Whole exome sequencing/whole genome sequencing (WES/WGS) showed that HBV infection was associated with DLBCL. Studies indicated several mutations in genes related to the AID/APOBEC enzymes in patients with HBV-associated DLBCL (32, 33). Ren et al. (32) detected the genes mutation in 275 patients of DLBCL, including 20% patients with HBsAg positive (56/275). And the tumor biopsy specimens of 229 patients were taken at diagnosis, and 46 were taken at relapse. The quantitation of viral DNA was performed in 44 HBsAg positive patients. Fourteen genes were confirmed to be preferentially mutated in the HBV infection group, including KLF2, TMSB4X, CD70, BCL-6, FAS, TNFRSF14, UBE2A, CD58, SGK1, ZFP36L1, CXCR4, FOXO1, CSK, and MSL2. In these genes, the enrichment of genes regulated by BCL-6, FOXO1, and ZFP36L1 and involved in signaling pathways, including BCR, janus kinase/signal transducer and activator of transcription (JAK/STAT), and nuclear factor kappa B (NF-κB), contributed to the gene expression signature ( Figure 3 ).

BCR signaling activates downstream oncogenic pathways, including NF-κB or PI3K (34). The PI3K-Akt-mTOR signaling cascade is known to be dysregulated and represents a major regulator of cell survival-proliferation (35). NF-κB regulates the proliferation of B cells and promotes the expression of proto-oncogenes (Myc and BCL-6) and cellular anti-apoptotic protein (BCL-2), as well as the secretion of cytokines, including tumour Necrosis Factor alpha (TNFα), lymphotoxin-α, and interleukin-6; it further positively regulates the JAK/STAT3 pathway and cell proliferation and promotes tumorigenesis. As a key transcription factor, BCL-6 plays a crucial role in the clearance of HBV (36). Furthermore, BCL-6 is a crucial gene of HBV-associated DLBCL. There was no significant difference between the HBsAg positive and HBsAg negative groups in the percentage of GCB vs. non-GCB patients. The incidence of GCB DLBCL was 37.5% vs. 38.1%, respectively; non-GCB was 62.5% vs. 61.9%, respectively (32). Zhang et al. showed that HBV was important in DHL/THL DLBCL and that the incidence of Myc/BCL-6 DHL was higher than that of Myc/BCL-2 DHL with HBV infection (37). A previous study of ours showed that the rate of Myc rearrangements in patients with HBV-associated DLBCL was significantly higher than that in HBV-free individuals (38). Moreover, HBx simultaneously activated the Src/Ras/ERK pathway, promoting proliferation and anti-apoptosis owing to the accumulation of ROS and endoplasmic reticulum stress (39). Therefore, HBV-associated DLBCL is involved in the signaling pathways, including BCR, NF-κB, and JAK/STAT3, and further dysregulates oncogenes, including BCL-6, Myc, which are associated with poor prognosis. Moreover, following HBV infection, the human body activates the abnormal BCR signaling pathway, which could be more directly related to HBV-associated DLBCL.

In the reactive phase, HBV could not be completely eliminated and induce immune escape, which is associated with the poor survival of patients with HBV-associated DLBCL. During chronic HBV infections, virus-specific T-cells appear deeply exhausted. Both CD8 and CD4 T-cells up-regulate co-inhibitory receptors, which can inhibit T-cell function following the cross-linking of their ligands, entailing T-cell exhaustion and tumor escape. For instance, the programmed cell death 1/programmed cell death ligand 1 (PD-1/PD-L1) cell signaling pathway is closely related to the functioning of the immune system, which is currently a research hotspot in the field of tumor immunotherapy. The activation of the PD-1/PD-L1 cell signaling pathway can suppress the function of the immune system and contribute to the immune escape of cancer cells (40). Our previous study showed that the incidence of PD-1 expression among patients with HBV-associated DLBCL was 4.3-fold higher than that among HBV-free individuals (40.0% vs. 9.4%; p=0.010). Moreover, the median OS and PFS were the worst in PD-1-positive patients with HBV-associated DLBCL. These results indicate that the dismal prognosis of patients with HBV-associated DLBCL may be related to the high rate of PD1 expression (15) ( Figure 3 ).

Moreover, the number of CD4-positive lymphocytes has been reported to decrease significantly after chemotherapy in patients with HBV-associated DLBCL, and the CD4:CD8 ratio decreased for a longer time in the aforementioned population than in HBV-free individuals, elucidating the reason behind the poor prognosis of patients with HBV-associated: The host immune system weakened the tumor surveillance effect, leading to rapid disease recurrence and poor prognosis (41). Moreover, Ren et al. (32) showed that CD70, TNFRSF14, and CD58 were the main mutated genes among patients with HBV-associated DLBCL, leading to a decrease in T-cell infusion in the tumor microenvironment and thereby weakening tumor immune surveillance and accelerating their escape.

In summary, the continuous high expression of multiple co-inhibitory molecules is the key cause of the depletion of HBV-specific T-cell function. Blocking these molecules can significantly restore HBV-specific T cell function and subsequently remove the virus in vivo. Therefore, the blocking of co-stimulatory molecules might represent a novel strategy for the clinical treatment of HBV infection.

HBx does not bind to DNA, and HBx-activated RNA polymerase (pol) I-, II-, and III-dependent promoters directly interact with some transcription factors and stimulate signal-transduction pathways. The HBx protein can initiate epigenetic modifications to dysregulate noncoding RNAs expression, which, consequently, can regulate downstream epigenetic changes throughout the pathogensis of HBV-associated DLBCL. Bruni et al. (42) found that patients with HBV-associated indolent B-NHL have dysregulated miRNA. Chen et al. (43) also showed that this dysregulation of miRNAs was closely related to the proliferation and differentiation of B-cells. Of note, miRNAs regulate through histone deacetylase (HDAC) (44) or the NF-κB pathway (45). While HDAC inhibitors can break this negative regulation by promoting the expression of miR-34a (46). Several studies have shown that epigenetic modification play a significant role in patients with HBV-associated DLBCL, which have a higher frequency of the CREB-binding protein (CREBBP) mutations (27.2%) compared to the general population (47). CREBBP mainly regulates the activity of histone acetyl transferase (HAT), maintains the activity of HAT domain, regulates transcription factors, and affects immune regulation. However, CREBBP mutation (loss-of-function) inactivates the HAT domain and impairs acetylation-deacetylation balance, thereby increasing BCL-6 expression ( Figure 3 ). Nevertheless, the selective inhibition of HDAC3 reverts the molecular phenotype of CREBBP mutations (48, 49). Moreover, SIRT1 plays an important role in the acetylation of B-cell germinal center and could lead to an imbalance of acetylation/deacetylation. SIRT1 could cause the deacetylation of BCL-6, p53, and other transcription factors (50) and up-regulate the expression of BCL-6 (51). BO et al. showed that chronic stimulation with HBsAg promoted the viability of the human B lymphoblastoid cell line through regulation of the SIRT1-NF-κB pathway (52) ( Figure 3 ). In conclusion, noncoding RNAs and epigenetic regulation may play roles in the pathogenesis of HBV associated DLBCL.

HBx elicits a DNA damage response. It binds to p53 in the nucleus, inhibits the expression of p53-responsive genes, promotes the phosphorylation (inactivation) of Rb, lowers the activities of CDK inhibitors, promotes vascular regeneration, and inhibits p53 function through Rb, thereby inhibiting cell apoptosis and cell cycle (53). HBx itself can inhibit p53 from functioning as a cell cycle blocker, thereby promoting cell survival. Moreover, the overexpression of HBx significantly reduces the activation of CHK2 signals induced by S-period blockers and p53 and p21 in patients with HBV-associated DLBCL, consequently reducing the sensitivity of chemotherapy drugs and suggesting that CHK2 may be a potential factor in HBx-induced chemotherapy resistance (54). Li et al. (55) found that HBx directly up-regulated the expression of lncNBAT1, a long non-coding RNA (lncRNA) that is closely associated with the chemotherapy outcomes of patients with HBV-associated DLBCL. The up-regulation of lncNBAT1 reduced the sensitivity of DLBCL cells to the chemotherapeutic agents that induced S-phase arrest, whereas the knockdown of lncNBAT1 significantly relieved the chemo-resistance of HBx-expressing DLBCLs ( Figure 3 ).

Therefore, patients with HBV-associated DLBCL have a poor prognosis owing to genomic instability, immune escape, epigenetic regulation, and chemo-resistance, leading to an uncontrolled cell cycle, obstruction of apoptosis, and increased frequency of gene mutations. Moreover, many signal pathways, including BCR, JAK/STAT3, and the NF-κB pathway, are involved in the pathogenesis of HBV-associated DLBCL ( Table 1 ).

With advances in epigenetic research in recent years, the use of epigenetic regulators has gradually increased in clinical practice. Several clinical trials have confirmed the effectiveness and safety of HDAC inhibitors, including chidamide, in first-line consolidation and rescue therapy for the recurrence of lymphoma. Chidamide is a selective inhibitor of histone deacetylases (HDACs), which is mainly targeted at class I HDACs, and has a regulatory effect on the epigenetic function of tumor abnormalities. At our center, the First Hospital of Jilin University, Changchun, China, a clinical trial ( Table 2 ; registered in www.clinicaltrials.gov under number NCT04661943) of chidamide maintenance treatment was conducted for patients with HBV-associated DLBCL. The inclusion criteria were achievement of complete response and complete response lasting for 1 year following systemic treatment before enrollment. The trial aimed at exploring the efficacy of epigenetic regulators in patients with HBV-associated lymphoma.

Such as PI3K, BCL-6, and BCL-2 inhibitors, and the immunomodulator lenalidomide have shown efficacy in treating recurrent and refractory DLBCL (57–60); however, clinical trials on the efficacy of these treatments for HBV-associated DLBCL are warranted.

CAR-T for the failure of multiline therapy of DLBCL has been demonstrated to have significant efficacy. Patients with HBV-associated DLBCL undergoing CAR-T cell therapy did not increase the risk of severe cytokine release syndrome (61). However, Yang et al. showed that patients with HBV-associated DLBCL receiving CD19-CAR-T cell therapy are at risk of HBV reactivation, especially in HBeAg-positive patients; the close monitoring of HBV-DNA levels and adequate antiviral prophylaxis are essential for the prevention of HBV reactivation (62). HBV-associated lymphoma presents immune escape-related gene mutations such as CD70, and targeted CD70 CAR-T may have potential efficacy.

PD-1 expression was higher in patients with HBV-associated DLBCL than in HBV-free individuals. Thus, immune-checkpoint inhibitors may have a satisfactory effect on patients with HBV-associated DLBCL. However, there is no standard first-line therapy, and clinical trials exploring specific treatment regimens for HBV-associated DLBCL are rare compared with those for other virus-related lymphomas. Further studies to explore more effective therapy regimens are warranted.