The receptor PD-1 is a protein having 288 amino acids with a N-terminal IgV-like domain. It is also known as CD279 and was first observed in IL-3 lacking LyD9 (murine hematopoietic progenitor) and 2B4-11 (murine T-cell hybridoma) cell lines (Ishida et al., 1992). It shares 13% sequence similarity with induced T-cell co-stimulator, 15% homology with CD28 and 20% similarity with CTLA4. The constitutive expression of PD-1 is present in immature thymocytes, activated CD4⁺and CD8⁺T-cells, B-cells, DCs, NK cells (Carreno and Collins, 2002). The expression of PD-1 is induced on APCs, monocytes and DCs via transcription factors namely NOTCH, Forkhead box protein (FOXO1), interferon regulatory factor (IRFs) and nuclear factor of activated T-cells (NFAT) (Ahmadzadeh et al., 2009). Additionally, IL-10 and TGF-β could stimulate the PD-1 expression in chronic infections. The increased expression of PD-1 is a marked feature of exhausted T-cells (Staron et al., 2014).

For the expression of PD-1 gene, the conserved upstream regulatory regions namely CR-B and COR-C are crucial. In the CR-C region, there is a binding site which is connected to the NFATc1 (NFAT2) in TCD4 and TCD8 units (Li et al., 2015). Additionally, in the CR-B region, c-FOS is associated leading to increased expression of PD-1. When NFATc is stimulated, it binds to the pdcd1promoter leading to expression (Youngblood et al., 2011). Moreover, IFN-α in association with IRF9 results into the expression of PD-1 by binding to the pdcd1 promoter leading to impairment of T-cells. In case of chronic infections, PD-1 is known to express in T-CD8 cells which are exhausted and is followed by binding of FOXO1 transcription factor to PD-1 promoter for increasing its expression (Xiao et al., 2012). In cancer cells as well, the tumor cell leakage enhances the expression of c-FOS subunit of AP-1, which in turn, enhances the PD-1 expression. However, PD-1 can act in two different ways i.e., it can be harmful and beneficial both (Salmaninejad et al., 2018). It plays a protective role in decreasing the regulation of harmful immune responses or acts as an immunosuppressor and regulates immune tolerance. On the other hand, it can also lead to the dilation of cancer cells by interfering with the protective immune response (Han et al., 2020).

PD-L1 is a ligand of co-inhibitory receptor PD-1 and is coded by chromosome 9p24.1 located on the CD274gene. It is also called as B7 homolog 1 due to its homology with B7-1, B7-2, and CD274 (Sanmamed and Chen, 2014). Under physiological conditions, constitutive PD-L1 expression takes place in several tissues, mainly in the MSCs, activated T-cells, B-cells, monocytes, DCs, bone marrow derived mast cells and several immune privileged organs (Sharpe et al., 2007). Additionally, PD-L1 expression can be stimulated by γ-chain cytokines and IL-21 in T-cells and CD19⁺B cells respectively. Lipopolysaccharide (LPS) or B-cell receptor activation in B-cells is also known to stimulate PD-L1 expression. IFN-γ in monocytes and non-lymphoid cells like endothelial cells can also stimulate the expression (Ohaegbulam et al., 2015). PD-L1 is extremely conserved evolutionarily which reveals its functional importance. The PD-L1 is often seen at the site of inflammation and on tumor cells of distinctive origin which suggests the wide disposition of PD-L1 in various cellular localizations (Ji et al., 2015). These involve membranous PD-L1 (mPD-L1), nuclear PD-L1 (nPD-L1), serum PD-L1 (sPD-L1), cytoplasmic PD-L1 (cPD-L1) and exosomal PD-L1. It is present as an acquired immune mechanism in cancer cells to skip anti-tumor responses. It is linked with immune surroundings rich in CD8 T-cells, Th1 cytokines, chemokines, and interferons. Studies revealed that IFN-γ results into increased expression of PD-L1 in acute myeloid leukemia via MEK/ERK and MYD88/TRAF6 pathways (Bellucci et al., 2015). IFN-γ activates protein kinase D isoform 2 (PKD2), which is crucial for regulating PD-L1. The inhibition of PKD2 prevents PD-L1 expression and induces an antitumor immune response. NK cells secrete IFN-γ via Janus kinase (JAK) and by activating signal transducer and activator of transcription 1 (STAT1) transcription pathways resulting into enhanced PD-L1 expression (Bellucci et al., 2015). Additionally, PD-L1 expression is regulated by IFN-γ released by T-cells via JAK/STAT/IRF1 pathway. PD-L1 is the protein receptor encoded by the CD274 gene, and is located on chromosome 9 in humans. The mRNA expression of PD-L1 takes place by the production of two alternative transcripts of CD274 followed by translation of PD-L1 protein receptor. The longer transcript contains seven exons having a coding sequence of 800 bp and codes for a 33-kDa protein. PD-L1 is a membrane bound glycoprotein with a big extracellular region having immunoglobulin (Ig)-like domains, a small cytoplasmic domain of 30 amino acids and a hydrophobic transmembrane domain (Garcia-Diaz et al., 2017; Khurana et al., 2021). Exon 1 encodes for the 5′-UTR while exon seven codes for the 3′UTR and an intracellular domain. Another transcript is produced through alternative splicing and lack of third exon produces a small 160 amino acid isoform of PD-L1 without having IgV-like domain. Moreover, the promoter region of PD-L1 contains CpG methylation sites alongwith a 220-bp region of epigenetic regulation. The 3′-UTR region of PD-L1 is longer and involves several cis acting elements, which are associated with mRNA decay (Han et al., 2020). Furthermore, PD-L2 is another ligand of PD-1 which shares around 60% sequence homology with PD-L1 in humans (Latchman et al., 2001). As compared to PD-L1, the expression of PD-L2 is limited up to activated DCs, macrophages, bone-marrow derived mast cells and B-cells (Zhong et al., 2007). The expression of PD-L2 could be stimulated by LPS and B-cell receptors in B-cells, and granulocyte-macrophage colony-stimulating factor (GM-CSF), IL-4 on DCs. Both the PD-1 ligands are present in tumor cells and in chronic infection, but the expression of PD-L2 is lower as compared to PD-L1 (Ohigashi et al., 2005).

The association of PD-1 with its ligands are crucial for regulating a balance among autoimmunity and immune tolerance, alters anti-viral and anti-tumor immune response (Sun et al., 2018). PD-1 transmits signals when it is linked to B-cell receptor (BCR) or TCR and hence, results in the formation of co-inhibitory microclusters with TCR and CD28 (Yearley et al., 2017). It is followed by the inhibition of various processes like preventing cytokine formation, glucose consumption and proliferation of T-lymphocytes. Furthermore, it prevents the expression of transcription factors linked with effector function such as GATA-3, Eomes and T-bet (Nurieva et al., 2006). The ligation of PD-1 results in decreased phosphorylation of protein kinase C, CD3 and ZAP70 (Sheppard et al., 2004). It also prevents the induction of ERK in T-cells and B-cells, and inhibits the phosphorylation of PLC-γ2, Syk, Igβ and calcium mobilization (Freeman et al., 2000). Hence, PD-1 ligation blocks signaling regulated by TCR and hampers the functioning of PI3K-Akt, Hedge-hog, Wnt and Ras/MEK/ERK pathway as shown in Figure 1 (Okazaki et al., 2001).

PD-1 prevents the induction of PI3K-Akt pathway by maintaining the PTEN kinase and phosphatase activity, which results into inhibition of Cdk2 and Skp2 expression as well as an enhancement in p27^kip1. The binding outcome of PD-1 with PD-L1 varies depending upon the cell type (Parry et al., 2005). In the case of T-cells, PD-1 inhibits its functioning by inhibiting several processes such as disturbing the cell cycle progression and preventing the formation, proliferation and survival of cytokines (Patsoukis et al., 2012). Moreover, PD-1 is linked with TGF-β regulated T_regs signaling resulting into their proliferation (Butte et al., 2007). Furthermore, Ras PD-1 controls SWAT3 and gives enhanced effect along with TGF-β. PD-1 ligation also fluctuates the metabolic environment of T-cells and in turn produces an oxidative environment (Tkachev et al., 2015). PD-1 signaling is extensively studied and reviewed in literature. However, our understanding of PD-1 signaling in CLDs is still limited (Dermani et al., 2019). PD-1/PD-L1 blockade based anti-tumor therapy is widely used in various types of cancers. Depending upon the type of cancer, the expression and signaling of PD-1/PD-L1 varies. For instance, in HCC patients, the expression of PD-1 is upregulated in CD8⁺ T cells and an increased number of circulating and tumor-infiltrating PD-1⁺/CD8⁺ T cells are seen which are associated with progression of HCC (Shi et al., 2011; Jubel et al., 2020). In breast cancer, PD-L1 expression is linked to epithelium-to-mesenchymal transition (EMT) in human breast cancer stem cells (BCSCs). It has been reported that the expression of PD-L1 is higher in estrogen receptor (ER)α-negative breast cancer, while a subsequently low expression is seen in ERα-positive breast cancer cell lines. In pancreatic ductal adenocarcinoma (PDAC), enhanced expression of PD-1 was seen in peripheral CD8⁺ T cells, while in bladder cancer it has been reported that autophagy related 7 (ATG7) protein regulates the expression of PD-L1 protein and overexpression of ATG7 results in increased PD-L1 protein levels by stimulating autophagy-dependent degradation of FOXO3A. Additionally, in colorectal cancer (CRC), expression of PD-L1 is generally common in metastatic CRC. PD-L1 expression promotes tumor cells skipping the surveillance of the immune system and increases functioning of T_reg in CRC, thus promoting metastasis (Dong et al., 2017; Li et al., 2020).

Hepatocellular carcinoma (HCC) is considered a very common neoplasia of the liver. It is placed in the second position for causing cancer associated mortality and occupies the 16th position as a common cause of death globally. Based on intensive research in oncology, it is considered that immunotherapy is by far the most effective therapy for HCC. Immunotherapy is effective for HCC as liver is an immune privileged organ in which every immunotherapy based drug has its individual pharmacokinetic profile. Additionally, liver can tolerate to immune response to antigens which is maintained by naive T-cell activation and also by several other immunosuppressive processes. Moreover, HCC is a type of cancer which is linked with inflammation. Immune response takes place by coordination among the stimulatory and inhibitory signals. Among inhibitory signals, PD-1 and PD-1 ligands are considered as most effective and have gained wide attention (Cristescu et al., 2018). In physiological conditions, PD-1/PD-L1 is expressed for regulating self-tolerance and inhibits immune stimulation by T-cell activation. However, cancer cells also show PD-1 resulting into immune escape. Indeed, drugs targeting PD-1 and PD-1 ligands are known to stimulate attentive antitumor effects in HCC as shown in Figure 2. It has become one of the most successful mechanisms in treating HCC in the past few years (Ringelhan et al., 2018). Targeting PD-1 and its ligands is promising due to the possibility of PD-1/PD-L1 staining in HCC patients after surgical resection with prognostic implications (Elsegood et al., 2017). Moreover, serum PD-1 can also be used as a means to monitor prior reoccurrences and to recognize the treatment outcomes. The PD-1/PD-L1 can also be able to serve individual specific treatment for HCC (Wang et al., 2011).

HCC generally takes place due to the occurrence of chronic liver disease (chronic hepatitis infection), metabolic disbalancing or alcoholism. All these promotes exhaustiveness of T-cell consumption and immunosuppressive condition of liver (Shrestha et al., 2018). In tumor growth and progression, immune checkpoints namely PD-1/PD-L1 are highly involved. Studies have shown that HCC patients show high expression of PD-1 in CD8⁺ T-cells and display increased circulating and tumor-infiltrating PD-1⁺/CD8⁺ T-cells, which can be used for disease prediction and to assess postoperative recurrence (Shi et al., 2011). The interaction between activated PD-1 on T-cells and PD-L1 regulates the downstream signaling of T-cell receptor and CD28 co-stimulator signals by adding phosphoryl group to the cytosolic immune receptor tyrosine dependent switch motif resulting into the placement of src homology region 2 domain having phosphatases 1 and 2 (SHP1/2) and slam associated protein (Gao et al., 2009). Additionally, SHP1/2 removes phosphate groups from the TCR and CD28 signaling molecules such as ZAP70 and PI3K, preventing activation of T-cells, increases production of cytokine, promote the expression of pro-apoptotic molecules, and finally lead to apoptosis or anergy of T-cells (Jung et al., 2017). The expression of PD-L1 in cancers headed to exhaustion and unresponsiveness in T-cells, offer immune escapism and progression of tumor. The abrogation of PD-L1 on tumor cells can also increase the sensitivity towards T-cell mediated killing. Intrinsic signaling of PD-L1 is not explored as PD-1, but the treating macrophage with anti-PD-L1 antibodies showed the enhancement in the activity of mTOR pathway and analysis of RNA-seq showed an enhancement in multiple macrophage inflammatory pathways (Kudo, 2019). The immunotherapies focusing on PD-1/PD-L1 showed significant anti-tumor results with very less side effects in patients in advanced stages of cancer. These are reported to enhance the multiplication of tumor-infiltrating lymphocytes and generate a more clonal TCR population in the T-cell population which work against the cancer cells (Nishida and Kudo, 2018).

PD-1 and PD-L1 are largely studied and clinically proven immune checkpoint (Table 1). Studies revealed that PD-L1 prevents tumor cells from direct attack of cytotoxic T-cells. The association of PD-1 present on the cancer cell surface and PD-1 present over CD8⁺T-cells leads to apoptosis and energy in CD8⁺T-cells (Xu-Monette et al., 2017). The expression of the high-mobility group box (TOX) DNA-binding factor in thymocyte selection overlap with the expression of PD-1 and can promote the phenotype and longevity of the exhaustive T-cells. Additionally, PD-L1 can also act as an anti-apoptotic in cancer cells as this may lead to the development of resistance in cancer cells. The inhibitory immune checkpoint expression could be dysregulated in the tumor region, which results into the progression of T-cell mediated immune response via cancer immunotherapy. It has been found that PD-1 pathway downregulates the activation of T-cells in peripheral tissues in the later stages. It suggests that blocking this pathway could be useful in getting possible results. The Food and Drug Administration (FDA) approved anti-PD-1 and anti-PD-L1 antibodies for commercial usage (Gong et al., 2018).

The intrinsic PD-L1 pathway is abnormally stimulated in many cancers. There are various factors which regulate PD-L1 in cancerous cells such as epigenetic regulation, genetic modifications, oncogenic and tumor suppressor pathways, inflammatory cytokines and various other factors.

When chronic viral infection takes place, a prolonged persistence of viral antigen is observed. As a result of which there is continuous activation of antigen specific T-cells which induces the entry of antigen specific T-cells into a stage known as T-cell exhaustion (Wang et al., 2011). The impaired T-cells lose various effector activities such as decreased amount of cytokine production specifically IL-2, and decrease in their cytotoxicity and proliferative potential. Moreover, exhaustion of T-cells can lead to increase in the expression of co-inhibitory receptors (Mataki et al., 2007). The expression of PD-1/PD-L1 on antigen specific T-cells and exhaustion of T-cells is studied in various chronic and acute viral infections including hepatitis. Every year around 257 million people are chronically infected with hepatitis B virus (HBV) and an estimate of around 800,000 people die every year because of liver cancer and cirrhosis (Cho et al., 2017). However, the infection with HBV can be prevented via vaccination. Nevertheless, in some cases, the infection is resistant to the treatment. In HBV infection, the PD-1 expression is enhanced in HBV-specific T-cells. The expression of PD-1 is negatively associated with T-cell responses. It has also been studied that PD-1 expression on HBV specific cells in earlier stages of infection is associated with increased serum ALT concentration, hence, pointing that early expression of PD-1 may serve as a biomarker of liver injury (Yao et al., 2007). Recent studies also depicted same outcomes when analysis of PD-1 expression on HBV-specific CD4⁺T-cells were carried out. The blockage of PD-1 pathway is considered as crucial in treating the infectious disease. The outcome and efficiency of PD-1 blocking differ significantly in various studies and among pathogens. In chronic HBV infection, the blocking of PD-1 enhances the proliferation of T-cells and increases the generation of cytokines such as IFN-γ and IL-2 by HBV specific T-cells taken from the liver of patients and peripheral blood region with chronic infection of hepatitis B (Urbani et al., 2008). Tang et al., reported that PD-1 blocking can considerably alter the functioning of CD4⁺T-cell as compared to CD8⁺T-cell function. The PD-1/PD-L1 pathways were earlier reported to downregulate the functioning of HBV specific T-cell function in a transgenic mice model. In this study, the T-cells generated IFN-c which later decreased inspite of the antigen presence in the liver followed by an increment in the expression of PD-1 (Hofmeyer et al., 2011). Blocking of this pathway could delay the downregulation of virus-specific T-cells. Moreover, the occurrence of virus specific T-cells in peripheral blood of patients showed enhanced PD-1 expression and were functionally exhausted while the recovered patient showed lower expression of PD-1. The exhaustion stage of T-cells is reversible i.e., after the blocking of PD-1/PD-L1 pathway, the impaired T-cells are recovered completely (Jin et al., 2010). Similar pattern is observed in case of HCV infection i.e., they also exhibit enhanced expression of PD-1 with decreased effector functions however, by blocking the PD-1/PD-L1 pathway, the situation is restored. Apart from enhanced PD-1 expression, the PD-L1 expression is also increased in hepatic APCs which are responsible for hypo-responsiveness of T-cells in chronic hepatitis infection (Wang et al., 2011). Moreover, along with enhanced PD-1 expression, the dysfunctional HBV/HCV specific CD8⁺ T-cells in the liver exhibit decreased expression of CD28 and CD127, revealing much severe condition (Gehring et al., 2009; Hsu et al., 2010; Li et al., 2020). In contrary, the peripheral dysfunctional T-cell displayed a good expression of CD127, which possibly suggests a lower functional exhaustion (Okazaki and Honjo, 2007). The HBV/HCV infection in liver resulted in exhausted T-cell but the blocking of PD-1/PD-L1 generated different responses in both types of infection. Studies showed that the liver resident HBV specific T-cells are recovered much easily functionally after PD-1/PD-L1 blockade as compared to T-cells which are present at the periphery (Nakamoto et al., 2008). However, no functional restoration of HCV specific T-cell was monitored after PD-1/PD-L1 blockade. These contradictory results signify the specificity of virus specific T-cells in different stages of disease. As discussed earlier, chronic hepatitis viral infection leads to increased PD-1 expression which is linked with dysregulation of T-cells and persistent load of viral antigen thereby favoring chronic hepatitis infection while limiting the immunopathogenesis (Dai et al., 2014). Studies also revealed that chronic HBV infected patients when provided with PD-1 inhibitors, an increment in HBV specific CD8⁺ T-cell is observed (Jin et al., 2010). Immune checkpoint mediated immune restoration can also be associated with severe liver injury and inflammation, liver failure and chances of HCC. The immunosuppressive molecule, PD-1, is associated with the course of HCC and HBV infection. It has been documented that HBV infection stimulates immunosuppression and as the infection progresses, it promotes peripheral immune tolerance which results in oncogenesis, because of impaired surveillance of immune system. T_regs plays a major immunosuppressive role as they secrete cytokines namely TGF-β, IL-10, and IL-35, and also inhibit the activity of Th₁ or Th₂. It has also been reported that in HBV⁺ HCC, the number of T_regs were more as compared to HBV⁻ HCC patients. The increased expression of genes associated with IL-10 pathway, FOXP3, and the immunosuppressive molecules namely cytotoxic T-lymphocyte-associated protein 4 (CTLA-4) and lymphocyte-activation gene 3 (LAG3) were seen in HBV-infected HCC. Additionally, resident memory T cells and myeloid-derived suppressor cells were also enriched in HBV-infected HCC. All these cells promote continuous immune suppressive effects in HBV which finally progresses into the development and progression of HCC. Additionally, the genome of virus is integrated in the DNA of hepatocytes and produces viral proteins. Hepatocytes expressing viral protein can be targeted by nonspecific and uncontrolled immune response. However, the immune checkpoint inhibitor mediated toxicity can be managed by treatment with corticosteroids (Cho et al., 2017).

PD-1 co-inhibitory receptor aids in immunoregulation by decreasing the initial activation of T-cell, hampers T-cell effector functions and differentiation (Dyck and Mills, 2017). It has been well documented that signaling of PD-1 through T-cells restricts the immune mediated tissue injury during the course of infection. Additionally, PD-1 also restricts the activation of self-reactive T-cells (Sage et al., 2018). The immunoregulation of PD-1during inflammation allows foreign microorganisms to escape the immune defense mechanisms of the host. Increase in the expression of PD-1 is noticed in acute liver infections caused by viruses (i.e., HBV and HCV), and bacterial infections (Sharpe et al., 2007). During acute liver injury, specific T-cells get activated with upregulation of various cell types via pattern recognition receptor signaling or indirectly by allowing the release of inflammatory cytokines. The function of PD-1 in acute infection revealed that PD-1 is associated with proving protection against lethal immunopathology (Dong et al., 2019; Allawadhi et al., 2021). However, PD-1 is not much explored in acute liver injury, and few studies suggest that the expression of PD-1 acts as a crucial biomarker for predicting the outcomes of associated diseases (Erickson et al., 2012). By utilizing the expression of PD-1 as a marker of activation or dysfunction of immune cells in acute injury may offer several benefits when considering the treatment strategies to follow.

The inhibitors targeting PD-1/PD-L1 have been known to play considerable role in cancers. Herein, we have discussed some of the important inhibitors with potent activity.

Fatigue is considered the most common side effect of anti-PD-1/PD-L1 agents. It has been revealed that Nivolumab administration resulted in fatigue in 16–24% of patients. Additionally, it has been observed that anti-PD-1 agents showed 16–37% chances of fatigue while anti-PD-L1 agents showed 12–24% incidence (Brahmer et al., 2015). However, the incidence of fatigue was increased and range from 21 to 71% when anti-PD-1/PD-L1 agents were administered with other chemotherapeutic drugs, anti-angiogenic agents and targeted therapies. Fatigue associated with combinational therapy can also generate other systemic manifestations like illness and cytokine release (Bendell et al., 2015). The mechanism of associated fatigue is unknown, however, it is suggested that it is not dose dependent (Weber et al., 2015a).

Fever and chills are generally associated with other immunotherapeutic agents also including vaccines, immune modulating antibodies or targeted therapies. The basic mechanism behind these toxicities is the release of cytokines and non-specific stimulation of the immune system and related response. However, these are managed easily by providing antipyretics and non-steroidal anti-inflammatory drugs (Weber et al., 2015a).