth of human MM transplanted tumorinvivo, and significantly improve overall survival of transplanted tumor NOD/SCID mice. Additionally, joint therapy of PD-1 inhibitors and CAR-T cells could ease side effects of CAR-T cell therapy alone, and joint therapy of CAR-T and PD-1 inhibitors will be a new hope to conquer cancer. Above results show that joint therapy of PD-1 inhibitors and immune cells could target the location of tumor and effectively reduce the toxic and side effects in the process of tumor treatment.

5 Conclusions and prospects

In recent years, immunotherapy, along with surgery, radiotherapy and chemotherapy, has become the fourth largest treatment for cancer. At present, inhibitors of immune checkpoint have been used in anti-tumor immunotherapy. Via the researches for many years, it is found that the existence of PD-1 is not conducive to the anti-tumor immunity of the body, and it could effectively inhibit tumor growth by applying PD-1 inhibitor monomer, and coordinating with other stimulus signals (endostatin, TIM-3 and SNA) or immunotoxic cell (CIK cells, NK cells, CTLs cells and CAR-T cells). However, the regulatory factors and related signaling pathways of PD-1 are not clear. Additionally, PD-1 inhibitors have different effects on different types of tumors, and accurate assessment on the patients’ conditions is needed for clinical application. The therapy of PD-1 inhibitors has slow onset and long duration, could induce autoimmune diseases, and cause pneumonia, hepatitis, colitis and endocrine disorders. It needs combining with related clinical practice theory to carry out more systematic and deeper research on PD-1 from animals (constructing nude mouse tumor model and zebrafish model), cells (such as various cell experimental models) and molecular level (such as target molecules in various signal pathways), thereby laying the basis for further promoting the research of PD-1 becoming target molecules of tumor immunotherapy.