Adoptive cellular therapy (ACT) based on T cells has been extensively studied (53). However, the ideal strategy should appropriately stimulate both cellular immunity and humoral immunity. Indeed, activated B cells also have the potential to kill tumor cells via antibody-dependent and -independent mechanisms (Figure 1) (54–59), which provides the possibility for ACT based on B cells. Our laboratory previously reported the tumor-killing abilities of B cells primed in vivo and activated in vitro (56–59). These activated B cells were adoptively transferred into tumor-implanted mice, leading to significant tumor regression in primary subcutaneous tumors as well as metastatic pulmonary tumors (56). This phenomenon has been confirmed in three tumor histologies, including breast cancer, melanoma, and sarcoma (56–59). In addition to producing tumor-specific antibodies that killed tumor cells via complement-dependent cytotoxicity, activated B cells harvested from tumor-draining lymph nodes (TDLN) were able to induce tumor-cell lysis in an antibody-independent way (58, 59). Of note, combined adoptive transfers of activated TDLN B cells and T cells resulted in stronger antitumor responses than the transfer of either cell population alone (56, 57).

The potential mechanisms of B cell-mediated direct killing effect appear to be multiple. Tao et al. found that effector B cells can directly kill tumor cells via the Fas/FasL pathway, and was inhibited by IL-10 (58). In that study, IL-10^-/- B cells mediated a stronger antitumor effect compared to wild-type B cells, which was verified by IL-10 neutralization utilizing antibody (58). Furthermore, B cell killing of tumor cells was impaired by anti-FasL antibody in a dose-dependent manner (58). In a separate study, Xia et al. defined additional mechanisms involved in cancer ACT based on activated TDLN B cells (59). In that study, IL-2 significantly amplified the therapeutic activity of transferred effector B cells (59). Additional in vitro experiments demonstrated that activated B cells provoked direct cytotoxic action on tumor cells through CXCR4/CXCL12 pathways, while without cell contact, B cell-secreted perforin also led to tumor cell cytotoxicity (59).

Additional publications support the fact that B cells can act as effector cells with tumor-killing capacity. Kemp et al. demonstrated that the stimulation of human peripheral blood mononuclear cells with CpG-A oligodeoxynucleotide (ODN) led to high levels of functional B cells, which mediated tumor cell lysis via inducing expressions of functional TRAIL/Apo-2 ligand on B cells (60). Penafuerte and colleagues reported that activation of B cells with a chimeric protein consisting of the ectodomain of TGF-β receptor II and IL-2 resulted in effector B cells that induced potent antitumor immunity (61). In addition, it was found that CD73 inhibition elicited sustained B cell-mediated tumor regression in melanoma-bearing mice (62). Together, these studies suggested that with multiple cellular and molecular events, B effector cells can mediate therapeutic antitumor function via direct killing of the tumor cells.

B cells may function as antigen-presenting cells (APCs) to engage with T cells in situ, and thereby trigger anti-cancer responses (Figure 1). It has been reported that human B cells can efficiently present peptides to CD4⁺ T cells after being activated by CD40 ligand and pulsed with tumor antigen (63). In agreement with this finding, we have reported that activated B cells administered in the ACT will confer host T cell immunity via its antigen-presenting function utilizing a murine congenic model where donor vs. host cells was tracked (57). Intriguingly, Colluru and colleagues successfully applied B cells as a tool to present tumor DNA to CD8⁺ T cells as a vaccine, which elicited an antitumor effect in vivo (64). In another report, B cells were reported to infiltrate into the brain tumor site, where they promoted T cell-induced tumor killing as APCs (65).

Bruno et al. observed that TIL-Bs isolated from human NSCLC tumors efficiently presented antigens to CD4⁺ TILs (33). Interestingly, two subgroups of TIL-Bs with opposite functions were described as activated (CD19⁺CD69⁺CD27⁺CD21⁺) or exhausted (CD19⁺CD69⁺CD27⁻CD21⁻) TIL-Bs (33), which induced Th1 phenotype CD4⁺IFN-γ⁺ or Treg phenotype CD4⁺Foxp3⁺ cells, respectively (33). In addition, B cells from doxorubicin-treated cancer patients were found to have enhanced expression of CD86 with improved APC ability (66).

A recent study reported by Jiang et al. demonstrated that CD19⁺ TIL-Bs expressing APC-characteristic markers were found to be co-localized with activated CD4⁺ TIL-Ts in human bladder cancer and was associated with improved survival (67). Using an antigen-presentation assay in this study, TIL-Bs markedly promoted proliferation and activation of CD4⁺ TIL-Ts, which was completely abrogated by HLA-DR blockade (67). In breast tumor-bearing mice, ICB-treated tumors presented a striking elevation in APC B cells (10). When B cells were depleted by anti-CD20 or anti-CD19 antibodies, the effector memory and central memory subsets of CD4⁺ T cells, as well as the effector and effector memory subsets of CD8⁺ T cells, were remarkably decreased (10). More recently, the abundance and spatial distribution of CD86⁺ antigen-presenting B cells have been described in tumor samples of cancer patients, indicating that they are elevated in TILs, localized in TLS, and enriched in tumors with increased TLSs (68). In vitro study further revealed that the specific B cell subpopulation can trigger T cell-mediated immune responses, confirming that they are professional APCs in tumor immunity (68).

Taken together, these findings suggested that TIL-Bs can serve as APCs to mediate antitumor responses, providing a better chance of B cells in developing additional immunotherapeutic strategies.

Multiple lines of evidence have demonstrated that B cells act as an antitumor player via antibody-dependent cytotoxicity to tumor cells. Reports have established that B cells within tumor-related TLSs can convert to antibody-producing memory B cells or plasma cells after exposure to tumor-associated antigens (Figure 1) (19, 69). Thus, TIL-Bs usually serve as an efficient source of tumor-related antibodies, which can be tested in the serum and tumor tissue from cancer patients (69–71). Importantly, detection of serum autoantibodies has been proposed as early clues for human malignancies (72). Additionally, increased tumor-infiltrating plasma cells have been reported to be correlated with better prognosis in ovarian cancer (73), breast cancer (74), colorectal cancer (74), NSCLC (74), and gastric cancer (75), which further support the antitumor property of tumor-targeting antibodies.

Indeed, tumor-infiltrating plasma cells are able to produce oligoclonal, somatically mutated antibodies with the capacity to bind tumoral antigens, provoking the specific cytotoxicity to tumor cells. Our preclinical studies showed that immunoglobulin G (IgG) harvested from tumor-bearing mice bound selectively to tumor cells, triggering tumor cell lysis via complement-dependent cytotoxicity (CDC) (57–59). More recently, we further demonstrated that the immunotherapies targeting integrin β4 induced host immunity against cancer stem cells via specific CDC (76). In a separate study, Carmi et al. revealed that allogeneic tumor rejection was mediated in the murine model at the early stage of tumor development by naturally occurring tumor-specific IgG antibodies, which prompted DCs to internalize tumor antigens and subsequently activate T cells, initiating a cytotoxic effect to control tumor growth (77). More recently, Hollern reported that tumor-associated IgG was essential in mediating the response to ICBs (10). In that study, ICB-induced B cell activation resulted in the generation of class-switched plasma cells and increased tumor-specific serum IgG (10). Both the loss of antibody secretion and neutralization of FcR during ICB therapy diminished the initial antitumor response in breast tumor-bearing mice (10). This work for the first time identified a crucial role for antibody secretion in the function of ICB therapy.

Very recent studies have shown that intratumoral B cell-generated autoantibodies induce tumor cell apoptosis via targeting proteins on the surface of tumor cells (12, 13). Mazor et al. have reported that tumor cells derived from patients of high-grade serous ovarian carcinoma were frequently coated with IgGs produced by intratumoral antibody-secreting cells (12). These tumor-reactive antibodies can target MMP14 on the surface of tumor cells and induce NK cell-mediated antibody-dependent cellular cytotoxicity (ADCC) (12). Another study has also shown that high levels of IgG-producing plasma cells and IgG-coated and apoptotic tumor cells coexist in TLS⁺ tumors, suggesting antibody-mediated anti-tumor immunity (13). These findings provide the direct proof of tumor-killing activity of autoantibodies generated by B/plasma cells.

Collectively, these findings strengthened the premise that humoral responses can have potent antitumor activities via tumor-specific antibody-facilitated tumor cell lysis.

Bregs, first reported by Morris (80), are special subsets of B cells with immune-suppressing properties for maintaining immune tolerance. Recently, a new wave of studies has emerged on tumor-infiltrating Bregs, which appears to be disadvantageous to cancer patients’ outcomes. In breast cancer, the co-localization of Bregs and Tregs was associated with shorter metastasis-free survival (52). Additionally, high infiltration of programmed cell death-1 (PD-1)^hi Bregs presenting in advanced-stage HCC, correlated with early recurrence (78). Similarly, programmed death-ligand 1 (PD-L1)⁺ Bregs were found to be enriched in human metastatic melanoma tumors compared to primary lesions (79).

Within various secondary lymphoid tissues, naïve B cells are activated once exposed to antigens, and then proliferate and form GCs, where they receive signals to grow and differentiate into memory B cells and plasma cells. As a subpopulation of B cells, Bregs have been shown to arise at multiple stages during B cell development (81). Typically, the phenotypes in human tumors are enriched within the CD38⁺ transitional (78, 82–85) and CD27⁺ memory B cell population (Table 1) (86, 88, 90). In addition, a novel type of Bregs identified by Shalapour et al. share the phenotypes with IgA⁺CD138⁺ plasma cells (Figure 1) (91). Meanwhile, the precursors of several Bregs (e.g., adenosine-producing B cells) remained to be clarified (92). Also, some Bregs were characterized by functional molecules, such as PD-1 (78), PD-L1 (79, 88), T-cell immunoglobulin and mucin domain-1 (TIM-1) (83, 93), granzyme B (85), and CD5 (89). Together, it is thought that all B cell subpopulations can obtain an inhibitory property with appropriate stimuli (81), making their descriptions even harder due to multiple subtypes.

Despite the confusing distinct phenotypes, Bregs can be functionally defined by their capacities for the generation of immunosuppressive cytokines, including IL-10 (86), IL-35 (82), and TGF-β (Figure 1) (84). For example, a protumorigenic PD-1^hi Breg subset in human HCC exhibited a unique CD5^hiCD24^−/+CD27^hi/+CD38^dim phenotype, which caused T-cell dysfunction and fostered tumor progression via IL-10-dependent pathways (78). In another study, a granzyme B⁺ Bregs with a CD19⁺CD38⁺CD1d⁺IgM⁺CD147⁺ expression signature was found in various human solid tumors (85). They can be induced by IL-21 and generate high amounts of IL-10 (Figure 1) (85). Similarly in HCC, the intratumoral TIM-1⁺ Bregs with a CD5^hiCD24^-CD27^-/+CD38^+/hi phenotype generated high levels of IL-10 and exerted robust repressive functions against CD8⁺ T cells (83). Another type of IL-10-secreting Bregs (B10) was characterized by a CD19⁺CD24^hiCD38^hi phenotype in gastric cancer (84). Besides, IL-35-producing CD1d^hiCD5⁺ Bregs accumulated in pancreatic cancer and supported tumor cell growth, underlining the potential value of IL-35 as a therapeutic target (87).

Several studies report that Bregs not only directly fostered tumor growth, but also acted on other immune compartments in the TME. Mostly, Bregs negatively regulate the antitumor immune responses of T cells. Among different types of Bregs, B10 has been shown to suppress the secretion of proinflammatory cytokines by CD4⁺ Th cells and induce CD4⁺FoxP3⁺ Tregs (52, 84, 86). Furthermore, an in vitro study demonstrated that B10 inhibited the proliferation and activation of T cells (52, 84, 86). IL-35-producing Bregs induced pSTAT3⁺CXCR3^-CD8⁺ T cells in pancreatic cancer (87). Additionally, Bregs promote tumor progression via interplay with myeloid-derived suppressor cells, tumor-associated macrophages, and natural killer cells (Figure 1).

However, the immunosuppressive characteristics and mechanisms of tumor-infiltrating Bregs are still largely unclear, which remains a central focus of investigation.

In addition to Bregs, local or systemic tumor-associated autoantibodies may also exert a protumoral effect through initiation and maintenance of inflammation (91), tissue remodeling (94), and angiogenesis (94). Clinical cohort studies exhibited a close association of serum p53 antibody with poor prognosis in breast cancer patients (95). Other serum antibodies were also reported to correlate with poor survival in ovarian and pancreatic cancer (96). Generally, elevated immunoglobulins (97) and high IgG3-1 switches (98) have been identified as unfavorable prognostic biomarkers in renal cell carcinoma. These data from a clinical perspective suggest that some tumor-specific antibodies might promote tumor processes.

It is important to note that antibody functionality, protumor or antitumor, may be determined by the antibody isotype. Remarkably, a study using TCGA RNA-seq data indicated the association of high fractions of IgA, IgD, or IgE with a poor prognosis in human melanoma (99). A low proportion of intratumorally produced IgA was specifically correlated with improved overall survival for KRAS mutation lung adenocarcinoma (71). Karagiannis et al. reported that antigen-specific and nonspecific IgG4 hampered IgG1-induced antitumor activities through the deactivation of FcγRI, which explained the inverse correlation between serum IgG4 and patient survival (100). Additionally, RNA-seq data have shown that the relative proportions of IgG3, in contrast to IgG1, was either neutrally or negatively associated with outcome in melanoma (99).

In summary, existing evidence indicates that some phenotypes of immunoglobulin are associated with worse outcomes in cancer patients. Animal studies also illustrate that tumor-specific antibodies can promote tumor development by dismantling antitumor immunity. However, the mechanisms underlying antibody-mediated protumor properties have not yet been fully investigated. More work needs to be conducted in this field to elucidate the obstacles to cancer immunotherapy.

Previous studies in our laboratory and others have demonstrated that activated B cells can specifically and directly kill tumor cells (16, 58, 59), supporting the potential of adoptive B cell therapies. It has been well established that adoptive T cell therapy, such as CAR-T cell therapy (2), is capable of inducing high clinical response rates in hematological malignant disorders (53), whereas attempts to treat patients with solid tumors using CAR-T cells are still in their infancy. Preclinical studies have shown that the co-transfer of activated T cells and B cells led to a greater extent of tumor regression than either cell population alone (56, 57), suggesting effector B cells can function as a valuable adjunct in T cell-based immunotherapy. Moreover, adoptive transfers of other types of B cells, like TIM-4⁺ B cells (101), GIFT4-programmed B cells (102), and CpG-ODN activated B cells (103), also have capacities to elicit tumor regression. These studies offer novel cancer immunotherapeutic approaches based on B cells.

Resting B cells with a poor capacity of antigen presentation can be activated by diverse stimuli to obtain immunostimulatory ability. CD40 signals are the strongest inducer of antigen-presenting B cells to trigger potent T-cell immunity (104). CD40-activated B (CD40-B) cells are thought to be an encouraging alternative to DCs as skilled APCs for antigen-specific cancer immunotherapy (105). In contrast to DCs, a greater number of B cells can be easily harvested from human peripheral blood after in vitro expansion in the presence of CD40L, without the loss of APC functionality (106). CD40 signal activation markedly enhances the antigen-presenting ability of B cells, powerfully inducing vigorous expansion of antigen-specific T cells and homing to the secondary lymphoid organs (55, 105). Importantly, tumor-implanted mice were preventively immunized by tumor antigen-primed CD40-B cells, resulting in a protective antitumor response against melanoma and lymphoma (105). In summary, CD40-B cells emerge as an attractive tool to develop additional immunotherapeutic agents. The design of clinical trials using CD40-B cells to confirm this potential antitumor function seems worthy.

Effector B cells also can be induced or activated by cytokines or chemotherapy drugs, which can be utilized to enhance B-cell-mediated antitumor responses. Li and colleagues found that IL-2 augmented the antitumor responses of adoptively transferred TDLN B cells (59). A separate study reported that IL-17A improved the tumor-killing activity of B cells in esophageal squamous cancer (16). In addition, Lu et al. demonstrated that chemotherapy in breast cancer patients induced a novel ICOSL⁺ B cell subset with antitumor capacity by enhancing the effector T cells to Tregs ratio (9). Similarly, doxorubicin can induce CD86 expression on the surface of B cells to elevate their antigen-presenting capacity in urinary bladder cancer patients (66). Thus, conventional drugs, as well as cytokines, could be a favorable option utilized to induce effector B cells and improve the antitumor efficacy of immunotherapy involved with B cells.

Since several lines of evidence indicate that some subpopulations of B cells possess tumor-promoting activities, the anti-CD20 antibody (rituximab) has been used to deplete the total B cell population for the treatment of solid tumors in small cohorts of patients with melanoma (107) and advanced colon cancer (108). Although it was difficult to estimate the therapeutic value from such small cohorts of patients, it seemed that B cell-depletion therapy was not associated with significant toxicity (107, 108). In another study, rituximab therapy combined with low-dose IL-2 in patients with melanoma and renal cell carcinoma did not show any clear benefit of the B cell depletion (109). Of note, a recent study demonstrated that B-cell depleting or inhibiting treatment in mammary tumor-bearing mice significantly weakened the antitumor immunity of combined ICB immunotherapy (10). In addition, most of the cohort studies revealed the positive association of TIL-Bs with better outcomes in various solid tumors (14). Therefore, more precise immunotherapies targeting specific B cell subsets (e.g., Bregs) instead of the total B cell population would be more rational for further investigations.

Indeed, strategies targeting Bregs have been reported to be effective in murine tumor models. For example, Li et al. demonstrated that IL-10 deletion or blockade notably improved the therapeutic effect of TDLN B cells in an ACT model (58). Additionally, a recombinant plasmid pcCD19scFv-IL10R containing the gene of the extracellular domain of IL-10R1 and anti-CD19 single-chain variable fragment significantly decreased B10 cells and Tregs, which may offer a new immunotherapeutic strategy targeting Bregs (110). Interestingly, both conventional chemotherapy (66) and novel molecular targeted therapy (111) were reported to serve as Breg inhibitors via the downregulation of IL-10 secretion. IL-35, another critical immunosuppressing factor derived from Bregs, has been proved to be a potential immunotherapy target. Mirlekar reported that specific deletion of IL-35 in B cells overcame immunotherapy resistance to anti-PD-1 through activation of CD8+ T cells in murine pancreatic cancer model (80).

In short, B cell-based immunotherapy should consider both antitumor and protumor functions of B cells. Therefore, the basic strategy is to precisely target specific B cell subsets by activating/transferring antitumor effector B cells or suppressing/eliminating protumor inhibitory B cells.