Revolutionizing Cancer Therapy With Bispecific Antibodies

WHITE PAPER INTRODUCTION W���� P����n� �n� I�n�va�i�� A�v�n�� Bi��e�i���� techsupport@acrobiosystems.com 1 Innovation Way, Newark, DE 19711 +1 800-810-0816 (US / Canada) +86 400-682-2521 (Asia & Pacific) In recent decades, one of the most significant breakthroughs in cancer therapy has been the introduction of monoclonal antibodies, especially immune checkpoint inhibitors (ICIs) that target the PD-1/PD-L1 pathway. These inhibitors have demonstrated remarkable efficacy and versatility in treating a range of cancers.1 However, the clinical application of ICIs faces several challenges, including the relatively low overall response rates, a high incidence of immune-related adverse events (irAEs), and the issues of both primary and acquired resistance.1,2 To address these limitations, bispecific antibodies (bsAbs) have emerged as a promising alternative. to overcome some of the limitations associated with monoclonal antibody (mAb) therapy, including ICIs.3 BsAbs offer enhanced therapeutic potential by precisely linking immune cells with tumor cells, thereby improving targeted engagement and minimizing off-target effects. By simultaneously binding two distinct antigens or epitopes, bsAbs increase therapeutic specificity, reduce drug resistance, and lower the incidence of severe side effects.4 Unlike traditional combination therapies that use multiple monoclonal antibodies and carry and pose increased risk of toxicity, bsAbs provide a more controlled and focused treatment , enhancing safety and efficacy.5 This review discuss the classification of bsAbs targeting Immune checkpoints are broadly classified into two primary types: co-stimulatory and co-inhibitory molecules. Abnormal expression of inhibitory checkpoints on cancer cells plays a critical role in immune evasion and drug resistance. BsAbs targeting these checkpoints can be categorized into three main groups: 1) those that target dual inhibitory checkpoints, 2) those that target both co-stimulatory and inhibitory checkpoints, and 3) those that target immunomodulatory checkpoints along with non-checkpoint molecules.2 CLASSIFYING BISPECIFIC ANTIBODY TARGETING IMMUNOMODULATORY CHECKPOINTS Targeting dual inhibitory checkpoints Bispecific Antibodies Targeting Immunomodulatory Checkpoints for Cancer Therapy The concept of therapeutically inhibiting two immune checkpoints has driven the rational design of bsAbs capable of targeting two inhibitory checkpoints simultaneously, either on the same cell or on different cells. This innovation builds upon the success of immune checkpoint blockade (ICB) therapies, offering enhanced clinical benefits that have been observed in patients receiving combination ICB treatments. Figure 1. BsAb classification2 Fab A Fc VH VL CL CH B Knobs-into-holes Duobody DVD-Ig IgG-(scFv2) C BiTE DART TandAb immunomodulatory checkpoints and discusses their emerging applications in cancer immunotherapy. PD-1 x CTLA-4 bsAbs Cadonilimab W���� P����n� �n� I�n�va�i�� A�v�n�� Bi��e�i���� techsupport@acrobiosystems.com 1 Innovation Way, Newark, DE 19711 +1 800-810-0816 (US / Canada) +86 400-682-2521 (Asia & Pacific) Bispecific Antibodies Targeting Immunomodulatory Checkpoints for Cancer Therapy Page 2 of 9 Combining PD-1 and CTLA-4 targets in a single bsAb has shown stronger anti-tumor activity compared to individual mAbs.2 Studies have shown that combining mAbs drugs targeting CTLA-4 like Ipilimumab with mAbs drugs targeting PD-1 like Pembrolizumab or Nivolumab improves therapeutic outcomes in patients with advanced renal cell carcinoma (RCC), melanoma, and non-small cell lung cancer (NSCLC).2 However, these mAbs drug combinations are often associated with an increased risk of severe irAEs. Notably, PD-1×CTLA-4 bsAbs exhibit enhanced anti-tumor efficacy while presenting more manageable irAEs.6,7 Cadonilimab (AK104) is the world's first approved PD-1 × CTLA-4 bsAb targeting dual inhibitory immune checkpoints. Its approval for the treatment of relapsed or metastatic cervical cancer underscores the feasibility and potential of bsAbs as a novel therapeutic approach.2 In patients with recurrent or metastatic cervical cancer, frontline therapy combining Cadonilimab with platinum-based chemotherapy -with or without Bevacizumab-has demonstrated a progression-free survival (PFS) advantage, successfully meeting the primary endpoint of the Phase III AK104-303 trial (NCT04982237).8 Recently, Cadonilimab received approval in China for treating patients with recurrent or metastatic cervical cancer who had previously failed platinum-containing chemotherapy. Notably, findings from the Phase III COMPASSION-15 trial, Vudalimab In addition, another PD-1 × CTLA-4 bsAb drug, Vudalimab (XmAb20717), exhibits increased affinity for PD-1/CTLA-4 dual-positive cells. Both pre-clinical and clinical findings have shown manageable side effects.10 Furthermore, phase II clinical trials of Vudalimab Alone or in combination with Chemotherapy or targeted therapy for patients with metastatic castration-resistant prostate cancer (mCRPC) have been initiated.11 MEDI5752 MEDI5752, unlike Cadonilimab, is a distinct anti-PD-1 × CTLA-4 bsAb fusing an anti–PD-1 mAb and Tremelimumab's variable binding domains on a DuetMab backbone, and preferentially binds to PD-1+ T cells in the TME. This design is enhanced with triple amino acid mutations in the human IgG1 constant heavy chain to decrease Fc-mediated immune effector functions.12 A phase II dose-escalation and dose-expansion study indicated that MEDI5752 has promising antitumor activity in patients with advanced RCC.2 Figure 2. The bsAbs targeting immunomodulatory checkpoints. The checkpoint-targeted bsAbs are mainly divided into three categories: targeting dual inhibitory checkpoints (①); targeting co-stimulatory and inhibitory checkpoints (②); and targeting immunomodulatory checkpoints and non-checkpoint targets (③④).2 PD-1 CTLA-4 TIM-3 LAG-3 1 2 Dual inhibitory checkpoint bsAbs Co-stimulatory and inhibitory checkpoint bsAbs T cell Tumor cell TAA and immunomodulatory checkpoint bsAbs PD-1 TGF- , VEGF PD-L1 PD-L1 PD-1 CD137 CTLA-4 4 TAA GITR ICOS CD137 OX40 PD-1 CTLA-4 3 Growth factors/cytokines and immunomodulatory checkpoint bsAbs presented at the AACR Annual Meeting 2024, further highlight Cadonilimab's potential. The trial demonstrated that PD-1 × CTLA-4 bsAb, in combination with chemotherapy, significantly improves both progression-free and overall survival for patients with untreated, HER2-negative, locally advanced or metastatic gastric or gastroesophageal junction (GEJ) cancer. This benefit was observed even in patients with PD-L1-low tumors, marking an improvement over chemotherapy alone.9 Tebotelimab The dual immunomodulator Tebotelimab (MGD013) targets LAG-3 and PD-1, blocking PD-1 interactions with PD-L1 and PD-L2 while also preventing LAG-3 from binding to MHC II. Its DART structure enables synergistic T cell activation.12 Tebotelimab exhibits a more robust capacity for T cell activation and cytokine secretion compared to individually blocking PD-1 or LAG-3 signaling.15 A recent clinical study has preliminarily confirmed the safety and efficacy of Tebotelimab as monotherapy or in combination with Margetuximab—an anti-HER2 monoclonal antibody—in patients with HER2-positive recurrent or refractory advanced solid tumors.16 New progress in nanoantibodies W���� P����n� �n� I�n�va�i�� A�v�n�� Bi��e�i���� techsupport@acrobiosystems.com 1 Innovation Way, Newark, DE 19711 +1 800-810-0816 (US / Canada) +86 400-682-2521 (Asia & Pacific) Bispecific Antibodies Targeting Immunomodulatory Checkpoints for Cancer Therapy Page 3 of 9 Figure 3. The structure and properties of Z15-0 nanobody13 Figure 4. Clinical trial design and patient flow16 Zeng et al engineered Z15-0-2, a bispecific nanobody against PD-1 and CTLA-4, demonstrating marked antitumor efficiacy in mice. They optimized its mRNA sequence to enhance expression, positioning Z15-0-2 as a promising candidate for advanced cancer immunotherapy. The strategy, empolying lipid nanoparticles (LNP) for mRNA delivery (LNP-mRNA), could surpass current immune checkpoint inhibitors in efficacy.13 PD-1 x LAG-3 bsAbs Lymphocyte activation gene-3 (LAG-3) monoclonal antibodies (mAbs) represent the fifth class of immune checkpoint inhibitors approved by the FDA for cancer treatment.14 The co-expression of LAG-3 and PD-1 is closely associated with exhausted effector T cells in ovarian cancer. Blocking LAG-3 and PD-1 simultaneously has been shown to enhance the cytotoxicity of CD8+ T cells, suggesting a promising therapeutic strategy.2 Clinical investigations are underway for several bsAbs targeting LAG-3 and PD-1 across various cancer types. MTD/MAD EOC (up to 40 pts) Cervical (up to 16 pts) TNBC (up to 40 pts) NSCLC (up to 32 pts total, 16 CPI-naive + 16 post-CPI) SCCHN (up to 32 pts total, 16 CPI-naive + 16 post-CPI) SCLC (up to 16 pts) Cholangio (up to 16 pts) DLBCL (up to 20 pts, of which 10 post-CAR-T) HER2+ locally advanced or metastatic solid tumors HER2+ breast cancer (up to 30 pts) HER2+ GC/GEJ cancer (4 pts) Other HER2 + cancer (up to 30 pts) Tebotelimab 300 mg IV + margetuximab 15 mg kg –1 IV (both Q3W) 1,200 mg 800 mg 400 mg 120 mg 30 mg 10 mg a 3 mg a 1 mg a Tebotelimab 600 mg IV + margetuximab 15 mg kg –1 IV (both Q3W) Separate HC C escalation: 120 mg →40 0 mg →600 mg (Q2W) 3+3 design Flat dosing Q2W: single-patient cohorts followed by 3+3 design 3+3 dose escalation Dose escalation (tebotelimab 1–1,200 mg IV Q2W) Cohort expansion (tebotelimab 600 mg IV Q2W) Dose escalation Previously treated unresectable, locally advanced or metastatic solid tumors of any histology Tebotelimab monotherapy (Q2W) Combination of tebotelimab and margetuximab (Q3W) Cohort expansion (tebotelimab 600 mg IV Q3W + margetuximab 15 mg kg –1 IV Q3W) MTD/MAD Treatment discontinuation ( n = 53) • PD (n = 39) • Adverse event ( n = 10) • Death (n = 2) • Withdrawal by subject ( n = 0) • Physician decision ( n = 1) • Completed ( n = 0) • Other ( n = 1) Tebotelimab monotherapy enrollment ( n = 277) Dose escalation ( n = 54) Not treated ( n = 1) Not treated ( n = 7) Cohort expansion ( n = 223) Evaluable patients Safety (n = 53) Response (n = 44) Evaluable patients Safety (n = 216) Response (n = 181) Treatment ongoing ( n = 0) Treatment ongoing n = 4) Combination tebotelimab + margetuximab enrollment ( n = 89) Dose escalation ( n = 20) Not treated ( n = 0) Not treated ( n = 5) Cohort expansion ( n = 69) Evaluable patients Safety (n = 20) Response (n = 17) Evaluable patients Safety (n = 64) Response (n = 55) Treatment ongoing ( n = 1) Treatment ongoing ( n = 8) Treatment discontinuation ( n = 212) • PD (n = 153) • Adverse event (n = 28) • Death (n = 11) • Withd rawal by subject ( n = 8) • Physician decision ( n = 5) • Completed ( n = 4) • Other ( n = 3) Treatment discontinuation ( n = 19) • PD (n = 14) • Adverse event (n = 3) • Death (n = 0) • Withd rawal by subject ( n = 1) • Physician decision ( n = 1) • Completed ( n = 0) • Other ( n = 0) Treatment discontinuation ( n = 57) • PD (n = 46) • Adverse event (n = 5) • Death (n = 1) • Withd rawal by subject ( n = 3) • Physician decision ( n = 1) • Completed ( n = 0) • Other ( n = 1) FS118 FS118, targeting LAG-3 and PD-L1, exhibits potential in activating exhausted immune cells and overcoming resistance to PD-L1 blockade. By simultaneously binding to LAG-3 and PD-L1, FS118 disrupts key interactions such as PD-1/PD-L1, CD80/PD-L1, and LAG-3/MHC-II, effectively reversing T cell suppression and promoting cytokine production in CD4+ and CD8+ T cells.17 In colorectal cancer (CRC) syngeneic mouse models, FS118 enhances the anti-tumor immune response and reduces LAG-3 expression on T cells, indicating its potential as an effective immunotherapy.2 ABL501 ABL501, a bsAb targeting LAG-3 and PD-L1, promotes dendritic cell maturation by inhibiting PD-L1 signaling, enhancing CD8+ T cell cytotoxicity.18 In a humanized mouse model, ABL501 shows a dose-d ependent anti-tumor effect by rejuvenating immune cells.19 Additionally, bispecific antibodies targeting LAG-3 × CTLA-4, such as XmAb22841, are also under clinical investigation. RG7769 RG7769, a heterodimeric TIM-3 × PD-1 bsAb developed by Roche, has a high affinity for PD-1 and low affinity for TIM-3. It selectively targets PD-1+ T cells and PD-1+ TIM-3+ T cells, enhancing IFN-γ secretion and boosting the anti-tumor activity of tumor-infiltrating T cells.22 A Phase II study (NCT04785820) is comparing RG7769 with Nivolumab in advanced esophageal squamous cell carcinoma. Another early-phase study on a TIM-3 × PD-L1 bsAb was terminated due to unexpected drug immunogenicity issues.23 W���� P����n� �n� I�n�va�i�� A�v�n�� Bi��e�i���� techsupport@acrobiosystems.com 1 Innovation Way, Newark, DE 19711 +1 800-810-0816 (US / Canada) +86 400-682-2521 (Asia & Pacific) Bispecific Antibodies Targeting Immunomodulatory Checkpoints for Cancer Therapy Page 4 of 9 TIM-3 × PD-1 TIM-3 (T-cell immunoglobulin and mucin domain 3) is a key co-inhibitory receptor expressed on various immune cells Figure 5. The ligands for LAG-3, TIM-3, TIGIT and their interactions14 and frequently co-expressed with PD-1. This co-expression, notably in tumor-infiltrating lymphocytes (TILs), results in reduced T cell activity and cytokine production, aiding in immune dysfunction and tumor immune escape.14 Several ligands for TIM-3 have been identified, including Galectin-9 (Gal-9), high-mobility group protein B1 (HMGB1), phosphatidylserine (PtdSer), and carcinoembryonic antigen cell adhesion molecule 1 (CEACAM-1).14 Galectin-9 is the primary ligand for TIM-3, promoting cell death in TIM-3-expressing T cells.20 TIM-3 upregulation on regulatory T cells contributes to T cell exhaustion, characterized by functional impairment and altered gene expression.21 IC Ligands Expression Mechanism of action LAG-3 MHC-II B cells, MON-Mø, DCs, some activated T cells Tim-3 TIGIT Negatively regulates T cell responses CD155 DCs, T cells, B cells, macrophages Increasing the IL-10 secretion CD113 Liver, testes, lungs, placenta, and kidneys Inhibition of T cell and NK cell activity CD112 Hematopoietic and non-hematopoietic tissues Inhibiting the activation of T cells and NK cells Nectin4 Tumor cells Inhibiting NK cell activity Fap2 Tumor cells Inhibiting NK cell toxicity and T cell activity FGL-1 FGL-1 protein is primarily secreted from hepatocytes Inhibiting antitumor immune responses α-synuclein Neurons, heart, muscle, and other tissues Gal-3 Inhibiting antitumor T cell responses Tumor cells, macrophages, epithelial cells, fibroblasts, activated T cells LSECtin Inhibiting antitumor T cell responses Liver, tumor-associated macrophages, and other tumor tissues Gal-9 APC, MDSCs, Naive CD4 T cells, plasma PtdSer Released from apoptotic cells LAG-3 can recognize α-synuclein fibrils and affect its endocytosis and intercellular transmission, contributing to PD Gal-9 mainly induces calcium to flow into the intracellular area of Th1 cells and induces apoptosis PtdSer and TIM-3 binding contributes to the clearance of apoptotic bodies and antigen cross-presentation by Tim-3+ DCs CEACAM-1 DCs, monocytes, macrophages, and activated T cells CEACAM-1/TIM-3 complex formation has a crucial role in regulating autoimmunity and antitumor immunity HMGB1 Proliferating tissues or estrogen stimulated cancer cells Blocking activation and suppresses innate immune responses to nucleic acids W���� P����n� �n� I�n�va�i�� A�v�n�� Bi��e�i���� techsupport@acrobiosystems.com 1 Innovation Way, Newark, DE 19711 +1 800-810-0816 (US / Canada) +86 400-682-2521 (Asia & Pacific) Bispecific Antibodies Targeting Immunomodulatory Checkpoints for Cancer Therapy Page 5 of 9 TIGIT × PD-L1 TIGIT (T cell immunoreceptor with Ig and ITIM domains), an inhibitory IgSF checkpoint, impedes immune cell activation by competing with essential co-stimulatory receptors like CD226 and CD69. TIGIT interacts with ligands such as PVR and PVRL2 on tumor, T, and antigen-presenting cells (APCs). The approval of Tiragolumab, an anti-TIGIT antibody combined with Atezolizumab for NSCLC with high PD-L1 expression, highlights TIGIT's potential as a target that can enhance the effectivenessof other ICIs.24 A bsAb targeting TIGIT and PD-L1 not only elevates IL-2 secretion but also improves survival rates in preclinical models.25 Additionally, PVRIG, another immune checkpoint that independently binds to PVRL2, plays a complementary inhibitory role. A human bsAb targeting both TIGIT and PVRIG has shown strong anti-tumor activity.26,27 Several bsAbs, including HLX301 and PM1022, which target TIGIT and PD-L1, are currently in phase I clinical trials.25-28 GITR(L) × PD-1 or GITR × CTLA-4 As a member of the TNFR superfamily (TNFRSF), glucocorticoid-induced TNF receptor (GITR) acts as a co-stimulatory checkpoint, promoting T cell activation and counteracting regulatory T cell (Treg) immunosuppression.2 Research reveals that combining anti-GITR antibodies with anti-PD-1 mAbs reduces TIGIT expression and regulating tumor-infiltrating T cells.29 This study paves the way for exploring bispecific constructs targeting GITR and other immune checkpoints. To overcome resistance, a multimeric GITR ligand is combined with an anti-PD-1 mAb to create a 4-1BB × PD-L1 4-1BB, also known as CD137 or TNFRSF9, plays a crucial role in activating and expanding T cells. Clinical trials have examed the safety and efficacy of 4-1BB agonistslike Urelumab and Utolimumab.2 However, the use of these agonists alone can lead to severe toxicities, limiting the development of 4-1BB monoclonal antibodies.5 A bispecific construct could potentially mitiage this issue by selectively regulating the stimulatory effects of 4-1BB agonists in specific immune cells, offering a way to reduce toxicity while enhance therapeutic outcomes.2 Simultaneously targeting co-stimulatory and inhibitory checkpoints bsAbs ICIs rely on pre-existing immune responses, making the combination of co-stimulatory agonists and checkpoint inhibitors pivotal for enhancing therapeutic outcomes. BsAbs bridge tumor and immune cells, restoring T cell functions for improved clinical efficacy while minimizing co-stimulation-indued side effects . This strategy, targeting checkpoints like 4-1BB, OX40, and ICOS, marks a transformative advancement in cancer immunotherapy.2 Immune co-stimulation primilary revolves around the B7-CD28 and TNFR families. CD28 and ICOS are key co-stimulatory receptors in the B7-CD28 family, while OX40, CD40, CD27, 4-1BB, GITR, and CD30 are part of the TNFR family.12 ABL503 By utilizing in trans cell bridging and modifying the Fc region, bsAb such as ABL503 (PD-L1 × 4-1BB) can enhance targeting precision while minimizing systemic side effects. Preclinical studies have shown that ABL503 is well-tolerated, with low risk of liver toxicity, and demonstrates superior activity compared to a combination of the corresponding monoclonal antibodies.32 GEN1046 BNT311/GEN1046 (Acasunlimab) is an investigational PD-L1 × 4-1BB bsAb that combines Genmab's proprietary DuoBody® technology and BioNTech’s immunomodulatory antibodies.³³ By conditionally activates 4-1BB on T cells and NK cells, it triggers an anti-tumor response, inhibiting tumor growth by enhancing T cell proliferation and the secretion of cytokines such as IFN-γ, IL-10, and CXCL10.2 In a Phase II trial evaluating Acasunlimab, alone or in combination with Pembrolizumab in patients with previously treated metastatic non-small cell lung cancer (mNSCLC), results indicated that the combination of GEN1046 with Pembrolizumab demonstrated a manageable safety profile and promising efficacy. These findings were presented at the 2024 American Society of Clinical Oncology (ASCO) Annual Meeting.33 Figure 6. The data of Acasunlimab in ASCO 202433 bispecific construct, enhancing T cell infiltration while reducing the presence of Treg cells and exhausted T cells.30 Beyond PD-1, CTLA-4 is another inhibitory checkpoint considered for dual targeting. ATOR114, a GITR × CTLA-4 bsAb, enhances T and NK cell activation, depletes Treg cells and GITR+ tumor cells. This promising dual-targeting approach awaits validation of its efficacy in clinical trials.31 Unconfirmed ORR Arm A: mono 31% Arm B: Acasunlimab + low-dose Pembro 25% 30% Unconfirmed DCR 50% 65% 75% Confirmed ORR 13% 21% 22% 0 18% 33% mDoR 6-mth PFS 2 mth 6 mth Arm C: Acasunlimab + high-dose Pembro NR Note: Arm A, 100 mg Q3W x 2 cycles then 500 mg Q6W Arm B, 100 mg + pembro 200 mg Q3W Arm C, 100 mg + pembro 400 mg Q6W W���� P����n� �n� I�n�va�i�� A�v�n�� Bi��e�i���� techsupport@acrobiosystems.com 1 Innovation Way, Newark, DE 19711 +1 800-810-0816 (US / Canada) +86 400-682-2521 (Asia & Pacific) Bispecific Antibodies Targeting Immunomodulatory Checkpoints for Cancer Therapy Page 6 of 9 Moreover, a number of 4-1BB × PD-L1 bispecific antibodies, such as ATG-101 (NCT04986865), INBRX-105(NCT03809624) and QLF31907 (NCT05150405), are presently undergoing phase I clinical trials to assess their effectiveness in individuals with advanced solid tumors.2 OX40 × PD-L1/CTLA-4 OX40, also known as CD134, is transiently expressed on CD4+ and CD8+ T cells following T cell receptor activation, while it is constitutively expressed on intra-tumoral Treg cells.2 In mouse models of ovarian cancer, combining PD-1 blockade with OX40 stimulation has been shown to enhance immune cell infiltration and elicit a tumor-specific T-cell response, highlighting the potential of combining OX40 with other immune checkpoints.34 KN-052, developed by Jiangsu Alphamab, is an IgG-like bispecific antibody targeting both OX40 and PD-L1. It demonstrates robust binding to both targets, inhibits PD-L1/PD-1 and PD-L1/CD80 interactions, and enhances immune responses in preclinical models. This leads to increased tumor growth inhibition in mouse models of colon cancer, with promising pharmacokinetics observed in cynomolgus monkeys.35 ICOS × PD-L1 ICOS, a member of the IgSF, provides a stimulatory signal linked to TCR activation and is consistently expressed in Treg cells. KY105, a novel ICOS × PD-L1 IgG1 bsAb, activates ICOS in a PD-L1-dependent manner. In murine models of colorectal cancer, KY105 has shown promising anti-tumor efficacy by reducing ICOS+ Treg cells and enhancing IFN-γ secretion.2 Targeting immune checkpoints and TAAs Targeting Immune Checkpoints and GFs/Cytokines The lack of tumor specificity in ICIs monotherapy can lead to nonspecific targeting of tumor cells and unpredictable irAEs. Tumor-associated antigens (TAAs) serve as a "navigation system," directing immune cells within the tumor microenvironment (TME) while sparing normal tissues. Combining TAAs with immunomodulatory checkpoints presents a potential solution to these challenges.2 By linking immune cells with tumor cells that express high levels of TAAs, bsAbs can activate immune cells' anti-tumor responses and induce apoptosis in tumor cells, effectively combating tumors while minimizing irAEs.25 Extensively studied TAAs such as EpCAM, EGFR, and HER2 have been customized for specificity in bispecific molecules.38 EpCAM, present in various normal epithelial tissues, also serves as a biomarker for cancer stem cells. A CD40 × EpCAM bsAb has been engineered to activate dendritic cells (DCs), enhancing the priming of tumor-specific T cells and extending overall survival in murine models.39 Receptor tyrosine kinases belonging to the ErbB family (EGFR, HER2, HER3, HER4) can facilitate tumor initiation and invasion when aberrantly activated.40 A recently developed symmetrical bsAb targeting EGFR and PD-1 has shown promising results. This anti-PD1 × EGFR bispecific antibody not only inhibits EGFR signaling but also elicits antibody-dependent cellular cytotoxicity (ADCC) in tumor cells, leading to tumor regression in xenograft and syngeneic colorectal cancer models.41 Beyond bsAbs that crosslink TAAs and checkpoints, several bsAbs targeting growth factors (GFs) and cytokines are currently undergoing clinical trials. With a unique design, the bsAb targeting OX40 and CTLA-4 depletes Tregs within the tumor microenvironment while activating effector and memory T cells.36 ATOR-1015, a bsAb targeting OX40 and CTLA-4, depletes Tregs in tumors, activates effector T cells, and shows promise in early-phase trials for advanced solid tumors, reducing tumor growth while improving survival across various cancer models.37 The specificity of pro-tumor growth factors and cytokines can reinforce tumor-killing effects while reducing drug resistance.2 These factors modulate the immunosuppressive TME by influencing immune cell differentiation and function. For instance, TGF-β can drive naive T cells to become Tregs, independently of IL-6, thus hampering anti-tumor immunity.2 Numerous GF and cytokine-targeted drugs have been approved for clinical use, either alone or in combination with ICIs. For example, combined anti-VEGF and antiPD-1/PD-L1 therapies have demonstrated enhanced patient responses across various cancers.42 Integrating GFs and cytokines into immune checkpoint -targeted bsAbs not only enhances immunotherapy efficacy but also directs these agents precisely to tumor sites.² Bispecific antibodies targeting checkpoints and GFs/cytokines, such as YM101 (PD-L1 × TGF-β) and AK112 (PD-1 × Figure 6. The structure of KN052 (Source: the website of Jiangsu Alphamab) Targeting Immune Checkpoints and Non-checkpoint Targets BsAb W���� P����n� �n� I�n�va�i�� A�v�n�� Bi��e�i���� techsupport@acrobiosystems.com 1 Innovation Way, Newark, DE 19711 +1 800-810-0816 (US / Canada) +86 400-682-2521 (Asia & Pacific) Bispecific Antibodies Targeting Immunomodulatory Checkpoints for Cancer Therapy Page 7 of 9 The development of bsAbs targeting immunomodulatory checkpoints represents a transformative approach in cancer therapy, offering a promsing alternative to traditional monclonal antibody treatments. These bsAbs provide several key advantages, including he ability to block two distinct inhibitory signals, activate s specific immune cells within the immunosuppressive TME, and deliver localized stimulatory or inhibitory effects in specific tumor tissues. These characteristics have the potential to yield more effective and safer treatment options for cancer patients. One of the central strengths of bsAbs lies in their capacity to deliver both inhibitory and stimulatory signals. By targeting two immune checkpoints simultaneously, bsAbs can enhance T cell activation while overcoming T cell exhaustion, leading to a more potent anti-tumor response. This dual blockade not only VEGF), are in development, offering potential to overcome immune tolerance in the TME.2 In a Phase I study, AK112 exhibited an overall response rate (ORR) of 23.5% in patients with platinum-resistant/refractory epithelial ovarian cancer, with favorable safety profiles.43 The combination of AK112 with chemotherapy yielded encouraging clinical benefits, including improved progression-free survival (PFS) and ORR, with manageable toxicity in NSCLC patients.44 TGF-β initially suppresses tumor growth but can later contribute to drug resistance and metastasis. Studies indicate that heightened TGF-β activity can reduce the effectiveness of anti-PD-1/PD-L1 therapies. Combining TGF-β blockade with PD-1/PD-L1 inhibition has shown synergistic effects.45 YM101 promotes antigen presentation, enhances T cell infiltration, and boosts tumor cell destruction. It exhibits superior anti-tumor activity compared to individual anti-TGF-β or anti-PD-L1 treatments in both in vitro and in vivo models.46 DISCUSSION restores T cell function but also promotes a more favorable balance between effector T cells and Tregs, which is essential for sustaining anti-tumor immunity over time. Moreover, bsAbs offer precision by exerting localized effects in specific tumor tissues, which can minimize systemic exposure and reduce the risk of adverse effects commonly associated with immune checkpoint inhibitors. This targeted approach enables enhanced therapeutic efficacy while improving safety profiles. For example, innovative bsAbs like ABL503 and GEN1046, designed to modulate the activity of 4-1BB and PD-L1, demonstrate how these agents can selectively enhance T cell responses while mitigating liver toxicity and other systemic side effects. Emerging clinical data underscores the potential of bsAbs to redefine oncology treatment paradigms. Ongoing clinical trials suggests bsAbs, when used in combination with established therapies such as chemotherapy and targeted therapies, may offer synergistic benefits, particularly for patients with advanced cancers who have exhausted standard treatment options. The combination of bsAbs with other modalities could significantly enhance overall treatment outcomes. However, challenges remain in the clinical development of bsAbs. The complex design , the need for precise dosing regimens, and potential immunogenicity require careful evaluation in clinical trials. Additionally, identifying the optimal patient populations who are most likely benefit from bsAbs will be critical to maximizing their therapeutic potential. In summary, bsAbs targeting immunomodulatory checkpoints represent a highly promising frontier in cancer immunotherapy. Their ability to block multiple inhibitory signals, deliver targeted immune activation, and minimize systemic side effects positions them as a potentially safer and more effective alternative to conventional therapies. Continued research and clinical trials will be critical in fully elucidating the capabilities of bsAbs and to optimizing their integration into the current cancer treatment landscape . PRODUCT RECOMMENDATIONS Comprehensive Related Products for Immuno-Oncology Therapy Immune Checkpoint Immune checkpoint proteins Cytokine targets Agonist target proteins PK assay kits Stable / Reporter cell lines Inhibitor screening kits W���� P����n� �n� I�n�va�i�� A�v�n�� Bi��e�i���� techsupport@acrobiosystems.com 1 Innovation Way, Newark, DE 19711 +1 800-810-0816 (US / Canada) +86 400-682-2521 (Asia & Pacific) Bispecific Antibodies Targeting Immunomodulatory Checkpoints for Cancer Therapy Page 8 of 9 References 1 Meng L, Wu H, Wu J,et al.Mechanisms of immune checkpoint inhibitors: insights into the regulation of circular RNAS involved in cancer hallmarks.Cell Death Dis.2024;15:3. 2 Zhang T, Lin Y, Gao Q. Bispecific antibodies targeting immunomodulatory checkpoints for cancer therapy. Cancer Biol Med. 2023 Mar24;20 (3):181–95. 3 Hosseini SS, Khalili S, Baradaran B, et al. Bispecific monoclonal antibodies for targeted immunotherapy of solid tumors: Recent advances and clinical trials. 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W���� P����n� �n� I�n�va�i�� A�v�n�� Bi��e�i���� techsupport@acrobiosystems.com 1 Innovation Way, Newark, DE 19711 +1 800-810-0816 (US / Canada) +86 400-682-2521 (Asia & Pacific) Bispecific Antibodies Targeting Immunomodulatory Checkpoints for Cancer Therapy Page 9 of 9 About ACROBiosystems ACROBiosystems Group, founded in 2010 and listed in 2021, is a biotechnology company aimed at becoming a cornerstone of the global biopharmaceutical industry by providing life science tools and business model innovation. ACROBiosystems spans across the globe and maintains over twelve offices, research & development centers, and production bases across the United States, Europe, and Asia. Numerous partnerships with well-known enterprises such as Pfizer, Novartis, and other academic institutes have been established. 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