Dual blockade immunotherapy targeting PD-1/PD-L1 and CTLA-4 in lung cancer

The genetic and cellular alterations characteristic of cancer offer pathways for the immune system to generate T-cell responses aimed at identifying and eradicating cancer cells. To enable effective immune-mediated destruction of cancer cells, the Cancer-Immunity Cycle [24] is initiated, striking a delicate balance between non-self recognition and prevention of autoimmunity. Within this cycle, dendritic cells (DCs) capture and process neoantigens that are produced and released by tumors. This process involves signals such as pro-inflammatory cytokines, which are crucial for directing the immune response and avoiding peripheral tolerance to tumor antigens. Following this, the DCs present these antigens to T-cells, thereby triggering and activating effector T-cell responses that are specific to neoantigens. The activated effector T-cells then migrate to the tumor tissue, specifically recognizing and binding to tumor cells through interactions between their T-cell receptors (TCRs) and antigen peptide major histocompatibility complexes (pMHCs), ultimately resulting in the immunological destruction of tumor cells. The death of these tumor cells releases additional neoantigens, thus perpetuating the cycle and further enhancing the response's breadth and depth [24].

However, in cancer patients, the functioning of the Cancer-Immunity Cycle is often suboptimal, due to tumor cells evading immune surveillance by suppressing the activation and effector functions of both the innate and adaptive immune systems [25]. For instance, neoantigens may not be recognized or may be misidentified as self-antigens [24], and the tumor microenvironment (TME) may inhibit the generated effector cells [26]. The advent of cancer immunotherapy has introduced a new paradigm for patients, facilitating the reactivation, amplification, and propagation of the Cancer-Immunity Cycle while minimizing unrestrained autoimmune inflammatory responses. Among these strategies, ICIs have become pivotal in restoring this cycle.

However, a significant challenge in cancer immunotherapy is the resistance to ICI treatment, with immune tolerance playing a critical role. The mechanisms of ICI resistance are categorized into tumor-intrinsic factors and tumor-extrinsic factors [30–32]. Intrinsic resistance arises when tumor cells possess either constitutive or acquired mutations in genes involved in immune regulation, affecting processes such as immune recognition, cell signaling, gene expression, and DNA damage response. Extrinsic resistance are related to TME. TME is composed of tumor-extrinsic factors, including various immune cells, stromal cells, vascular systems, extracellular matrices, and cytokines that influence treatment responses. Through impaired homeostasis between immune suppressive and pro-inflammatory cytokines and mediators, the dynamic interplay in the TME often lead to increased infiltration of myeloid-derived suppressor cells (MDSCs), tumor-associated macrophages, M2 macrophages, and regulatory T cells (Tregs) [31, 33–35]. Immunosuppressive cells and inhibitory cytokines in the TME impair anti-tumor immune responses [36, 37]. For instance, Tregs promote self-tolerance by inhibiting effector T cell (Teff) function through suppressive cytokines and direct contact, thereby limiting inflammation [38, 39]. The infiltration of Tregs into many tumor types has been observed, indicating the presence of an immunosuppressive environment, potentially leading to immunotherapy resistance due to the inability to increase Teffs or reduce Tregs [40–42]. Moreover, continuously evolving immune responses, ranging from antigen presentation to the cytotoxicity of cancer cells and the formation of immune memory, can facilitate evasion of anti-tumor immunity. The inability of T cells to proliferate and diversify may also contribute to ICI resistance [30].

The advent of dual blockade immunotherapy effectively overcomes immune tolerance, enabling bypass of resistance. The clinical benefits of dual blockade immunotherapy are potentially attributed to complementary mechanisms. Given the roles of PD-1/PD-L1 and CTLA-4 in the Cancer-Immunity Cycle, CTLA-4 inhibitors initiates T cell activation, whereas PD-1/PD-L1 inhibitors are involved in the reactivation of effector responses in later stages. Blocking both targets can prevent cancer cells from evading exhausted T-cell immunity while simultaneously delivering signals to antigen presenting cells (APCs) that activate T-cells in the early stages, therefore strengthening the immune response [43, 44]. Furthermore, CTLA-4 inhibitors have been demonstrated to deplete Tregs in the TME and enhance cytotoxic T lymphocyte (CTL)-mediated anti-tumor immunity through enhanced antigen recoginition [45]. Dual blockade immunotherapy also aids in reshaping immune memory, thus facilitating a long-term immune response [46–48]. Additionally, PD-1/PD-L1 inhibitors and CTLA-4 inhibitors exhibit no cross-resistance. Therefore, dual blockade immunotherapy offers substantial advantages over ICI monotherapy.

Numerous preclinical models have consistently demonstrated that ICIs targeting CTLA-4 and PD-1/PD-L1 can restore the recognition and eradication of tumor cells by T cells. In 1996, Leach et al. [15] demonstrated through murine tumor models that CTLA-4 inhibitors disrupted the signals that typically downregulated endogenous T cell responses, enabling the activation of usually unresponsive T cells. Wong et al. [49], in 2007, examined the effects of PD-1 inhibitors on the ex vivo expansion and functional capacity of human melanoma antigen-specific CD8⁺ CTLs. They found that PD-1 blockade enhanced the frequency and absolute numbers of CTLs, and increased the proportion of antigen-specific CTLs capable of recognizing melanoma targets through degranulation. Furthermore, a kinetic analysis of cytokine secretion revealed that PD-1 blockade also augmented the accumulation of type 1 and type 2 cytokines in the cultures.

Compared to monotherapy with a single ICI, the combination of PD-1/PD-L1 and CTLA-4 inhibitors is more than merely additive. Wei et al. [43, 50] reported that their synergistic effect significantly surpassed the sum of their effects when used individually. The combined use of dual immune checkpoint inhibitors significantly decreased the proportion of exhausted phenotype cytotoxic CD8⁺ T-cells, while simultaneously increasing the presence of active effector T-cells, including active CD8⁺ and CD4⁺ effector cells (Fig. 2C). This shift in the immune cell population from exhausted T cells to active effector cells has the potential to substantially enhance the overall immune response. Sun et al. [51] demonstrated that the combination therapy of PD-1 and CTLA-4 inhibitors significantly reduced the risk of tumor relapse and metastasis in mouse models, thereby markedly prolonging survival (p < 0.05). Similarly, Yeo et al. [52] observed that the use of a single T cell immunoglobulin and ITIM domain (TIGIT) antibody in mouse models did not produce a significant antitumor response, but dual ICI therapy combined with PD-L1 inhibitor Tecentriq elicited a discernible antitumor effect.

Similar findings were reported in the preclinical murine model experiments conducted by Curran et al. [53] In mice pre-implanted with B16-BL6 melanoma, vaccination with B16-Flt-3 ligand (Fvax) combined with a CTLA-4 inhibitor resulted in tumor rejection in 10% of the mice, while adding a PD-1 inhibitor to Fvax increased tumor rejection to 25% of the mice. The combination of CTLA-4 and PD-1 ICIs resulted in tumor rejection in 50% of the mice, more than doubling the efficacy of using either ICI alone. Incorporating a PD-L1 inhibitor into this regimen resulted in tumor rejection in 65% of the mice. Furthermore, the study revealed that the combined blockade of PD-1 and CTLA-4 enhanced the infiltration of Teffs, thereby producing a highly favorable ratio of Teffs to Tregs within the tumor. Additionally, a higher proportion of CTLA-4 and PD-1 positive T cells, which are normally inactivated, remained active.

In view of these findings and other preclinical studies [54], one can infer that the dual ICI therapy combining CTLA-4 with PD-1/PD-L1 is potentially superior to monotherapy with ICIs, providing a solid foundation for subsequent clinical trials.

In addition to the dual ICI combination therapy, with the continuous advancements in antibody engineering technology, BsAbs that bind two different epitopes on the same or different antigens offer an alternative approach in cancer therapy for enhancing immunity and reducing toxicity. The concept of merging multiple specific targets into a single antibody dates back to the 1960s [55]. Catumaxomab (anti-CD3 × anti-EpCAM) [21], the first BsAb approved for cancer treatment, heralded the beginning of a new era in cancer therapy with the advent of BsAbs. This innovative approach represents a significant step forward in the ongoing quest to optimize cancer treatments by enhancing both specificity and efficacy.

Natural antibodies consist of paired heavy (H) and light (L) chains, typically exhibiting a Y-shaped configuration. Papain can split them into two functional fragments, specifically the Fab and Fc segments. The Fab fragment comprises the antigen-binding site, while the Fc segment facilitates interactions with cells and effector molecules [56]. Through this mechanism, natural antibodies can participate in a variety of immune processes, including antibody-dependent cell-mediated cytotoxicity (ADCC), antibody-dependent cellular phagocytosis (ADCP), and complement-dependent cytotoxicity (CDC) [57]. However, the two antigen-binding sites of natural antibodies are identical, imparting monospecificity and bivalency characteristics. This structural uniformity restricts the scope of their immune engagement, thus necessitating the exploration of more versatile antibody designs in therapeutic applications.

Pang et al. [65] performed preclinical experiments on the bsAb Cadonilimab (AK104), targeting PD-1 and CTLA-4. This study showed that Cadonilimab, due to its tetravalent design, exhibits high binding affinity, especially in environments rich in PD-1 and CTLA-4, and can simultaneously bind to distinct cells expressing PD-1 and CTLA-4, demonstrating differential activity compared to ICI. Moreover, Cadonilimab, compared to the combination of anti-PD-1 and anti-CTLA-4, can activate T cells by increasing the secretion of interleukin-2 (IL-2) and interferon-γ (IFN-γ). These characteristics potentially contribute to reducing the toxicity of Cadonilimab and enhancing its antitumor activity. KN046, a bsAb targeting PD-L1 and CTLA-4, was shown in preclinical mouse model trials by Jiang et al. [66] to cause significant increases in the percentage of CD3⁺ CD4⁺ and CD3⁺ CD8⁺ T cells in the tumor and spleen, indicating a robust in vivo immune response. Additionally, residual tumor imaging showed that mice treated with KN046 experienced slower melanoma growth, further comfirming its antitumor efficacy. The outcomes of these preclinical experiments offer crucial evidence for the further advancement of clinical trials involving bsAbs.

Nivolumab and ipilimumab, representative drugs of PD-1 and CTLA-4 inhibitors respectively, served as the focus of the CheckMate 012 (NCT01454102↗) trial [79], a multi-cohort Phase I study. This pivotal study was the first to assess the efficacy and safety of dual ICI combination therapy in patients with advanced NSCLC. Participants in the study were divided into three groups, each receiving one of three distinct dosing regimens: nivolumab 1 mg/kg Q2W (every 2 weeks) + ipilimumab 1 mg/kg Q6W, nivolumab 3 mg/kg Q2W + ipilimumab 1 mg/kg Q12W, nivolumab 3 mg/kg Q2W + ipilimumab 1 mg/kg Q6W. The latter two regimens received particular focus in the trial. In these two cohorts, all treated patients (n = 77) exhibited an objective response rate (ORR) of 43%, significantly higher than the 23% ORR observed in patients treated with nivolumab monotherapy (n = 52). Notably, in subgroups defined by PD-L1 expression levels, the ORR of the dual immunotherapy consistently exceeded that of nivolumab monotherapy. Furthermore, the adverse reactions associated with the dual immunotherapy were tolerable, with grade 3–4 TRAEs at 37% and 33% respectively. The outcomes of this trial suggested that the combination therapy of nivolumab 3 mg/kg Q2W with ipilimumab 1 mg/kg Q12W or Q6W offered more promising clinical outcomes and improved safety profiles compared to previous treatments. This was particularly true for patients with tumor PD-L1 expression of 1% or higher, establishing a solid foundation for subsequent Phase II and III trials.

The Phase II trial CheckMate 568 [80] was designed to evaluate the efficacy and safety of first-line treatment with nivolumab in combination with low-dose ipilimumab in advanced NSCLC patients, and to investigate the correlation between treatment efficacy with PD-L1 expression and tumor mutational burden (TMB). The study enrolled 288 previously untreated advanced NSCLC patients, who received a regimen of nivolumab 3 mg/kg Q2W combined with ipilimumab 1 mg/kg Q6W. Results showed an ORR of 30% across all patients, with an ORR of 41% in patients with PD-L1 expression ≥ 1% and 15% in those with PD-L1 expression < 1%. Regardless of PD-L1 expression levels, patients with a TMB ≥ 10mut/Mb (n = 48: PD-L1 ≥ 1%, 48%; PD-L1 < 1%, 47%) showed benefits in ORR and progressive free survival (PFS) compared to those with TMB < 10 mut/Mb (n = 50: PD-L1 ≥ 1%, 18%; PD-L1 < 1%, 5%). However, no significant correlation was observed between PD-L1 expression and TMB. Grade 3–4 TRAEs were reported in 29% of the patients, with gastrointestinal toxicity being the most common (5%). This trial reaffirmed the efficacy and safety of the combination immunotherapy of nivolumab and ipilimumab as a first-line treatment in advanced NSCLC. It also highlighted TMB as a potential predictive biomarker for the efficacy of dual ICI combination therapy and evaluated its threshold value.

CheckMate 227 [81], an open-label Phase III clinical trial, classified patients with stage IV or recurrent NSCLC based on PD-L1 expression (≥ 1% or < 1%) and assigned them to either part 1a or 1b of the study. Following stratification, patients were randomized into three groups. In part 1a, the treatment options included nivolumab 3 mg/kg Q2W combined with ipilimumab 1 mg/kg Q6W, nivolumab monotherapy, or chemotherapy alone. In part 1b, the treatment regimens included nivolumab 3 mg/kg Q2W combined with ipilimumab 1 mg/kg Q6W, nivolumab plus chemotherapy, or chemotherapy alone. The two-year follow-up results [81] showed that in the part 1a group (patients with PD-L1 expression ≥ 1%), the median overall survival (OS) for the dual ICIs group and the chemotherapy group was 17.1 months and 14.9 months (p = 0.007) respectively, with two-year survival rates at 49% and 11%. In the part 1b group (patients with PD-L1 expression < 1%), the median OS for the dual ICIs group was nearly 5 months longer than that of the chemotherapy group (17.2 months vs. 12.2 months, HR = 0.62). Subgroup analysis [82] demonstrated that patients with TMB ≥ 10 mut/Mb receiving dual ICI combination therapy experienced superior ORR and PFS compared to the chemotherapy group (ORR: 45.3% vs. 26.9%, PFS: 7.2 months vs. 5.5 months). This trial further comfirmed that while the efficacy of dual ICI combination therapy was not significantly related to PD-L1 expression, high TMB expression might identify a population that benefits more from the dual ICI combination therapy. Thus, this study has transformed the therapeutic landscape for patients with PD-L1 expression < 1%, marking a new chapter in the dual immunotherapy treatment of advanced NSCLC.

Beyond NSCLC, numerous clinical trials are currently assessing the efficacy and application of dual immunotherapy with nivolumab and ipilimumab in SCLC. CheckMate 032 [83], a multicenter, multi-arm, open-label Phase 1/2 trial, aimed to assess the safety and efficacy of nivolumab monotherapy and nivolumab in combination with ipilimumab dual immunotherapy in SCLC patients who had progressed after one or more previous treatments, including platinum-based chemotherapy. This trial enrolled 216 SCLC patients, treated with either nivolumab 3 mg/kg Q2W (n = 98) as monotherapy or nivolumab combined with ipilimumab in different regimens (1 mg/kg plus 1 mg/kg (n = 3), 1 mg/kg plus 3 mg/kg (n = 61), or 3 mg/kg plus 1 mg/kg (n = 54)) Q3W as dual immunotherapy. The trial results showed that the ORR and median PFS of dual immunotherapy (1 mg/kg plus 1 mg/kg: ORR 33%; 1 mg/kg plus 3 mg/kg: ORR 23%, median PFS 2.6 months; 3 mg/kg plus 1 mg/kg: ORR 19%, median PFS 1.4 months) were generally better than those of nivolumab monotherapy (ORR 10%, median PFS 1.4 months). However, compared to monotherapy, dual immunotherapy also resulted in a higher incidence of grade 3/4 TRAEs. This trial demonstrated the effectiveness and manageable safety of dual immunotherapy in SCLC patients who have failed platinum-based chemotherapy, offering new hope for these patients with limited treatment options.

The double-blind Phase III trial CheckMate 451 [84] enrolled 834 patients with extensive stage SCLC (ES-SCLC) who had not progressed during first-line platinum-based chemotherapy to evaluate the efficacy and safety of dual immunotherapy with nivolumab and ipilimumab. Patients were randomized into three groups, each receiving either nivolumab 1 mg/kg combined with ipilimumab 3 mg/kg Q3W for four cycles, followed by maintenance nivolumab 240 mg Q2W (n = 279), nivolumab monotherapy at 240 mg Q2W (n = 280), or placebo (n = 275). The trial findings showed that, compared to the placebo, neither the dual immunotherapy nor the nivolumab monotherapy significantly improved OS. PFS in the dual immunotherapy group showed a slight improvement over the placebo (1.7 months vs. 1.4 months, HR: 0.72). This study indicated that for SCLC patients who relapse after at least one platinum-based chemotherapy, dual immunotherapy with nivolumab and ipilimumab may be beneficial. However, as a maintenance therapy, it did not achieve the anticipated efficacy.

Concerning the combination of PD-L1 inhibitors and CTLA-4 inhibitors, durvalumab plus tremelimumab represents one of the most notable therapeutic regimens. Durvalumab functions as a PD-L1 monoclonal antibody that blocks the interaction between PD-L1 and PD-1 with high affinity and selectivity [85], while Tremelimumab is a selective human IgG2 monoclonal antibody targeting CTLA-4 [86]. In the Phase 1b study (NCT02000947↗) [87], the efficacy of durvalumab plus tremelimumab dual immunotherapy in advanced NSCLC patients was assessed for the first time. This study administered escalating dose regimens of durvalumab plus tremelimumab dual immunotherapy in 102 previously untreated advanced NSCLC patients (durvalumab: 3 mg/kg, 10 mg/kg, 15 mg/kg, or 20 mg/kg Q4W, or 10 mg/kg Q2W; and tremelimumab: 1 mg/kg, 3 mg/kg, 10 mg/kg Q4W).The trial determined that most TRAEs were manageable without discontinuing treatment at the 1 mg/kg dose of tremelimumab. As the dose of tremelimumab was gradually increased, a higher frequency of TRAEs was noted, but without an increase in clinical efficacy. Of the 102 patients, 37 (36%) encountered severe TRAEs. The study revealed that durvalumab at 20 mg/kg Q4W in combination with tremelimumab at 1 mg/kg Q4W exhibited manageable tolerability and anti-tumor activity, regardless of PD-L1 expression, laying the groundwork for dosing in Phase III studies.

The Phase III MYSTIC study [88] conducted the initial clinical trial to evaluate the efficacy of durvalumab combined with tremelimumab dual immunotherapy against standard chemotherapy and durvalumab monotherapy in previously untreated advanced NSCLC patients. The study enrolled 1118 previously untreated patients, all of whom had a performance status (PS) of 0–1 and were negative for EGFR/ALK, and who were then randomly assigned to one of three groups: durvalumab 20 mg/kg Q4W as monotherapy, durvalumab 20 mg/kg Q4W in combination with tremelimumab 1 mg/kg Q4W, or platinum-based doublet chemotherapy. The study showed that in patients with PD-L1 expression ≥ 25%, neither of the two immunotherapy achieved the primary endpoint of a significant survival benefit compared to chemotherapy. Furthermore, regardless of PD-L1 expression, the dual ICI combination therapy failed to demonstrate a significant survival benefit in terms of PFS (HR: 1.05, p = 0.705) and OS (HR: 0.85, p = 0.202) compared to monotherapy and chemotherapy. However, in the subgroup analysis, patients with a bTMB ≥ 20 mut/Mb saw a notable improvement in OS with the dual ICI combination therapy compared to monotherapy and chemotherapy (21.9 months vs. 12.6 months vs. 10 months), identifying TMB as a potential predictive biomarker for the efficacy of durvalumab plus tremelimumab dual immunotherapy.

ARCTIC [89], a Phase III randomized open-label study, assessed the efficacy of durvalumab combined with tremelimumab dual immunotherapy in advanced NSCLC patients who had undergone at least two systemic treatments, including at least one platinum doublet chemotherapy. The trial consisted of two independent substudies based on PD-L1 expression (≥ or < 25%). Substudy B randomly assigned 469 patients with PD-L1 expression < 25% to receive either durvalumab plus tremelimumab dual immunotherapy, monotherapy with either ICI, or standard treatment. The results demonstrated that, compared to chemotherapy, the dual immunotherapy showed some numerical improvement in median OS (11.5 months vs. 8.7 months, HR 0.80, p = 0.109) but no difference in median PFS (both 3.5 months, HR 0.77, p = 0.056).

Despite evidence showing that dual ICI combination therapy significantly enhances therapeutic efficacy compared to ICI monotherapy or chemotherapy alone in various tumor types [67, 69, 74, 81], offering improvements in secondary resistance, prolonged responses, and significant survival benefits, its safety profile and limitations have been the focus of ongoing concern and attention. While the safety of dual ICI combination therapy remains within a tolerable range, several large-scale clinical trials, including CheckMate 9LA [91] and CheckMate 227 [81], have shown that the incidence of grade 3/4 TRAEs with dual ICI combination therapy is comparable to, or even higher than, that of the control chemotherapy group, especially at the grade 3/4 level. The Phase III clinical study CheckMate 9LA [91] demonstrated that the incidence of grade 3/4 TRAEs in the combination therapy group was 47%, higher than in the chemotherapy-alone group (38%). Among these, the dosage of CTLA-4 inhibitors was identified as one of the main factors affecting safety, requiring a delicate balance between dose reduction for safety and dose escalation for efficacy. The most common TRAEs associated with dual ICI combination therapy are dermatological (34% for any grade and 4.2% for ≥ grade 3), endocrine (23.8% and 4.2%), gastrointestinal (18.2% and 2.4%), and hepatic (15.8% and 8.2%) events [91]. Moreover, the population benefiting from dual ICI combination therapy seems limited, with multiple clinical trials [80, 82, 88] suggesting more pronounced survival benefits in populations positive for biomarkers such as TMB and PD-L1, and the range of tumor types benefiting from this approach is relatively narrow. Given the potential for increased TRAE risk with dual ICI combination therapy, the opportunity for combination with other drugs and treatment modalities (such as chemotherapy, targeted therapy, radiation, etc.) remains quite limited, as it may further elevate the risk of TRAEs.

KN046 is a novel bsAb targeting PD-L1 and CTLA-4, capable of inhibiting the interactions between PD-L1 and PD-1, as well as between CTLA-4 and CD80/CD86. Preclinical studies [66] have demonstrated that KN046 can mediate the depletion of regulatory T-cells in TME, thereby enhancing anti-tumor immune responses and reducing immunosuppression. A Phase I study of KN046 [102] explored its efficacy, safety, and tolerability in patients with advanced solid tumors. The study enrolled 100 patients, including 59 patients with nasopharyngeal carcinoma and 36 patients with NSCLC. During the dose-escalation phase of the trial, KN046 was administered at doses of 1, 3, 5 mg/kg Q2W, 5 mg/kg Q3W, and 300 mg Q3W, based on a modified toxicity probability interval method. In the dose-expansion phase, the recommended dose was administered. 14.0% of patients experienced grade 3/4 TRAEs, with the most common TRAE being a rash (33.0%). The trial results indicated an overall ORR of 12.5% and a median response duration of 16.6 months. Notably, patients with high expression of CD8 and PD-L1 exhibited a better prognosis. This trial confirmed the promising efficacy and tolerability of KN046 in advanced solid tumors and established a RP2D of 5 mg/kg Q2W for subsequent studies.

The Phase II study (NCT03838848↗) [103] focused on assessing the efficacy and safety of KN046 in advanced NSCLC patients who had failed or were resistant to platinum-based chemotherapy. The study enrolled 64 patients, divided into Cohort A (n = 30) and Cohort B (n = 34), who received KN046 intravenously at doses of 3 mg/kg and 5 mg/kg, respectively. The results indicated that the ORR for Cohorts A and B were 13.3% and 14.7%, respectively, with a median PFS of 3.68 months for both groups. OS was 19.70 months and 13.04 months for Cohorts A and B, respectively. However, the incidence of grade 3/4 TRAEs was observed at 42.2%. This trial confirmed that both dosing regimens of KN046 exhibited promising efficacy and safety in advanced NSCLC patients who had either failed or were resistant to platinum-based chemotherapy.

CTX-8371, developed by Compass, is a bsAb targeting PD-1 and PD-L1, featuring a 2 + 2 symmetric structure and employing common light chain technology. It functions by converting PD-1 positive cells to PD-1 negative cells through multiple pathways, thereby enhancing anti-tumor efficacy. Its Phase I clinical trial (NCT06150664↗) is currently in progress in patients with solid tumors, utilizing a 3 + 3 dose-escalation design ranging from 0.1 to 10 mg/kg across five dosage levels. AK112 is a humanized IgG1 bsAb targeting PD-1 and VEGF [110]. Its Phase II trial [109] aimed to evaluate the efficacy and safety of AK112 in combination with chemotherapy in advanced NSCLC patients. The trial results demonstrated that AK112 combined with platinum-based doublet therapy showed promising anti-tumor activity and safety, serving as the first-line treatment in advanced NSCLC patients without driver mutations as well as in patients with EGFR functional mutations who have failed previous EGFR-TKI therapy, and in advanced NSCLC patients who had failed prior systemic platinum-based chemotherapy and PD-1/PD-L1 inhibitor therapy. This provided a valuable potential treatment option for these patients, which was approved for marketing in May 2024. Amivantamab is a bsAb targeting EGFR and MET, with clinical studies indicating significant efficacy in NSCLC patients with EGFR Exon 20 insertions [111]. It has been granted accelerated approval by the Food and Drug Administration (FDA) for second-line treatment in NSCLC with Exon 20 insertions, becoming the first targeted therapy globally for this mutation.

For patients with SCLC, Tarlatamab is a bsAb targeting delta-like ligand 3 (DLL3) and CD3. In 2023, a Phase 2 trial (NCT05060016↗) [112] involving previously treated SCLC patients indicated that Tarlatamab, administered once every two weeks at doses of 10 mg or 100 mg, exhibited antitumor activity and durable objective responses. Furthermore, it demonstrated favorable survival outcomes with an ORR of 40% in the 10-mg group and 32% in the 100-mg group. The median PFS was 4.9 months and 3.9 months in the 10-mg and 100-mg groups, respectively. And the estimates of OS at 9 months were 68% in the 10-mg group and 66% in the 100-mg group. Besides, this study revealed no new safety concerns compared to the Phase 1 trial. Based on the favorable outcomes of this clinical study, the FDA expeditiously approved Tarlatamab as an innovative therapy for ES-SCLC on May 16, 2024, suitable for patients whose disease continues to progress after platinum-based chemotherapy. This approval also signifies the introduction of the first bsAb targeting DLL3 in the field of SCLC treatment. Additionally, clinical trials are in progress for FPI-2068 targeting EGFR × c-MET (NCT06147037↗), more bsAbs targeting DLL3 in SCLC [113–115], and others.

For both NSCLC and SCLC, chemotherapy remains one of the primary first-line treatment methods. It is noteworthy that, during the process of tumor treatment, chemotherapeutic agents exert bidirectional effects on the immune system, causing systemic immune suppression while also eradicating specific immune cells to aid in the reconstruction of a new immune system [130]. On the one hand, drugs such as cyclophosphamide, gemcitabine, and platinum compounds enhance the antigenicity of tumor cells through mechanisms such as calreticulin exposure, autophagy induction, mobility group box 1 protein, and ATP release [131–133]. On the other hand, chemotherapy can increase the sensitivity of tumor cells to immune attacks by enhancing their visibility to the immune system. Moreover, chemotherapeutic agents such as anthracyclines and oxaliplatin interact with DNA replication and repair mechanisms, triggering immunogenic cell death (ICD) and antigen-specific responses [134]. These interactions between chemotherapy and the immune system provide theoretical support for the potential enhanced efficacy of combining chemotherapy with immunotherapy.

In the context of dual ICI combination therapy, numerous clinical trials have explored the clinical efficacy of combining dual ICIs with chemotherapy. The Phase II clinical trial, CheckMate 568 [80], was divided into two parts: the first part treated patients with nivolumab and ipilimumab, while the second part investigated the combination of nivolumab and ipilimumab with chemotherapy. The results indicated that, although the median OS showed minor differences (part 1: 20.83 months, part 2: 19.35 months), the median PFS was significantly prolonged with the combination therapy (part 1: 5.19 months, part 2: 10.81 months). The Phase III study, CheckMate 9LA [119], further examined the clinical efficacy of the combination of nivolumab and ipilimumab with chemotherapy. Patients were randomized into two cohorts: one to receive the combination therapy and the other to receive chemotherapy alone. The findings revealed that the combined treatment led to an extended survival benefit compared to chemotherapy alone (median OS: 14.13 months vs 10.74 months; median PFS: 6.83 months vs 4.96 months). Similar outcomes were observed in the Phase II clinical trial for durvalumab and tremelimumab (NCT03057106↗) [135], where the cohort treated with durvalumab and tremelimumab combined with chemotherapy demonstrated extended median OS and PFS compared to the dual ICIs treatment cohort (median OS: 16.6 months vs 14.1 months; median PFS: 7.72 months vs 3.22 months). These studies collectively highlight the potential of combining dual ICIs with chemotherapy, with many clinical trials currently ongoing to further explore this combination strategy (NCT03158129↗, NCT03994393↗, NCT03975114↗, NCT03043872↗).

Research on the combination treatment strategy of bsAbs and chemotherapy is currently in its early exploratory phase. AK112, a bsAb targeting PD-1 and VEGF, has shown promising antitumor activity and safety in its Phase II trial [109]. This efficacy extends to first-line treatment in advanced NSCLC patients without driver mutations, patients with EGFR functional mutations who have failed prior EGFR-TKI therapy, and those who have failed previous systemic platinum-based chemotherapy and PD-1/PD-L1 inhibitor therapy. Numerous clinical trials investigating various bsAbs are actively underway, including Cadonilimab targeting PD-1 and CTLA-4 (NCT04647344↗, NCT05377658↗), QL1706 (NCT05487391↗), and MEDI5752 (NCT03530397↗). Overall, the prospects for developing combination treatment strategies involving immunotherapy and chemotherapy appear promising.

Besides chemotherapy, radiotherapy is another common first-line treatment method for lung cancer, operating on the principle of directly damaging tumor cells through ionizing radiation. There is substantial evidence indicating that radiotherapy can trigger both local and systemic immune responses through various mechanisms, thereby exerting multifaceted impacts on tumor immunity. For instance, radiotherapy can induce ICD in tumor cells and release damage-associated molecular patterns (DAMPs), which in turn promote the maturation of DCs [136]. These mature DCs can then present tumor antigens to CD8⁺ T cells, initiating specific immune responses [137]. Additionally, radiotherapy can exhibit immunosuppressive characteristics [138–140]. These interactions between radiotherapy and the immune system provide a theoretical basis for combining these two treatment modalities.

The ‘abscopal effect’, referring to the regression and rejection of unirradiated and distant tumor lesions triggered by radiation [141], has been observed in tumors including lung adenocarcinoma [142]. The underlying mechanism for this phenomenon likely involves radiotherapy enhancing the antigen presentation of tumor cells, thereby increasing the production of CD8⁺ T cells. These newly generated T cells are transported through the bloodstream to distant sites, thereby affecting tumors outside the irradiated area [143, 144]. Notably, several studies have demonstrated that immunotherapy can enhance the abscopal effect, while radiotherapy can amplify the efficacy of immunotherapy [140, 145–147]. This interaction provides new avenues and opportunities for combined treatment strategies in immunotherapy.

A randomized Phase II study [148] investigating recurrent SCLC evaluated the efficacy of durvalumab and tremelimumab with or without stereotactic body radiotherapy (SBRT). Eighteen patients were randomly assigned to either Group A, receiving durvalumab and tremelimumab, or Group B, undergoing immunosensitizing SBRT followed by durvalumab and tremelimumab treatment. The results indicated that Group B had an extended median OS compared to Group A (Group A: 2.8 months, Group B: 5.7 months). Numerous clinical trials exploring the combination of dual ICIs with radiotherapy are currently underway, including nivolumab and ipilimumab combined with radiotherapy (NCT04013542↗, NCT02696993↗), durvalumab and tremelimumab combined with radiotherapy (NCT02888743↗, NCT03923270↗) and others. However, clinical studies investigating the combination of bsAbs with radiotherapy are relatively rare.

Targeted cancer therapy involves the suppression of cancer growth, progression, and metastasis by interfering with specific molecular targets. This strategy includes inhibiting tumor cell proliferation, intervening in the cell cycle, promoting tumor cell differentiation, suppressing metastasis, inducing apoptosis, and obstructing tumor angiogenesis, representing a revolutionary treatment approach [149]. However, targeted therapy currently faces a significant challenge as a substantial number of patients develop resistance to the treatment. Recent studies indicate that targeted cancer therapy can trigger ICD in tumor cells, thereby enhancing the efficacy of ICIs [149]. This discovery has enabled the combination of targeted cancer therapy with immunotherapy, offering a new approach to overcome drug resistance and enhance clinical efficacy.

The Phase I/II MEDIOLA study [150] assessed the efficacy of durvalumab and olaparib combination therapy in patients with metastatic breast cancer harboring germline BRCA1 or BRCA2 mutations. The results demonstrated that after 12 weeks of combination therapy, 80% of the patients exhibited positive disease control and acceptable safety profiles. Additionally, the Phase III clinical trial, IMspire150 study (NCT02908672↗) [151], assessed the clinical efficacy of vemurafenib and cobimetinib in combination with atezolizumab, an anti-PD-L1 monoclonal antibody, in patients with advanced or metastatic melanoma carrying BRAF V600 mutations. The study outcomes indicated that the combination of targeted therapy with immunotherapy significantly prolonged PFS (15.1 months vs 10.6 months), confirming the feasibility and advantage of the combined treatment approach. Currently, numerous clinical trials exploring the combination of bsAbs and targeted therapies are in progress (NCT05816499↗, NCT04646330↗, NCT05420220↗, NCT04777084↗).

Beyond the combined treatment methods mentioned above, many potential combination therapies merit exploration, offering additional possibilities for enhancing the clinical efficacy of immunotherapy. Adoptive immunotherapy using genetically modified T cells expressing antigen-specific chimeric antigen receptors (CARs) is an emerging cancer treatment method with significant promise [152–154]. CARs are synthetic receptors that redirect T cells to tumor surface antigens [155, 156], possessing the unique ability to recognize various cellular targets (including both protein and non-protein entities) and activate T cells without the need for antigen processing and presentation, thereby circumventing human MHC restrictions [157]. Although CAR-T cells can directly eliminate tumor cells, they remain susceptible to inhibition by immune checkpoints. This has brought into focus the strategy of combining CAR-T cell therapy with ICIs. Several preclinical studies [158, 159] using mouse tumor models have validated the feasibility of combining CAR-T cell therapy with ICIs, indicating that ICIs may be an effective strategy to enhance the clinical benefits of CAR-T cell therapy. Clinical trials for this combination therapy are also currently in progress (NCT04003649↗, NCT03726515↗).

In the context of SCLC, DLL3 serves as an inhibitory ligand in the Notch signaling pathway and is highly expressed on the surface of SCLC tumor cells, correlating with the progression of SCLC [113, 160, 161]. Rova-T, a humanized monoclonal antibody targeting DLL3, has demonstrated significant antitumor activity in recurrent SCLC, with manageable safety profiles [162]. Within the immune system, DLL3 plays a crucial role in regulating T cell development. Its absence can induce the activity of Notch signaling, which, in conjunction with TCR signaling, promotes T cell differentiation [163]. This suggests that combining Rova-T with immunotherapy is potential for enhancing the clinical benefits against SCLC. Regrettably, a Phase 1/2 study of Rova-T combined with nivolumab ± ipilimumab for treating 2L + ES-SCLC patients (NCT03026166↗) indicated that, despite its activity in 2L + ES-SCLC, Rova-T with nivolumab and/or ipilimumab were not appropriate due to dose-limiting toxicities (DLTs) [164].

Additionally, various emerging therapeutic modalities, such as CAR-NK cell therapy [165–167] and macrophage-targeted therapy [168, 169], hold the potential for combination with immunotherapy, offering an expanded array of options and possibilities for cancer treatment.