Advancing interleukin-21 in cancer immunotherapy: Mechanisms of action and clinical translation

Abstract

Interleukin-21 (IL-21) is a pleiotropic cytokine that coordinates both adaptive and innate immune responses. It enhances the cytotoxic activity of CD8+ T cells and natural killer cells, regulates the suppressive function of regulatory T cells, and promotes B cell-mediated antibody production, thereby showing therapeutic potential in antitumor immunity. Current evidence suggests that IL-21 can postpone or reverse T cell depletion by downregulating inhibitory receptors such as programmed cell death protein 1 and T cell immunoglobulin and mucin domain-containing protein 3 and reducing levels of the transcription factor TOX. This implies that combining IL-21 with immune checkpoint inhibitors or chimeric antigen receptor T cell therapies may produce synergistic effects. IL-21 encounters significant clinical translation challenges, particularly its limited in vivo half-life and systemic toxicity that constrain dosing. Researchers are exploring sophisticated engineering approaches to mitigate these constraints. Strategies include developing prolonged-activity IL-21 variants or tumor-targeted formulations through protein engineering, alongside creating advanced delivery systems that minimize biodistribution and reduce unintended effects. This review synthesizes IL-21’s biological roles and molecular pathways in anti-tumor immunity, examines developments in combining IL-21 with both conventional and novel immunotherapeutic approaches, and assesses current barriers to clinical translation. We then propose innovative molecular engineering and delivery mechanisms that could potentially enhance safety and efficacy of IL-21-based combination therapies in oncology.

Key Words: Interleukin-21; Anti-tumor immunity; Combination therapy; Oncology; Cancer immunotherapy; Clinical translation

Core Tip: Interleukin-21 (IL-21) mediates dual antitumor effects by enhancing effector immunity and reducing immunosuppression, primarily through the Janus kinase-signal transducer and activator of transcription 3 pathway, which regulates multiple immune cell types. It has also shown synergistic effects in combination with various cancer therapies, including immune checkpoint inhibitors and adoptive cell therapy. However, clinical translation has been hindered by its short half-life and dose-limiting systemic toxicity. This review summarizes the regulatory mechanisms of IL-21, progress in combination regimens, challenges in clinical development, and recent strategies to optimize its therapeutic use. Our goal is to provide a theoretical basis for developing safer and more effective IL-21-based combination immunotherapies for cancer.

INTRODUCTION

Over the last decade, cancer immunotherapy has experienced a revolution. Immune checkpoint inhibitors (ICIs), such as anti-programmed cell death protein 1 (PD-1)/programmed death ligand 1 (PD-L1) antibodies, have led to improved clinical outcomes in many cancers[1,2]. However, durable responses (complete response/partial response > 24 months) to ICI therapy is only 15%-30% to absent adult patients with melanoma or renal cell carcinoma treated with first-line PD-1-based regimens. Primary resistance is prevalent in PD-L1 negative tumors, whereas acquired resistance develops in 20%-30% of initial responders within 2 years[3,4]. In addition, grade 3-4 immune-related adverse events (irAEs) are observed in 40%-60% of patients treated with dual ICIs [anti-PD-1 + anti-cytotoxic T lymphocyte-associated protein 4 (CTLA-4)] vs 10%-20% with anti-PD-1 monotherapy across several phase III trials. Organ-specific irAEs include colitis, pneumonitis, and endocrinopathies; most severe cases are treated with corticosteroids. Treatment discontinuation due to irAEs occurs more frequently with combination regimens than with monotherapy and fatal irAEs are reported in < 1% patients. Another hurdle is the immunosuppressive TME containing infiltrating immunosuppressive cells and functionally exhausted effector lymphocytes[5]. These challenges underscore the necessity of developing rational combination strategies to enhance both the potency and safety of cancer immunotherapies.

Interleukin-21 (IL-21) is a type of I cytokine. It is mostly secreted by the subsets of activated CD4+ T cells especially T follicular helper cells (Tfh) and T helper 17 cells (Th17). Natural killer T (NKT) cells produce it as well. The receptor complex of this cytokine is expressed through a receptor complex; this is one with the alpha chain of the IL-21 receptor (IL-21R) and the common gamma chain. It can be written in some instances as γ-c[6-8]. IL-21 is a pleiotropic immune modulator. What that means is that it influences the development of various immune cells, differentiation, proliferation, and function of various immune cell types, including T cells, B cells, and natural killer (NK) cells[9]. By enhancing the cytotoxicity of effector T cells and NK cells, promoting antibody production by B cells, and modulating regulatory T cell (Treg) activity, IL-21 plays a pivotal role in both antiviral and antitumor immunity[10]. This is why IL-21 seems to have a very promising future in cancer immunotherapy because it has such properties.

Nevertheless, the clinical application of native IL-21 is limited by its pharmacokinetic properties and safety profile. For example, it has a brief half-life in the body and when administered, it causes systemic toxicity. The recombinant human IL-21 had a terminal half-life of approximately 3.0 hours after a 30 μg/kg intravenous infusion (NN028-1614) was given. In clinical trials, the maximum tolerated dose (MTD) of intravenous rhIL-21 was found to be 30 μg/kg per day over 5 days, followed by a 9-day period of rest. Grade 3 lab problems, such as high transaminases, neutropenia and thrombocytopenia, were toxicities that limited the dose[11]. Consistent results were reported in NCT00095108, 62% of patients had adverse events that were grade 2, and 12% had grade 3 adverse events, most often patients experienced fever and fatigue[12]. The whole-body toxicity usually manifests itself as symptoms like the flu, including grade 1-2 fever, chills, and fatigue. Such issues are generally not permanent and disappear within a few days following the administration of the dose (Table 1).

Table 1 Summary of clinical trials evaluating interleukin-21 in cancer.

Cancer type	Phase/status	Combination/intervention	Registration number
Malignant melanoma	Phase II	IL-21	NCT00514085; NCT00336986
Melanoma skin cancer	Phase II	Recombinant IL-21	NCT00601861
Metastatic or recurrent melanoma	Phase II	IL-21	NCT01152788
Ovarian cancer	Phase II	Recombinant IL-21; Caelyx	NCT00523380
Pancreatic cancer	Phase II	Recombinant human IL-21 expressing oncolytic vaccinia virus injection (hV01)	NCT07006077
Metastatic renal cell carcinoma	Phase I/II	Recombinant IL-21; sorafenib	NCT00389285
Renal cell carcinoma	Phase I/IIa	Recombinant human IL-21; sunitinib	NCT00617253
Advanced cancer	Phase I	Recombinant fusion protein of IL-21 and humanized anti-human serum albumin VHH antibody	NCT05296772
GPC3-positive solid tumors; liver cancer; ATRT and CNS rhabdoid tumors	Phase I	IL-15 and IL-21 armored GPC3-CAR T cells	NCT06198296; NCT04093648; NCT07513194
Melanoma	Phase I	Recombinant IL-21; ipilimumab	NCT01489059
Metastatic malignant melanoma and metastatic kidney cancer	Phase I	Recombinant IL-21	NCT00095108
Metastatic melanoma	Phase I	Recombinant IL-21	NN028-1614
Non-Hodgkin’s lymphoma	Phase I	Recombinant IL-21; rituxan	NCT00347971
Solid tumors	Phase I	IL-21; anti-PD-1	NCT01629758

IL-21: Interleukin-21; VHH: Variable domain of the llama heavy chain of heavy-chain antibody; ATRT: Atypical teratoid rhabdoid tumor; CNS: Central nervous system; CAR: Chimeric antigen receptor; PD-1: Programmed cell death protein 1.

To overcome these obstacles, many studies have also explored the potential approaches. For example, protein modification has been attempted in order to obtain a better or more durable IL-21 product with improved antitumor activity. Alternatively, delivery systems that maintain IL-21 within the tumor microenvironment (TME) have also been developed, as this can increase its concentration at the site of action and reduce systemic toxicity[13]. Finally, it has been shown that combining IL-21 with other immunotherapies, including ICIs, adoptive T-cell therapy [e.g., chimeric antigen receptor (CAR)-T cells] or oncolytic viruses, can enhance its antitumor effect and may overcome some of the limitations of using IL-21 alone[14].

Literature retrieval strategy

To search information, literature search was conducted primarily through PubMed using keyword combinations including ICIs (such as “PD-1”, “PD-L1”, “CTLA-4”), and we looked up IL-21-based strategies using pharmacokinetics, half-life, cytokine engineering, CAR-T, oncolytic virus, and finally combination therapy. The search included papers published from January 2000 up to December 2025. We only took peer-reviewed original articles, clinical trials from phase I to III, systematic reviews, and well-regarded reviews. We did not use conference abstracts, papers not in English, or any study not related to cancer immunotherapy. After going through them, 86 articles were included in what we are reviewing here.

This review systematically explores the biological functions and mechanistic foundations of IL-21 in antitumor immunity. It is especially interested in its influence on T cells, NK cells, B cells, and Treg. It then highlights recent advances in combination strategies integrating IL-21 with various immunotherapies, elucidating the underlying mechanisms of observed synergy. Finally, this review analyzes the key challenges in the clinical translation of IL-21, including the half-life and systemic toxicity of the drug, and future trends are proposed based on the promise of protein engineering, specific delivery and rational design of combinations.

THE BIOLOGICAL FUNCTIONS AND MECHANISMS OF IL-21

IL-21 is a cytokine that is pleiotropic and controls both innate and adaptive immunity. Its wide range of activity indicates cell-type-specific effects on immune subsets. Furthermore IL-21 also acts on immunosuppressive cells in the TME[15]. The enhancement of effector immune responses and suppression of suppressive pathways by IL-21 offers an obvious biological foundation to develop therapies in cancer immunotherapy.

The role and mechanism of IL-21 in T cell function

T cells are pivotal in adaptive antitumor immunity. IL-21 regulates various T cell subsets, including CD8+ cytotoxic T cell, CD4+ helper T cell and Tregs. It increases the activity of effector T cells in antitumor activities, suppresses the role of Treg cells in suppression and enhances the development of memory T cells to help build stronger immunity against tumors[9].

In CD8+ T cells, IL-21 cooperates with the other cytokines including IL-7 and IL-15 to control the growth, survival and effector activities of the memory T cell populations[16]. Mechanistically, IL-21 activates CD8+ T cells via the signal transducer and activator of transcription 3 (STAT3) signaling pathway, promoting the differentiation of naïve CD8+ T cells into central memory T cells and supporting the persistence of effector phenotype cells[17]. In antiviral immunity, IL-21 promotes the proliferation of virus-specific CD8+ T cells and downregulating TRAIL expression, enhancing their survival and contributing to memory CD8+ T cell formation[18]. Compared to IL-2 or IL-15, IL-21-stimulated CD8+ T cells exhibit superior cytotoxicity and effector cytokine secretion in vitro. This can be attributed to the capability of IL-21 to stimulate granzyme B expression but not proliferative activity[19].

IL-21 is also involved in the differentiation and the action of various types of helper T cells in CD4+ T cells. It is an autocrine Th17 inducer, which induces Th17 differentiation under the influence of IL-6 by activating STAT3, but inhibits Foxp3[20]. Th17 cells exhibit a dual role in tumor immunity, they may promote angiogenesis and tumor growth, while in other contexts, they mediate potent antitumor immune responses[21,22]. In addition to Th17 cells, IL-21 can be also used to control the activity of Tfh cells that are associated with the downstream effects on the germinal center responses and antibody production by B cells[23,24].

IL-21 is a powerful suppressor of the development and homeostasis of Tregs. Experimental studies indicate that IL-21 neutralization or absence promotes Treg expansion, while IL-21 overexpression reduces both the function and number of Tregs, diminishing their infiltration and activity within tumor tissues. IL-21 directly downregulates Foxp3, the master transcription factor of Tregs, and reducing their immunosuppressive function by inhibiting glycolytic metabolism[25]. Such effects will release the effector T cells from being restrained and decrease the immunosuppressive state of the TME. These results demonstrate that IL-21 plays a critical role in immunity against tumors. It enhances immune cell response of effectors and restricts some significant suppressive cells.

The role and mechanism of IL-21 in NK cell function

NK cells are the main effector of innate antitumor immunity, can directly lyse tumor targets and coordinate subsequent immune responses. IL-21 has a context-specific effect on NK cells. It will be able to significantly increase cytotoxicity of anti-tumors but in large doses or long-term exposure, it can cause apoptosis of NK cells. This response shows the significance of dosing and scheduling when using IL-21 in clinical practice.

IL-21 is a cytokine that enhances the activity of NK cells to kill tumor. IL-21 has been demonstrated to be capable of eliciting antitumor immunity in various preclinical murine models of tumors and the main effect of this process is attributed to the killing ability of the NK cell[26]. The IL-21 also activates the production of pro-inflammatory cytokines by the NK cells, including interferon-gamma (IFN-γ) and tumor necrosis factor-alpha (TNF-α), which will increase the antitumor immune response. IFN-γ can directly suppress the growth and spreading of tumor cells. Despite its potential to block the maturation of dendritic cells (DCs), it may enhance the antigen presentation of NK cells and therefore have an indirect effect on adaptive antitumor responses as well as enhancing the interaction between innate and adaptive immunity[26]. IL-21 also facilitates the differentiation of NK cells into memory-like phenotype with enhanced persistence and increased recall cytotoxicity upon rechallenge with the antigen[27]. In comparison, constant or high levels of IL-21 stimulation cause apoptosis of NK cells and decreased survival of NK cells in vivo that can result in their functional exhaustion and poor antitumor effects in the case of prolonged therapy[28].

The molecular mechanisms underlying these opposing effects converge on the Janus kinase (JAK)-STAT3 signaling pathway. When IL-21 binds to its receptor complex (IL-21Rα/γc), JAK-STAT3 signaling was activated, and then activating receptors upregulates and facilitate the synthesis of cytotoxic mediators like perforin and granzyme B[29,30]. In addition to activation of pro-inflammatory cytokines, including IFN-γ, memory-related genes like Kruppel-like factor 2 (Klf2) and C-X-C motif chemokine receptor are also induced by the activation of STAT3 which promotes the formation of memory-like NK cells[31-33]. With a long-term stimulation however, this same process can be stimulated in the form of negative feedback where the amount of FasL and BIM increases, as well as the decrease of anti-apoptotic protein Bcl-2, finally causing the apoptosis of the NK cell[34].

The role and mechanism of IL-21 in B cell function

B cells mediate humoral immunity through antigen-specific antibody production, and this process is strongly regulated by IL-21. IL-21 fine-tunes B cell responses by promoting proliferation driven by T cell-dependent signals while suppressing proliferation induced by T cell-independent stimuli, thereby favoring antigen-specific B cell expansion[35,36]. It is a key regulator of terminal B cell differentiation, driving class switch recombination and plasma cell generation. IL-21 particularly enhances the production of immunoglobulin G (IgG) and IgA, which contribute to antitumor effector functions[30].

Antibodies generated under the influence of IL-21 can promote tumor cell elimination through antibody-dependent cellular cytotoxicity, complement-dependent cytotoxicity, neutralization of tumor antigens that support proliferation and metastasis, and opsonization that facilitates macrophage-mediated phagocytosis[37]. However, the role of B cells in the TME is complex and context-dependent. Besides contributing to the development of antitumor humoral immunity, certain subsets of B cells can be involved in promoting tumor growth through pro-angiogenic antibody production or even immunosuppressive cell networks[38]. Therefore, the overall impact of IL-21 on B cell function must be assessed in the context of the specific immunological environment of each tumor type.

The IL-21-induced control of B cells is based on the interaction between Tfh cells and B cell surface, such as IL-21R, CD40[39]. The process is centered around the binding of IL-21 to IL-21Rα/γ heterodimer and subsequent activation of JAK-STAT3 signaling[40]. It is a pathway that cooperates with phosphatidylinositol 3-kinase (PI3K)-protein kinase B (Akt) signaling which is induced by the ligation of CD40 to form an interactive network in the fate decision of the B cells[41]. Activated STAT3 translocates to the nucleus, upregulating key transcription factors for plasma cell differentiation, such as Blimp-1 and X-box binding protein 1 (XBP1)[41,42]. Another function of STAT3 is also antibody class switch recombination through its binding to switch regions in immunoglobulin heavy-chain locus, where it facilitates IgM switching to IgG or IgA[43]. Concurrently, PI3K-Akt signaling, which is triggered by the action of CD40, enhances the levels of the anti-apoptotic protein Bcl-2, thus enhancing the survival and proliferation of the B cells, and induces Pax5, which leads to the production of memory B cells and long-term immunity against humoral diseases[42].

Summary of the mechanisms by which IL-21 regulates anti-tumor immunity

The actions of IL-21 on the immune lineages are primarily through binding to the IL-21Rα/γ heterodimer and activating the JAK-STAT3 pathway. The cell-specific effects are mediated by downstream targets of STAT3, including TOX and perforin in CD8+ T cells, NK receptor group 2 member D and Klf2 in NK cells, Blimp-1 and XBP1 in B cells, and their interactions with other signaling pathways.

IL-21 is a system-level effect on the antitumor immunity, and it contributes to the effector activity and suppression of immunosuppression. On one hand, through JAK-STAT3 activation, it amplifies cellular cytotoxicity (in CD8+ T and NK cells), promotes Tfh cell differentiation, and enhances antibody production (in B cells), thereby strengthening both cellular and humoral arms of effector immunity while fostering memory cell generation. Simultaneously, Foxp3 expression is suppressed by IL-21 and the suppressive TME is disrupted with the downregulation of the immunosuppressive response and proliferation of Tregs. Furthermore, IL-21 has also been associated with the innate and adaptive immune reaction which provides a positive feedback mechanism between the two types of immune responses. These mechanisms of regulation, which are characterized by the presence of molecular effects, have put IL-21 in a good position to become an effective agent in anti-tumor immunity. They also provide the argument that the use of immunotherapy using IL-21 is combined with other types of immunotherapies (Figure 1).

Figure 1 Pleiotropic mechanisms of interleukin-21 in antitumor immunity. IL-21: Interleukin-21; JAK: Janus kinase; STAT3: Signal transducer and activator of transcription 3; Treg: Regulatory T cell; IFN: Interferon; NK: Natural killer; NKG2D: Natural killer receptor group 2 member D; Klf2: Kruppel-like factor 2; CXCR6: C-X-C motif chemokine receptor 6; PI3K: Phosphatidylinositol 3-kinase; Akt: Protein kinase B; IgA: Immunoglobulin A; XBP1: X-box binding protein 1; IgG: Immunoglobulin G.

COMBINATION OF IL-21 WITH TUMOR IMMUNOTHERAPIES

Since its discovery in 2000, IL-21 has been recognized as an important regulator of adaptive and innate immunity, with clear roles in antiviral and antitumor responses. Because of these immunomodulatory effects, IL-21 has been studied as a therapeutic candidate for cancer. Preclinical work has examined IL-21 either as a direct treatment component or in combination with ICIs, cellular therapies, oncolytic viruses, and other treatment strategies, with early evidence of antitumor activity[9] (Figure 2, Table 1).

Figure 2 Strategic combination therapies of interleukin-21 for remodeling the tumor microenvironment. IL-21: Interleukin-21; ICIs: Immune checkpoint inhibitors; PD-1: Programmed cell death protein 1; CTLA-4: Cytotoxic T-lymphocyte antigen-4; TIM-3: T cell immunoglobulin and mucin domain-containing protein 3; IL-21R: Interleukin-21 receptor; TME: Tumor microenvironment; Tregs: Regulatory T cells; MDSCs: Myeloid-derived suppressor cells; CAR: Chimeric antigen receptor; ICR: Inverted cytokine receptor; STAT3: Signal transducer and activator of transcription 3; Th17: T helper 17 cell; OV: Oncolytic virus; NK: Natural killer; DC: Dendritic cell; GM-CSF: Granulocyte-macrophage colony-stimulating factor.

Synergistic effects of IL-21 in combination with ICIs

ICIs, particularly antibodies targeting CTLA-4, PD-1, or PD-L1, are now standard treatments for several advanced solid tumors[44]. Many patients, however, either do not respond or develop acquired resistance, which limits the overall clinical benefit of these agents[2]. Combination strategies that improve ICI efficacy have therefore become a major focus in cancer immunotherapy. IL-21, with its unique ability to reverse T cell exhaustion and remodel the TME, has been recognized as an ideal candidate for combination with ICIs to enhance antitumor activity[9,45].

Preclinical studies support a synergistic interaction between IL-21 and ICIs[46]. In mouse models of melanoma and lung cancer, recombinant IL-21 combined with anti-PD-1 or anti-CTLA-4 antibodies inhibited tumor growth more effectively than either treatment alone, mainly by increasing the proliferation and function of tumor-infiltrating CD8+ T cells[47,48]. Zhang et al[49] further showed that anti-CTLA-4 therapy, but not anti-PD-1 monotherapy, promotes cytotoxic reprogramming of exhausted PD-1+CD8+ T cells by augmenting IL-21 production from helper T cells. This reprogramming, which shifts the cells from a canonical exhausted state to an effector-functional phenotype, depends on differential activation of STAT1 and STAT3 signaling and involves modulation of the transcription factor TOX. In B16 melanoma models, genetic deletion of the IL-21R or antibody-mediated blockade of IL-21 signaling abolished the therapeutic effect of anti-CTLA-4 treatment, indicating that IL-21 is required for effective CTLA-4 blockade[49]. These findings clarify why IL-21-based combinations may improve therapeutic responses and suggest that activation of the IL-21 pathway could serve as a biomarker for ICI-based regimens[44]. Engineering strategies for targeted IL-21 delivery to tumor-reactive T cells have been shown to enhance anti-PD-1 efficacy and promote memory T cell formation, thereby supporting long-term antitumor immunity[50]. Additionally, Deng et al[51] reported that IL-21 remodels the TME by expanding a PD-1 intermediate T cell immunoglobulin and mucin domain-containing protein 3 (Tim-3) negative CD8+ T cell subset, thereby sensitizing tumors to anti-PD-1 therapy. In major histocompatibility complex class I-deficient tumor models, IL-21 combined with PD-1 and Tim-3 blockade enhances NK cell-mediated antitumor activity, providing a potential strategy for tumors that are less dependent on classical T cell responses[52]. These findings provide a robust mechanistic foundation for combining IL-21 with ICIs.

Combination of IL-21 with CAR-T cell therapy

CAR T cell therapy has produced major clinical benefits in hematological malignancies, but its activity in solid tumors remains limited[53,54]. Solid tumors contain a complex TME marked by immunosuppressive cells, physical barriers, and functional defects in CAR-T cells, including exhaustion, all of which reduce therapeutic efficacy[55]. To overcome these challenges, IL-21 has emerged as a key immunomodulatory cytokine for enhancing CAR-T cell therapy in solid tumors. By promoting CAR-T cell proliferation and cytotoxicity while simultaneously modulating the TME to improve infiltration and persistence, IL-21 presents a promising candidate for combination strategies.

Preclinical studies have shown that IL-21 can improve CAR-T cell function. One common strategy is to engineer CAR-T cells to secrete IL-21 Locally, thereby maintaining cytokine exposure within the tumor site. For instance, Liu et al[56] generated B7H3-targeting CAR-iNKT cells that secrete IL-21 and showed enhanced in vivo antitumor activity against renal cell carcinoma without evident toxicity. Similarly, IL-21-armed CAR-T strategies have exhibited potent tumor inhibition in melanoma models. Mechanistically, IL-21 co-expression improves CAR-T cell expansion and survival, promotes a central memory T cell phenotype, and reduces the accumulation of Treg and myeloid-derived suppressor cells in the TME, thereby improving CAR-T cell infiltration and durability[55]. In an esophageal squamous cell carcinoma model, epidermal growth factor receptor-targeted nanobody-based CAR-T cells co-expressing IL-21 showed strong antitumor activity in vitro and in vivo, together with increased IFN-γ and TNF-α secretion and higher frequencies of CD8+ T cells and central memory T cells[57].

In addition to direct cytokine co-expression, engineered antigen-presenting cells have been employed to expand functional lymphocyte populations. Portillo et al[58] showed that peripheral blood-derived Vδ1 T cells expanded with K562 feeder cells expressing membrane-bound IL-21 acquired strong innate antitumor activity. These cells controlled tumor growth in ovarian cancer xenograft models, and their activity was further improved after CAR engineering. Moreover, to convert immunosuppressive signals within the TME into activating cues, innovative cytokine receptor designs have been explored. Wang et al[59] developed a 4/21 inverted cytokine receptor by fusing the extracellular domain of the IL-4 receptor to the intracellular domain of the IL-21R and expressing it in CAR-T cells. In the presence of IL-4, this receptor converted an inhibitory cytokine signal into IL-21-like activation, drove CAR-T cells toward a Th17-like phenotype through STAT3 signaling, and produced durable tumor clearance in hepatocellular carcinoma models[59].

Encouraged by promising preclinical data, clinical translation of IL-21 and CAR-T cell combinations is underway. A phase I trial in patients with relapsed/refractory B-cell non-Hodgkin lymphoma evaluated CD19 CAR-T cells secreting a PD-1-targeted IL-21 fusion protein. Preliminary results from nine patients revealed an overall response rate of 77.8% and a complete response rate of 66.7%. All instances of cytokine release syndrome (CRS) were grade 1 or lower, suggesting a favorable safety profile alongside promising efficacy[60].

In general, IL-21 can be used to enhance the treatment of solid tumors with CAR-T cells because it has a positive impact on the growth and survival of T-cells, the ability to maintain memory phenotype, and the cytotoxicity. While early clinical results are promising, further studies are needed to optimize combination parameters, such as dosing, timing, and cellular design, to strike an ideal balance between efficacy and safety.

Combination of IL-21 with oncolytic virotherapy

Oncolytic virotherapy is a form of the virus-based therapy that can be used to kill the tumor cells by using engineered viruses, which are able to selectively lyse and infect the tumor cells as well as promote the antitumor immunity[61]. However, its efficacy as a monotherapy is often limited by the immunosuppressive TME[9]. IL-21, as a potent immunomodulatory cytokine, demonstrates synergistic potential in this context by enhancing tumor antigen release and presentation following viral oncolysis, promoting DC maturation, and directly increase the cytotoxicity of NK cells and CD8+ T cells to the tumor cells infected by the virus[62].

Preclinical studies combining IL-21 with oncolytic virotherapy have advanced significantly. A major strategy is to arm oncolytic viruses with the IL-21 gene, allowing sustained local cytokine expression in the TME[63]. Chen et al[61] engineered an oncolytic vaccinia virus expressing IL-21 (VV-IL-21). In melanoma models, VV-IL-21 monotherapy significantly inhibited tumor growth and prolonged survival compared with the parental virus. Mechanistic analyses showed that IL-21 expression increased the proliferation and IFN-γ production of tumor-infiltrating CD8+ T cells and enhanced NK cell cytotoxicity, thereby counteracting the immunosuppressive TME[61]. Xuan et al[64] later reported an oncolytic vaccinia virus co-expressing granulocyte-macrophage colony-stimulating factor (GM-CSF) and IL-21 (VVL-GL21) in pancreatic cancer models. Intratumoral or systemic administration of VVL-GL21 induced marked tumor regression, increased infiltration of DCs, macrophages, and T cells, promoted DC maturation, and shifted tumor-associated macrophages from an M2-like toward an M1-like phenotype. Notably, VVL-GL21 also promoted immune memory, prevented tumor recurrence, and showed synergy with anti-PD-1 blockade[64]. Together, these studies show that IL-21-armed oncolytic viruses can substantially increase antitumor immune activation.

Building on strong preclinical evidence, the clinical translation of oncolytic viruses engineered to express IL-21 is actively progressing. For example, a recombinant human IL-21-armed oncolytic vaccinia virus, hV01, developed by a Chinese company, has entered clinical testing in patients with advanced solid tumors. Available information indicates that the phase I trial is ongoing, with preliminary safety data suggesting that the treatment is well tolerated. However, detailed efficacy outcomes and comprehensive immunological profiles await formal publication.

In summary, combining IL-21 with oncolytic virotherapy offers a novel therapeutic strategy for solid tumors, leveraging multiple mechanisms such as enhancing antitumor immunity and remodeling the TME. The synergistic effects observed in preclinical studies have received preliminary validation in early-phase clinical trials. Nevertheless, larger-scale studies are required to optimize viral design, dosing schedules, and combination strategies, thereby achieving an ideal efficacy-safety balance and facilitating the clinical translation of this combination approach.

Combination of IL-21 with other therapeutic modalities

In addition to the immunotherapies previously discussed, IL-21 can be combined with many other immunotherapeutic and conventional therapies which may enhance efficacy by coupling tumor cell killing or antigen release with immune activation.

Combination of IL-21 with chemotherapy: The combination of IL-21 with chemotherapy has shown clear synergistic potential. While chemotherapy may suppress immune cell function and impede immunotherapy, emerging evidence demonstrates that rationally designed combinations can not only be feasible but also yield superior or additive antitumor effects. Preclinical studies have shown that IL-21 can be effectively combined with chemotherapeutic agents such as pegylated liposomal doxorubicin, oxaliplatin, and 5-fluorouracil. A key insight is that the timing of IL-21 administration significantly affects outcomes: Delayed IL-21 delivery following chemotherapy leads to superior antitumor activity, likely due to the transient lymphopenia induced by certain chemotherapies, which necessitates a period of immune recovery before immunostimulation[46]. Chemotherapy has also been associated with an increase in tumor antigen release due to the presence of immunogenic cell death, and IL-21 has also been implicated in enhancing effector activity of CD8+ T cells and NK cells, thus resulting in a feedback loop of antigen release and immune stimulation[65]. HepG2 co-treatment with recombinant human IL-21 and 5-fluorouracil (5-FU) at 48 hours resulted in 89.34% cytotoxicity, which is higher than the 27.71% of 5-FU alone, and less toxicity to normal fibroblasts[66]. Furthermore, enhanced antitumor effects have also been reported for IL-21 combined with 5-FU in colon cancer models, further supporting the feasibility of this strategy[30]. This implies that the treatment of IL-21 after the chemotherapy will enhance the immune surveillance against the tumors and lead to the destruction of the remaining cancerous cells.

Combination with targeted therapies: Clinical trials of IL-21 and the specific drugs have also shown positive results. The use of IL-21 in combination with vascular endothelial growth factor receptor tyrosine kinase inhibitors has been tested in a number of trials in renal cell carcinoma[45]. A phase I/II trial in pretreated metastatic renal cell carcinoma patients showed that IL-21 combined with sorafenib achieved an objective response rate of 21% and a disease control rate of 82%[67]. Notably, two patients sustained durable responses for over 41 months and 30 months, respectively, even after discontinuing treatment, suggesting that IL-21 may promote long-term immunological memory. Pharmacokinetic analyses showed that the immunostimulatory activity of IL-21 was preserved in the combination setting. In another phase I study including patients with metastatic melanoma (n = 24) and renal cell carcinoma (n = 19), recombinant IL-21 monotherapy had a MTD of 30 μg/kg, acceptable safety, and evidence of antitumor activity, including 1 complete response and 11 cases of stable disease in melanoma, and 4 partial responses and 13 cases of stable disease in renal cell carcinoma[68]. These findings support further evaluation of IL-21 with targeted therapies, particularly in advanced solid tumors such as renal cell carcinoma.

IL-21 as an immune adjuvant for tumor vaccines: IL-21 can also enhance antitumor immunity as an adjuvant for tumor vaccines. Tumor vaccines provide tumor antigens to induce antigen-specific immune responses, while IL-21 promotes the expansion and cytotoxic function of antigen-specific CD8+ T cells and supports immune memory. Preclinical studies have shown that IL-21 enhances the antitumor activity of a GM-CSF-expressing bladder cancer vaccine. Mechanistically, IL-21 promotes Tfh cell differentiation and regulates B cell antibody production, thereby coordinating cellular and humoral immunity[69]. Several preclinical studies are now examining IL-21 with peptide vaccines, DC vaccines, and other vaccine platforms in solid tumor models.

In summary, while IL-21 combinations with ICIs and CAR-T therapies both aim to enhance T-cell-mediated antitumor immunity, their synergistic mechanisms, target cell populations, and clinical considerations differ fundamentally. The role of IL-21 in ICI combinations is mainly to affect the endogenous T cells inside the TME as well as to restore the effector activity of the exhausted ones. In CAR-T complexes, the effect of IL-21 on the engineered cells that were given by the patient was primarily through enhanced expansion and maintenance of these cells and increased functionality. Oncolytic virus combination involves viral lysis of the tumour antigens, and the IL-21 induces an antigenic presentation and recruitment of the effector cells which results in a vaccine-like action.

This has practical implications in the translation of these mechanistic variations to clinical applications. ICI combinations are likely to be most appropriate to the patients with high tumor-infiltrating lymphocytes and the checkpoint expression that is applicable, and it should be closely observed in terms of cumulative immune-related adverse events. IL-21 timing and local concentration control are needed in CAR-T combination and the possible deterioration of cytokine release syndrome. The oncolytic virus combinations will have to consider the possibility of intratumoral administration as well as the effect of neutralizing antibodies. In precision medicine systems in the future, the type of tumors, the immunity microenvironment and individual factors of the patient will dictate the choice of the partners of IL-21 and their doses.

CURRENT STATUS AND CHALLENGES IN THE CLINICAL TRANSLATION OF IL-21

IL-21, a key member of the γc cytokine family, regulates both innate and adaptive immune responses to exert antitumor effects. Its considerable potential in combination therapies, particularly in enhancing ICIs, has established IL-21 as a central candidate in cancer immunotherapy[70]. However, the clinical application of native IL-21 is hindered by its inherent biological properties.

Short half-life

The half-life of IL-21 is quite short, with the time of half-life being in minutes and hours. Consequently, there is a high probability of requiring frequent administration or continuous infusion at higher dosages to achieve therapeutic concentrations, which adds to the burden on treatment as well as risks of cumulative toxicity[71]. More critically, the transient nature of IL-21 limits its spatial and temporal reach, undermining its ability to persistently modulate the TME. This limitation highlights the need for long-acting formulations developed through protein engineering to achieve sustained immune activation, which is crucial for overcoming local immunosuppression.

Systemic toxicity

As a potent immune activator, systemic IL-21 administration is generally associated with mild to moderate and transient adverse events. In phase I trials of intravenous recombinant human IL-21, dose-limiting toxicities were mainly reversible grade 3 laboratory abnormalities, including elevated transaminases, neutropenia, thrombocytopenia, and fatigue[11,12]. In a phase I/II study of IL-21 plus sorafenib in 52 patients with metastatic renal cell carcinoma (NCT00389285), the most common clinical adverse events were fatigue, diarrhea, fever, chills, hand-foot syndrome (HFS), and rash. Fever, chills, fatigue, nausea, and vomiting were usually transient and occurred during the weeks of IL-21 administration. The most frequent grade ≥ 3 adverse events were skin rash (29%), HFS (24%), and fatigue (9%), with rash typically presenting as a generalized maculopapular erythematous eruption within the first two weeks of treatment and progressing rapidly. Common grade 3-4 laboratory abnormalities included lymphopenia (69%), hypophosphatemia (58%), and elevated lipase levels (26%)[67] (Table 1).

IL-21, belonging to the IL family alongside IL-2, shares certain toxicity profiles, particularly those associated with systemic immune activation and inflammatory responses. IL-2 is well known to cause capillary leak syndrome (CLS), a severe form of systemic vascular leakage. No cases of CLS have been reported in IL-21 clinical trials to date, although this may partly due to the more limited clinical development of IL-21 compared with IL-2. Given the strong immunostimulatory activity of IL-21, CLS or related vascular toxicities should still be monitored carefully in future studies. Additionally, excessive immune activation could trigger autoimmune phenomena or exacerbate pre-existing inflammatory conditions. Therefore, achieving a balance between robust antitumor efficacy and systemic toxicity remains a central challenge in developing IL-21-based therapies.

Limited monotherapy efficacy

Despite the fact that IL-21 is effective in antitumor activity against tumors, it has not been so effective as a single agent to be used in clinical studies. Phase I/II trials in patients with advanced cancers reported only modest tumor regression or durable remission[9]. It does not mean that this gap is not due to weak immunostimulatory action. In fact, there are various obstacles such as the presence of immunosuppressive TME, short half-life of IL-21, and the lack of ability to target tumor cells following its systemic injection. Consequently, IL-21 alone often fails to overcome these immunosuppressive barriers. Other factors, such as tumor heterogeneity and inter-patient variability, further influence therapeutic outcomes. To enhance the efficacy of IL-21 on immune response and its combination, an increase in the intensity and selectivity of IL-21-induced immune responses would be needed.

Administration route and biodistribution

The mode of administration and the subsequent biodistribution also poses some pharmacokinetic and safety challenges. The predominant method intravenous infusion leads to rapid systemic distribution, causing off-target immune activation and toxicity in normal tissues while failing to achieve sustained, effective concentrations within tumors[72]. Optimizing delivery strategies, such as local or intratumoral administration, to concentrate IL-21 at the tumor site while minimizing systemic exposure is critical. Additionally, the potential for IL-21 to disrupt immune homeostasis requires precise control over its spatial and temporal activity. Addressing these challenges necessitates concerted efforts to optimize IL-21’s clinical application through molecular modifications, long-acting formulations, localized delivery, and combination therapies, as discussed in the following section.

OPTIMIZATION STRATEGIES AND FUTURE DIRECTIONS FOR IL-21

To overcome the clinical limitations of native IL-21, particularly its transient activity and systemic toxicity, recent research has centered on developing multi-dimensional optimization strategies. Main strategies involve protein engineering to enhance molecular characteristics, delivery systems to concentrate IL-21 at tumor sites, and combination regimens pairing IL-21 with complimentary therapies. By improving receptor affinity and stability, enabling targeted enrichment at tumor sites, and rationally integrating with other therapeutic modalities, these complementary approaches aim to amplify anti-tumor immune stimulation while minimizing adverse effects. This section reviews progress in these strategies and discusses future directions for clinical translation.

Protein engineering approaches

Protein engineering plays a pivotal role in enhancing the bioactivity and stability of IL-21. Targeted structural modifications can improve receptor binding affinity, enhance downstream signaling, and optimize pharmacokinetic properties. Recent studies have produced several IL-21 variants designed to overcome the limitations of the native cytokine.

Generation of high-affinity mutants: High-affinity mutants are used to enhance the IL-21 activity. The binding of IL-21 and IL-21R alpha chain can be enhanced through site-directed mutagenesis or directed evolution in order to promote downstream signaling. For example, a structure-guided variant known as IL-21V, incorporating four critical interfacial mutations (Q19 L, Y23N, V69T, K73Q), demonstrated significantly improved receptor affinity and enhanced STAT3 signaling activation. In murine tumor models, IL-21V exhibited broad-spectrum antitumor activity, suppressing the growth of melanoma and colorectal cancer even at reduced doses[73]. This concept is based on the cis-delivery principle of cytokine engineering where altered cytokines are targeted at specific subsets of immune cells with the aim of enhancing their efficacy and minimizing toxicity[74]. This research not only validates the feasibility of extending IL-21’s therapeutic window through rational design, but also provides a basis for dose reduction and toxicity minimization in clinical applications.

Half-life extension strategies: Half-life extension is another critical strategy for addressing the rapid systemic clearance of native IL-21. Fc fusion and PEGylation are the main approaches used to prolong the half-life of IL-21. The neonatal Fc receptor mediated recycling of IL-21-Fc fusion proteins allows the immunostimulatory action of the protein to be retained at a longer period of circulation. IL-21-Fc fusion protein can generate more effective immune cell response as well as antitumor effect than natural IL-21[75]. PEGylation will also increase the weight of molecules, hydrodynamic radius and decrease filtration through the kidneys and increase the half-life. This method has been used effectively with cytokines like IL-2 and IL-15 and could enhance pharmacokinetics without significant loss of bioactivity[76-78]. However, PEGylation can alter receptor binding kinetics, necessitating a careful balance between prolonged exposure and preserved biological potency.

Stability optimization and novel fusion designs: The stability and manufacturability of IL-21 can also be enhanced by protein engineering. Native IL-21 tends to aggregate and degrade in vitro, which may decrease activity and impair storage stability. Soluble production and formulation properties could be improved by stabilizing mutations and optimized expression systems such as yeast surface display and insect cell systems. New fusion designs can also add functional specificity. For example, fusion of IL-21 to tumor-targeting ligands can enrich the cytokine within the TME and increase local immune activation. In a recent study, an affinity-attenuated IL-21 variant was fused to an antibody targeting selected immune cell subsets, creating an immunocytokine that delivered biased signaling to intended cell populations and aimed to reshape antitumor immunity within tumors[79]. Furthermore, integrating IL-21 into CAR-T cell therapy offers unique advantages. Compared with IL-2, it carries a lower risk of systemic toxicity, making it a more suitable partner for CAR-T-based combination therapies[13].

In summary, protein engineering provides several avenues to optimize IL-21 for clinical use. By improving receptor affinity, extending half-life, and enhancing stability, engineered IL-21 variants promise to deliver more potent and localized immune activation at lower doses, with reduced systemic toxicity. These innovations are essential for translating IL-21 from preclinical research to clinical practice, positioning it as a safer and more effective immunotherapeutic agent.

Local delivery strategies

Local delivery is an important way to reduce systemic toxicity while preserving the immune activity of IL-21 within tumors. By confining IL-21 exposure to the TME, high local concentrations of cytokines can be achieved that activate tumor-infiltrating lymphocytes, with less systemic activation of the immune system. These methods include intratumoral injections, tumor-targeted delivery, nanocarrier-based delivery and engineered cell-based delivery.

Intratumoral injection provides a direct method of local administration. Delivering IL-21 directly into tumor tissue achieves high local concentrations rapidly, robustly stimulating tumor-infiltrating lymphocytes while limiting systemic effects. In murine models, intratumoral IL-21 increases the proliferation and function of these effector cells and improves antitumor efficacy. By contrast, systemic administration is more likely to induce inflammatory responses, highlighting the safety advantage of local delivery[51]. However, this approach is inherently limited to accessible, discrete lesions, presenting challenges for treating multifocal metastases or deep-seated tumors.

Tumor-targeted delivery systems have been developed to address these limitations by enriching IL-21 at tumor sites through conjugation to tumor-specific ligands. One strategy is to fuse IL-21 to antibodies that recognize tumor vasculature, thereby directing the cytokine to the TME. This design is supported in part by earlier evidence that IL-21 has anti-angiogenic activity and can act directly on tumor endothelial cells[80]. Other approaches use features of the TME to trigger cytokine release, including pH-sensitive or enzyme-degradable carriers that respond to acidic or protease-rich tumor conditions. Simultaneously, the molecular design of targeted fusion proteins is being refined. For example, Möller et al[81] evaluated the design of tumor-targeted IL-15/IL-21 trifunctional fusion proteins, assessing how variations in antibody formats, cytokine combinations, and spatial arrangements affect their functionality. They found that there was an increased activity of scFv-based fusions IL21 but no off-targets activity, which supports the specificity of delivering cytokines into targets and providing a framework for optimizing IL-21 fusion proteins.

There is also a system of nanocarriers to deliver IL-21 in localized areas. The nanoparticles are capable of entrapment of IL-21, which will not be degraded and will accumulate in the tumor cells by passive means (e.g., increased permeability and retention effect) or active targeting. Such systems may have the advantage of local sustained release and extend the activity of pharmacodynamics. For instance, IL-21 loaded lipids or polymeric nanoparticles can be injected intravenously and the cytokine released in the tumors. Preclinical experiments revealed that lipid nanoparticles containing mRNA encoding IL-21 could induce the local immune cell response, inhibit local as well as distant tumour growth, and the minimal exposure to the body makes it safe[72]. This method has been made even more plausible due to developments in the formulations of lipid nanoparticle delivery of mRNA[82]. In addition to conventional nanoparticles, innovative biomimetic platforms have emerged. Wu et al[83] developed neutrophil membrane-coated vesicles that display IL-21 on their surface and co-load oxaliplatin and a photosensitizer for multimodal therapy of esophageal squamous cell carcinoma. These vesicles exploit the tropism of the inflammatory neutrophil membranes and reach the tumor site and then release drugs in a photothermally manner. This design facilitates IL-21’s synergy with chemotherapy, promoting DC maturation and CD8+ T cell infiltration, and exemplifies a novel paradigm for local cytokine delivery.

Gene delivery and engineered cell-based delivery represent another distinct and impactful direction for localized IL-21 therapy. The viral vectors, plasmids, or other delivery systems are used to introduce the material with the encoding of IL-21 into tumor tissues or autologous immune cells, which is then expressed in vivo over time and local immunological responses. It is also possible to modify the immune cell (T cell or NK cell) by using an engineered cell-based delivery system to express IL-21 that can be released as a cytokine at the sites of tumors. These strategies have the potential to provide long term IL-21 expression in the tumour and also have the ability to move around the tumour and access disseminated disease through the migration of immune cells. CD19 CAR-NK cells were produced by He et al[84] genetically engineering to co-express IL-21. Compared to CAR-NK cells co-expressing IL-15, CAR-IL-21 NK cells secreted higher levels of effector cytokines and cytotoxic molecules, exhibited significantly enhanced cytotoxicity against lymphoma cells, and demonstrated improved in vivo persistence, resulting in suppressed tumor growth in xenograft mouse models. There was an increase in cytokine signaling and cytotoxicity in CAR-IL-21 NK cells as determined by transcriptomic analysis that indicates a specific functional state. However, gene and engineered cell-based delivery systems present challenges, including concerns about safety and potential immune responses, which require further investigation and optimization.

In summary, local delivery platforms have pioneered novel approaches for translating IL-21 into clinical use. By concentrating the cytokine within the TME, these strategies enhance anti-tumor immunity while minimizing systemic exposure and associated toxicities. Ongoing advancements in nanotechnology, targeting methodologies, and genetic engineering are expected to further refine these approaches, positioning locally delivered IL-21 as a promising therapeutic candidate.

Combination therapy design

As described above, IL-21 has demonstrated synergistic antitumor activity with ICIs, CAR-T cell therapy, oncolytic viruses, chemotherapy, targeted agents, and cancer vaccines in preclinical and early clinical studies. The clinical value of these combinations will hinge on thoughtful regimen design, since IL-21 needs to be utilized in a manner that enhances immune activation but does not amplify its shortcomings[85].

In combination therapy, the optimization of administration timing and dose adjustments is critical. In combination with ICIs, there is some evidence that administration of IL-21 prior to checkpoint blockade could be helpful since it has the ability to prime the immune cells prior to PD-1 or PD-L1 antibodies releasing inhibitory signals. In CAR-T cell therapy, IL-21 tends to be used after CAR-T cell infusion where it can promote expansion and persistence but not excessive inflammation prior to engraftment. Additionally, dosing regimens must be carefully calibrated according to the MTD of each agent to ensure manageable cumulative toxicity. This principle was highlighted in a phase I trial combining IL-21 with cetuximab, where dose-limiting toxicities were monitored to confirm the safety profile of the combination[86]. The initial monotherapy trials of IL-21 have also determined the maximum tolerable doses of intravenous and subcutaneous administration, which offers a significant foundation on the subsequent combination trial development.

In addition to dose optimization, the safety profile of IL-21-based combination regimens, especially the risk of cumulative or synergistic toxicity, warrants further rigorous investigation. It can be hypothesized that IL-21 has the potential to promote immune-related adverse effects when used in combination with ICIs, since it enhances the effector activity of T cells. There are not yet enough clinical trials to support the use of high-grade immunological complications in the combination of IL-21 and ICI but colitis, pneumonitis, and endocrinopathies must be monitored carefully. Early-phase clinical data derived from CAR-T therapy, such as CD19-directed CAR-T cells engineered to secrete PD-1-targeted IL-21, demonstrate that only low-grade CRS is observed in relevant trials. Nonetheless, to minimize potential risks, future trial designs should incorporate stepwise dose escalation, delayed IL-21 initiation following CAR-T infusion, and protocol-driven management of irAEs and CRS. These safety considerations are critical to preserving the synergy of these combinations while maintaining an acceptable toxicity profile.

The synergistic effect observed in different combination regimens is not only determined by the mechanistic properties of each monotherapy, but also affected by multiple variables including administration order, dose proportion and patient-specific immune microenvironment. These considerations collectively emphasize the necessity of developing efficient, customized designs for combination regimens. With the development of bioinformatics and big data analysis, it is possible to use computational tools to predict the effectiveness of synergistics and toxicity of all kinds of combinations used by researchers to develop more scientifically based therapeutic approaches. In addition, the concept of personalized medicine is guiding the development of combination therapy towards precision; the choice of the best partner and dosage regimen on the basis of individual immunological status, tumor molecular phenotype, and microenvironmental phenotypes may have positive effects on the index of the therapeutic effect of IL-21.

On the whole, the rational design of combination therapies significantly expands the clinical potential of IL-21. Combination of IL-21 with other therapeutic modalities enables effective anti-tumor immunotherapy without elevating the dosage of single agents, while concomitantly reducing adverse effects. As combination strategies are refined and personalized medicine frameworks become more deeply embedded in clinical practice, this approach is poised to become a cornerstone of future cancer immunotherapy, offering clinical benefit to a broader patient population.

CONCLUSION

IL-21 plays a critical role in antitumor immunity through enhancement of effector cytotoxicity, modulation of Treg function, and conditioning of adaptive immune responses. Its pairing with therapies such as CAR-T cells and ICIs has yielded promising synergy in preclinical models, though clinical translation is limited by a short serum half-life, dose-limiting systemic toxicity, and lack of monotherapy efficacy. To overcome these challenges, current strategies focus on molecular optimization through protein engineering, development of targeted delivery systems, and the rational design of combination regimens. Looking ahead, advances in bioengineering technologies are expected to further expand the therapeutic potential of IL-21. Future clinical studies should aim to clarify its mechanism of action across diverse tumor types, identify optimal combination partners, and establish effective dosing schedules. By creating safer and more precise therapeutic approaches, IL-21 has the potential to realize its full promise as an anti-tumor agent and significantly contribute to the evolution of more effective cancer immunotherapies.