Management of Hepatitis B Virus in Allogeneic Hematopoietic Stem Cell Transplantation

The high morbidity of HBV reactivation following allogeneic hematopoietic stem cell transplantation (allo-HSCT) is partially due to the intense immunologic potency of complex therapeutic regimens, the use of antithymocyte globulin and calcineurin inhibitors to prevent graft versus-host disease (GVHD), prolonged immune reconstitution, and hematological malignancies infected with hepatitis B virus (HBV). Immunosuppression results in the reactivation of HBV replication from covalently closed circular DNA (cccDNA) residing in hepatocytes. However, the role of viral mutations during HBV reactivation needs to be validated. All individuals scheduled to receive allo-HSCT or wish to donate stem cells should be screened for hepatitis B surface antigen (HBsAg), antibodies to hepatitis B core (anti-HBc), and HBV-DNA. HBsAg-positive recipients of allo-HSCT have a high risk of HBV reactivation; thus, they should receive prophylactic antiviral therapy. The high barrier to resistance nucleos(t)-ide analogs (NAs) seems to be superior to the low barrier agents. Resolved-HBV recipients have a lower risk of HBV reactivation than HBsAg-positive recipients. Although prophylactic antiviral therapy remains controversial, regular monitoring of alanine transaminase (ALT) and HBV-DNA combined with preemptive antiviral treatment may be an optimized strategy. However, optimal antiviral therapy duration and time intervals for monitoring remain to be established. Accepting stem cells from HBsAg-positive donors is associated with a risk of developing HBV-related hepatitis. The overall intervention strategy, including donors and recipients, may decrease the risk of HBV-related hepatitis following HSCT from HBsAg positive stem cells. In this review, we summarize the issues of HBV in allo-HSCT, including HBV reactivation mechanism, HBsAg-positive recipients, HBV-resolved infection recipients, and donor-related factors, and discuss their significance.


INTRODUCTION
Globally, an estimated 257 million people live with chronic HBV infection (1). The HBV carrier rate is high (6.2%) in the African and Western Pacific regions (2). In China, the prevalence rate of hepatitis B surface antigen (HBsAg) was estimated at 5-6%, and 4.38% of people 15-29 years of age are carriers (3,4). Researchers are aware of HBV reactivation (HBVr) complications in patients receiving chemotherapy, monoclonal antibody (especially anti-CD20 antibody) treatment, and other intensive immunosuppressive therapies. Since covalently closed circular DNA (cccDNA) persists in hepatocytes and other tissues, HBsAg-positive patients and historically HBV infected patients are at a risk of HBVr during immunosuppressive therapy (5)(6)(7). The strength of HBVr is determined by the degrees of immune control and virus immune activity in vivo. In addition, because of the intense immunologic potency of the complex therapeutic regimens, and the use of rituximab and high-dose glucocorticoids, which usually leads to cytopenia, the incidence of HBVr due to immunosuppression is much higher in hematological malignancies than in other diseases (8,9).
Guidelines have been recommended for patients with HBV infection undergoing immunosuppressive and cytotoxic therapy (8,(10)(11)(12)(13)(14). Hematopoietic stem cell transplantation (HSCT) technology has developed rapidly and is expected to become the mainstay treatment for patients with hematologic malignancies. Myeloablative conditioning regimens, antithymocyte globulin and calcineurin inhibitor treatment to prevent graft-versus-host disease (GVHD), high-dose glucocorticoids for GVHD therapy, prolonged immune reconstitution, evolving therapeutic treatments (e.g. ruxolitinib, rituximab, ibrutinib, and monoclonal antibodies) for chronic GVHD therapy, and the risk of donor HBV sources lead to a heighted risk of HBVr complication in hematological patients who accept allo-HSCT. Allo-HSCT is an independent risk factor for HBVr in patients with hematologic malignancies (15). However, knowledge regarding HBVr in allo-HSCT is not comprehensive and there are no standard guidelines for managing HBVr during allo-HSCT. Due to the high probability of HBV infection in hematological patients living in HBV epidemic area and the high frequency of HBVr during HSCT, it is necessary to review the developments made by studies on HBVr in allo-HSCT in recent decades. To comprehensively understand HBVr in allo-HSCT and help physicians deal with HBVr in allo-HSCT, here, we have summarized and reviewed the key issues in this domain.

DEFINITION OF HBV REACTIVATION
Previously there were no standard criteria for HBVr. For HBsAgpositive patients, HBVr was defined as a) 10-fold elevation of circulating HBV DNA compared with baseline levels before HSCT and b) detectable circulating HBV DNA in patients whose serum HBV DNA was undetectable before HSCT. In HBV-resolved patients, HBVr was defined as a) a positive result for the HBsAg test in a patient who previously tested negative (called reverse seroconversion, RS) and b) detectable circulating HBV DNA in patients with undetectable serum HBV DNA before HSCT (6,13,16,17) (Figure 1). In 2018, the American Association for the Study of Liver Diseases (AASLD) recommended stricter criteria for HBVr (10). For HBsAgpositive patients, (1) ≥2 log (100-fold) increase in HBV DNA compared to the baseline, (2) HBV DNA ≥ 3 log (1,000) IU/ml in a patient with previously undetectable levels, (given that HBV DNA levels fluctuate) or (3) HBV DNA ≥ 4 log (10,000) IU/ml, if the baseline level is not available. For patients who are anti-HBcpositive and HBsAg-negative, the criteria are: (1) detection of HBV DNA, or (2) reappearance of HBsAg ( Figure 1). Increase in alanine transaminase (ALT) levels to ≥3 times the baseline level and >100 U/L was deemed a hepatitis flare and the definition of HBV related to hepatitis was hepatitis flare plus HBVr.

HBV REACTIVATION MECHANISM
HBV enters the body and eventually enters hepatocytes through the key liver-specific receptor, sodium-taurocholate cotransporter (18). The nucleocapsid is inserted into the nucleus of hepatocytes and the DNA is converted into cccDNA (19). HBV cccDNA is stable and persistent in hepatocytes, which is the reservoir of HBVr despite serum clearance of HBV (20, 21) ( Figure 2). The host's immune response to HBV infection undergoes an inactive immune tolerance state, active state, and conversion to the immune control phase (22,23). HBV-specific T-cell responses suppress viral replication by both cytopathic effects and non-cytopathic cytokine pathways (24,25). B cells produce antibodies against HBV and inhibit the spread of HBV infection to other hepatocytes ( Figure 2). The first report of HBVr was made in 1975 by Wands in a patient with lymphoproliferative disease undergoing chemotherapy (26). It has been reported that HBV DNA restarts replication due to treatment-induced loss of immune control and immunosuppression (27)(28)(29).
In addition, HBV mutations in the major hydrophilic region of the S domain have been found in HBVr after HSCT in recent years (30)(31)(32)(33)(34)(35). However, there is still no evidence that immuneescape mutations occurred prior to reactivation, or that they are responsible for assisting in the viral reactivation process (30). The role of viral mutations and immune escape during HBVr needs to be validated. There was no phenomenon of HBVspecific CD8 T-cell exhaustion during HBVr reported. Hepatitis occurs after immune system reconstitution and destruction of HBV infected hepatocytes (36) (Figure 2). HBV immune response activities with amplification of HBV DNA cannot be detected in time since they precede ALT alterations and clinical symptoms. HBV-induced hepatocellular damage is considered to be the result of a complex interplay among HBV, hepatocytes, and immune cells of the host; thus, several HBVr patients may have little or no liver dysfunction, while others may have hepatitis flares, interruption of immunosuppressive drugs, hepatic failure, and even death. The serological alteration of HBV and the HBV DNA load in vivo cannot effectively reflect the clinical influence of HBVr after allo-HSCT (37). Recent studies have found a favorable prognosis for HBVr in allo-HSCT recipients (38-42). The RS of HBV-resolved patients facilitates hepatitis B surface antigen seroclearance following antiviral treatment (38, 39). However, previous studies had different HBVr definitions and heterogeneous patient characteristics. Therefore, it is necessary to reevaluate the clinical influence of HBVr in a homogeneous group following consistent HBVr criteria, such as the incidence of HBV-related hepatitis, liver-related mortality, non-relapse mortality, and interruption or reduction of primary immunosuppressants.

HBsAg-POSITIVE RECIPIENTS
HBsAg-positive recipients have been widely recognized as highrisk (>10%) for HBVr following allo-HSCT. A study from Mary Hospital of the University of Hong Kong in 2002 indicated that the historical control without prophylactic antiviral treatment had an incidence of 45% HBVr after allo-HSCT (43). A multicenter retrospective study from the Italian Group for Blood and Marrow Transplantation showed that two-year incidence of HBVr after allo-HSCT in HBsAg-positive recipients was up to 81% without prophylactic antiviral treatment (44). Because of the high risk of  However, data comparing the high barrier to resistance of NAs (ETV, TDF, or TAF) with low barrier agents (LAM, LdT, ADV) was limited. Physicians performing HSCT may be concerned whether the antiviral drug would influence the engraftment of neutrophils or platelets during allo-HSCT. A retrospective study in Brazil indicated that LAM/ETV/TAF treatment had no influence on neutrophil or platelet engraftment in allo-HSCT, which needs to be confirmed in large-sample size studies (52). Based on these studies, nearly all guidelines for the prevention of HBVr associated with immunosuppressive therapy had consensus regarding screening for HBsAg and HBcAb before accepting allo-HSCT treatments (11,13,53,54), and prophylactic antiviral treatment was recommended to decrease the risk of HBVr for HBsAgpositive recipients. However, there is no explicit view on screening for HBV DNA and hepatitis B surface antibody (HBsAb) before accepting allo-HSCT therapy. Additionally, the monitoring interval of serological HBV, the duration of prophylactic antiviral treatment, and choice of NAs remain unclear. One suggestion has been that referred monitoring should continue for 6 months after cessation of immunosuppressive therapy, with 2-month intervals potentially being appropriate (55). A recent publication suggested that serological HBV should be obtained at baseline and evaluated every 6 months during antiviral therapy. Additionally, it should also be checked at least monthly for the first 3 months after the cessation of antiviral therapy and every 3 months thereafter (56). Although there is no consensus on duration of antiviral treatment, the majority of recommendations for prophylactic antiviral treatment duration vary from 6 months to 12 months after discontinuation of immunosuppressive therapy (10,11,13,54).

HBV-RESOLVED RECIPIENTS
HBV-resolved patients were defined as HBsAg-negative but positive for antibodies against hepatitis B core antigen (57). While HBV was not actively amplifying in vivo, the persistent cccDNA in hepatocytes could be amplified again FIGURE 2 | The mechanism of hepatitis B virus (HBV) reactivation. Immune control phase, HBV-specific T-cell suppress viral replication by both cytopathic effects and non-cytopathic cytokine pathways, B cells produce antibodies against HBV and inhibit the spread of HBV infection among hepatocytes, HBV cccDNA persistent in hepatocytes. Immune suppressive phase, T-cells and B-cells are inhibited or eliminated by the immunosuppressive treatments, HBV DNA restarts replication due to treatment-induced loss of immune control. Maybe HBV mutations lead to immune-escape from HBV-specific T-cell, HBV DNA restarts replication. HBV reactivation occurs when HBV DNA amplify actively in vivo. Active immune phase, immune system reconstitution and T-cells attack HBV-DNA and HBV infected hepatocytes. The destructed hepatocytes release ALT and aspartate aminotransferase (AST).
when the patient's immune system is suppressed. Previous prospective studies have shown that HBV-resolved hematological patients who accept immunosuppressive treatment/chemotherapy have a risk of HBVr, with the incidence of HBVr varying from 8.3% to 41.5% (7,(58)(59)(60)(61)(62). However, the risk of HBVr in HBV-resolved patients who accept allo-HSCT is not well known. A multicenter retrospective study from Italy illustrated that 6/50 (12%) of HBV-resolved patients underwent HBVr after allo-HSCT without prophylactic antiviral treatment at a median of 12 (7-32) months after transplantation; the 5-year cumulative incidence of HBVr was 22% (63). A retrospective study from Chiba University found that 4/35 (11%) HBV-resolved patients developed HBVr (64) without prophylactic antiviral treatment. Another retrospective study from San Martino University Hospital indicated that 14/137 (10%) patients had HBVr within a median of 19 months (range 9-77 months) after HSCT (40) without prophylactic antiviral treatment. We found that 13/300 (4.3%) HBV-resolved patients developed HBVr at a median of 588 days (range 455-1,294 days) after HSCT without prophylactic antiviral treatment (39). A recent retrospective study with a similar sample size from the National Taiwan University Hospital reported that 21/485 (4.72%) HBVresolved patients presented HBVr at 16 months (range 8-50 months) after HSCT with no antiviral prophylaxis (42). A higher risk of HBVr was reported by a retrospective study at Hamanomachi Hospital, Japan: 18/69 (26.1%) HBV-resolved patients developed HBVr after allo-HSCT at a median of 440 days (75-1,829) without prophylactic antiviral treatment (65). The only prospective research in the domain of HBV-resolved patients undergoing allo-HSCT was conducted by the University of Hong Kong. The two-year cumulative incidence of HBVr was 40.8% (13/62) without prophylactic antiviral treatment, occurring at a median of 44 (8-100) weeks posttransplantation (41). Based on these studies, HBsAg-negative, anti-HBc-positive patients who underwent allo-HSCT also had a risk of HBV reactivation. However, it remains unclear whether prophylactic antiviral therapy can benefit HBV-resolved patients following allo-HSCT treatment as much as it benefits HBsAgpositive patients. A retrospective study from the University of Genoa evaluated 7 years' worth of single-center data on HBVresolved patients who received allo-HSCT; none of the 50 HBVresolved patients experienced HBVr while on prophylactic LAM treatment (66). However, another study indicated that although the majority of HBV-resolved recipients accepted antiviral treatment, the rates of HBVr in the HSCT group at one and seven years were 2.5% and 57.9%, respectively (38).
There are many controversial issues in HBV-resolved patients receiving allo-HSCT. The risk stratification of HBsAg-negative/ HBcAb-positive HSCT recipients and recommendations on antiviral treatment are inconsistent from different specialized associations ( Table 1). The Asian Pacific Association for the Study of the Liver recommended in 2015 that HBsAg-negative/ HBcAb-positive patients with undetectable serum HBV DNA should be followed carefully by means of ALT and HBV DNA testing, then treating with NAs therapy upon confirmation of HBVr. Most European and American specialized associations provide aggressive views on prophylactic antiviral therapy in HBV-resolved patients. The experts in the domain of immunosuppressive treatment-related HBVr recommend initiating prophylactic treatment or monitoring HBV DNA levels for HBsAg-negative/HBcAb-positive patients undergoing intermediate-risk immunosuppression (6,58). However, whether HBV-resolved patients receiving allo-HSCT belong to the high-risk or intermediate-risk group of HBVr is controversial. Previous studies reported a much lower incidence of HBVr in the HBsAg-negative/HBcAb-positive group than HBsAg-positive patients who underwent allo-HSCT. Large-sample prospective studies are needed to robustly investigate the incidence of HBVr in HBV-resolved patients undergoing allo-HSCT according to the 2018 AASLD definition of HBVr. Moreover, the effectiveness of prophylactic antiviral therapy in minimizing the risk of HBVr for HBVresolved HSCT recipients is not well known. There are no studies comparing high barrier NAs (ETV, TDF, or TAF) with low barrier agents (LAM, LdT, and ADV) in HBsAg-negative/ HBcAb-positive allo-HSCT recipients. The monitoring interval of serological HBV and duration of prophylactic antiviral treatment is also unclear ( Table 1).

UNDERLYING REASONS WHY CONTROVERSIES EXIST IN PROPHYLACTIC ANTIVIRAL THERAPY OF HBV-RESOLVED ALLO-HSCT RECIPIENTS
The protective role of HBsAb has been found in hematological patients receiving chemotherapy. A systematic review described the protective role of the HBsAb, with a lower HBV reactivation rate in HBsAb(+) patients compared with HBsAb(-) patients with hematologic disease (7.1% versus 21.8%; P < 0.001) (9). In a group of HBsAg(-)/HBcAb(+) patients with lymphoma, patients without HBsAb before rituximab-based chemotherapy had a higher incidence of HBV reactivation than those with HBsAb (68.3% vs. 34.4%; P = 0.01) (61). It was reported that exceeding the threshold HBsAb titer of 100 IU/ml was associated with a 0% rate of HBVr (68,69). However, few studies have investigated the role of HBsAb in HBcAb-positive patients who undergo HSCT. The role of HBsAb during HBVr in HSCT is unclear. We stratified 665 HBsAg-negative patients according to HBcAb/ HBsAb presence into four groups; the HBcAb(+)HBsAb(-) group had the highest risk of HBVr among the patient groups (15.7%; P < 0.001). The cumulative HBV reactivation rates were 5.3% in the HBcAb(+)HBsAb(+) group, 0% in the HBcAb(-) HBsAb(-) group, and 2.1% in the HBcAb(-)HBsAb(+) group, with no significant difference among these groups. HBsAb in HSCT recipients conferred a protective effect against HBVr (39). A recent retrospective study from Turkey also reported the protective role of HBsAb in HBVr during allo-HSCT (70). Twenty two HBV-resolved patients showed different two-year cumulative incidence of HBVr (20% vs 75%) between groups of HBcAb(+)HBsAb(+) and HBcAb(+)HBsAb(-) (70). Certainly, these need to be verified in larger prospective studies. Nearly all studies on HBVr in HBsAg-negative/HBcAb-positive allo-HSCT recipients reported that HBVr was a late phase complication ( Table 2). Additionally, nearly all studies recommended the administration of 6-12 months of antiviral prophylaxis. It is necessary to consider the value of early antiviral prophylactic treatment in preventing late phase complications of HBVr.
However, long-term antiviral treatment may cause resistance. The cumulative incidence of HBV resistance of anti-HBV drugs with a low resistance barrier (LAM, LdT, and ADV) is prevalent and growing over time in patients with chronic hepatitis B (54). LAM resistance occurs in up to 20% of patients after just one year of use (8). One study reported that an HBsAg-negative patient who underwent allo-HSCT using stem cells from an HBsAgpositive donor eventually acquired HBV infection due to a YYMD mutation as a result of long-term prophylactic treatment with LAM (73). One report suggested that HBV reemerges with T127P, F170FL, and S204R mutations with prophylactic LAM treatment, causing HBVr after HSCT (34). Furthermore, recent studies found favorable prognosis of HBVr in HBsAg-negative/HBcAb-positive HSCT recipients (38-42). HBVr in these groups can be controlled, and most HBVr patients acquire serologic clearance of HBsAg with regularly HBVr monitoring and preemptive antiviral treatment (38, 39). After antiviral initiation, HBV-resolved patients with reactivation showed a one year cumulative HBsAg clearance of 68.3% (38). No case of serologic clearance in an HBV-RS patient who recovered from HBVr converted to active HBsAg carriers has been reported. The liver-related mortality of HBVr was nearly zero. In a large prospective study monitoring HBV DNA monthly in HBV-resolved B cell lymphoma patients, no hepatitis due to HBVr was observed in patients who received antiviral treatment when HBV DNA levels were between 11 and 432 IU/ml (7). Another prospective study enrolled 83 HBsAgnegative/HBcAb-positive hematologic patients receiving anti-CD20 therapy. These patients were monitored every 4 weeks without antiviral therapy and every 2 weeks once HBV DNA was detectable. All patients with HBV DNA had RS or two-fold increase in upper limit of normal ALT received antiviral therapy. After therapy, ALT was normalized and HBV DNA returned to undetectable levels. There were no cases of clinical hepatitis, liver failure, or death (74). These data indicate that regular monitoring and preemptive antiviral therapy are effective methods for preventing HBVr-related hepatitis in HBV-resolved patients following immunosuppressive therapy.

DONOR RELATED FACTORS
It was first confirmed in 1995 that HBV can be transmitted via stem cells from HBsAg-positive donors to recipients during HSCT (75). A group at Queen Mary Hospital in Hong Kong pioneered the application of stem cells from HBsAg-positive donors in allo-HSCT (16,76,77). The incidence of HBV-related hepatitis in recipients who receive HBsAg-positive donor stem cells is in the range of 48% to 55.5% (16,76). Therefore, serological HBsAg positivity seems to be a contraindication for HSCT donors, due to the fear of HBV-related hepatitis. Selecting a suitable donor for a HSCT recipient from a pool of potential HBsAg-positive donors is an unresolved problem. The donor's predicted favorable factors for HSCT outcomes may conflict with the status of serologic HBsAg positivity. There are no standard guidelines for managing patients who receive stem cells from HBsAg-positive donors. The Fifth European Conference on Infections in Leukemia indicated that both the donor and recipient undergo antiviral treatment and that HBsAg-negative recipients are vaccinated to prevent HBV transmission (78). These measures lowered the incidence of HBV-related hepatitis to 6.9% of recipients who receive stem cells from HBsAg-positive donors, whereas the historical control group was 48% (16). We

ADOPTIVE IMMUNE TRANSFER
It was reported that donor HBsAb decreased the risk of HBVr in HSCT recipients (40). Univariate and multivariate analyses of HBVr risk factors confirmed the protective role of an HBVimmune/exposed donor (HR adjusted = 0.12, 95% CI 0.02-0.96; P = 0.045) (40). A recent large retrospective study indicated that the cumulative incidence of HBV-RS at 5 years was 16.3% and 8.4% for patients with or without donor anti-HBs, respectively. Multivariate analysis revealed that the independent risk factor for HBV-RS was allo-HSCT from donors lacking anti-HBs compared with other donors with anti-HBs antibodies (HR= 0.294; 95% CI, 0.13-0.85; P = 0.0117) (42). However, we stratified 565 HBsAg-negative donors according to HBcAb and HBsAb status into four groups. The cumulative HBVr rates at 5 years in the four groups were 5.3% for HBcAb(-)HBsAb(-), 5.1% in HBcAb(-)HBsAb(+), 3.8% in HBcAb (+)HBsAb(-), and 1.6% in HBcAb(+)HBsAb(+) (P=0.794). We did not find a protective role for HBV-immune/exposed donors in HSCT recipients (39). In addition, there were few reports of HBsAg clearance in HBsAg-positive patients after allo-HSCT. There were several case reports demonstrating that an HBV-immune/exposed donor with HBsAb can lead to serologic clearance of HBsAg in HBsAg-positive recipients (80,81). The factors influencing HBsAg clearance in HBsAg-positive patients following allo-HSCT are unclear. Additionally, there was no strong evidence that adoptive immune transfer plays a protective role in HBVr during allo-HSCT.

HBV VACCINE ISSUES
The Francophone Society of Bone Marrow Transplantation and Cellular Therapy recommended HBV vaccine for HBV-resolved recipients to prevent HBVr after allo-HSCT (82). A retrospective study from Hokkaido University Hospital enrolled 21 patients with HBV-resolved infection (83). They received a standard three-dose regimen of hepatitis B vaccine after discontinuation of immunosuppressants. The first vaccine was administered at a median of 15 months (range, 6-79 months) after transplantation. Nine of them tested positive for HBsAb. None of the 21 patients in the vaccine group developed HBVr, which indicated that HBV vaccination of HSCT recipients was a promising method for preventing HBVr. However, the following multicenter prospective clinical research of hepatitis B vaccine to prevent HBVr after allo-HSCT failed to find the protective role of hepatitis B vaccine in minimizing the risk of HBVr in HBVresolved recipients (31). Only 37% (10/27) of patients had HBsAb with three doses of hepatitis B vaccine 12 months after HSCT, and the 2-year cumulative incidence of HBVr was 27.3%. Encouragingly, a recent preliminary study showed excellent results with an anti-HBs seroconversion rate of 82% in HBsAg-negative pediatric and young adult recipients after HSCT at the median of 10.4 (range 3.0-22.4) months after the third vaccination (84). Another prospective study enrolled 86 adults that accepted a low dose of the HBV vaccine (10 mg/dose) at 6, 7, 8, 12 months after allo-HSCT. The proportion of recipients achieving anti-HBs antibody titers 100 mUI/ml was 64.6% (95% CI, 53% to 75%; n = 51/79) at 6 months after vaccine initiation and 56.8% (95% CI, 39.5% to 72.9%; n = 21/37) at 24 months after vaccine initiation (85). This study suggested a better efficacy of higher HBV vaccine antigen doses. However, the effectiveness of these vaccines in preventing HBVr remains to be evaluated.

OBI (OCCULT HEPATITIS B INFECTION)
OBI was defined as the presence of replication-competent HBV DNA in the liver and/or HBV DNA in the blood of people who test negative for HBsAg. These patients can be classified as seropositive OBI (HBsAb-positive or HBcAb-positive) and seronegative OBI (HBsAb-negative and HBcAb-negative) (86).
A study found that 19/124 (15.3%) HBsAg-negative donors were detected to have OBI by using the PCR method, for which 14/19 (73.7%) OBI donors were HBsAb-positive (77). Thus, transmission of stem cells from OBI blood donors is a risk factor for HBV-related hepatitis. Patients with OBI have a risk of HBVr when they receive cancer chemotherapy or other immunosuppressive therapies (86). The risk is high (>10%) in OBI patients receiving anti-CD20 containing regimens and myeloablative regimens for HSCT (41, 61,62,87). Considering the probability of OBI, both HSCT recipients and donors should be screened for HBV DNA before HSCT.

NEW THERAPEUTIC TREATMENTS
Several new treatment strategies have emerged for patients after allo-HSCT. These include CAR-T therapy and blinatumomab therapy for relapse, rituximab, ruxolitinib, ibrutinib, and other monoclonal antibodies for chronic GVHD treatment. However, as these strategies target B cells and/or T cells, they may also cause HBVr. Indeed, rituximab is known for resulting in HBVr (59). HBV reactivation has also been reported after CAR-T therapy in patients with current or past HBV infection (88)(89)(90)(91). Cases of HBVr have been associated with ibrutinib treatment for hematological malignancies but not for patients after allo-HSCT (92)(93)(94). Therefore, physicians should carefully monitor the ALT and HBV DNA in chronic HBV infection or resolved HBV infection patients when applying new therapeutic treatments after allo-HSCT.

FUTURE DIRECTIONS
Several unresolved issues remain regarding HBVr during allo-HSCT, which require more work in future studies. The new DNA sequence of HBVr in the process of HBVr needs to be found to redefine the role of virus mutation itself in the mechanism of HBVr. The frequency of monitoring and the duration of NAs prophylactic administration in preventing HBVr during allo-HSCT remains unclear and needs to be established urgently. Many consensuses still recommend LAM as the first-line option. The data regarding the comparison of high barrier (ETV, TDF, or TAF) NAs with low barrier agents (LAM, LdT, ADV) in the prophylaxis of HBVr in HSCT is limited. The frequency of developing drug resistance during long-term antiviral treatment, especially for low barrier agents, in HSCT is unclear. These data may change the first-line recommendation of prophylactic antiviral treatment for HBVr in the future. Prophylactic antiviral therapy for HBV-resolved patients is controversial and large-sample prospective studies are needed to investigate the incidence of HBVr in these patients following allo-HSCT. The protective role of HBsAb in HBVr during allo-HSCT and the adoptive immune effect of donors for decreasing HBVr are awaiting further exploration. Regularly monitoring of ALT and HBV DNA and preemptive antiviral treatment in HBV-resolved patients need to be verified. We believe that HBsAb-positive patients may be the most likely stratified group of resolved HBV infection patients who need not accept prophylactic antiviral therapy for preventing HBVr. The dosages and schedule of vaccines in preventing HBVr in HBV-resolved recipients remains to be evaluated in prospective trials. The clinical influence of HBVr during allo-HSCT requires further investigation, such as the interruption of immunosuppressants, severe hepatitis, liverrelated mortality, and non-relapse mortality. We portray an agenda for future further research in HBVr during allo-HSCT ( Table 3).

ACKNOWLEDGMENTS
We would like to thank Editage (www.editage.com) for English language editing.

Group Further research Purpose
HBsAg-positive recipients The frequency of monitoring and the duration of NAs prophylactic administration Establish antiviral therapy duration and monitored time intervals Comparison effectiveness and resistance of high barrier drugs and low barrier agents in the prophylaxis of HBVr Promote to first-line recommendation of prophylactic antiviral treatment with high barrier agents HBsAg clearance in HBsAg-positive patients after allo-HSCT accepting stem cells from HBsAb-positive donors To verify the adoptive immune role and the influence factors of HBsAg Seroclearance New therapeutic treatments (e.g. ruxolitinib, ibrutinib, monoclonal antibodies) To investigate the latent risk of HBVr during treatments HBsAg-negative/HBcAbpositive recipients Prospective studies to reflect the real cumulative rate of HBVr without prophylactic treatment To investigate the incidence of HBVr in HBV-resolved patients undergoing allo-HSCT Prospective studies to investigate the protective role of HBsAb (both recipients and donors) To explore the protective role of HBsAb in HBVr The frequency of monitoring and the duration of NAs prophylactic administration Establish antiviral therapy duration and monitored time intervals HBV vaccine dosage and schedule for minimizing the risk of HBVr Explore and establish valid method of preventing HBVr with vaccine method New therapeutic treatments (e.g. ruxolitinib, rituximab, ibrutinib, monoclonal antibody)  (5)