CD73 is a key enzyme in the adenosine-mediated immunosuppressive pathway and represents an attractive target for cancer immunotherapy. Here, we aimed to develop novel anti-CD73 antibodies with dual mechanisms of action by simultaneously inhibiting enzymatic activity and promoting receptor internalization to more effectively disrupt the adenosine barrier within the tumor microenvironment.
Two monoclonal antibodies, CR201 and CR202, were generated and characterized for cross-species reactivity, inhibition of both membrane-bound and soluble CD73 enzymatic activity, and domain-specific epitope recognition. Functional assays were performed to evaluate reversal of AMP-mediated T-cell suppression, restoration of IFN-γ secretion, and induction of CD73 internalization. In vivo efficacy was assessed using an A375 melanoma xenograft model.
CR201 and CR202 specifically bound human and cynomolgus CD73 and recognized distinct structural domains, with CR202 targeting the N-terminal domain and CR201 binding the C-terminal domain. Functionally, CR202 demonstrated greater potency against membrane-bound CD73, whereas CR201 achieved complete inhibition of soluble CD73 activity. Both antibodies effectively restored T-cell proliferation and IFN-γ production and promoted internalization of surface CD73. In vivo, treatment with either antibody significantly suppressed tumor growth without observable toxicity.
CR201 and CR202 are domain-specific, dual-mechanism anti-CD73 antibodies that integrate potent enzymatic blockade with receptor internalization, thereby enhancing antitumor immune responses. These findings highlight the therapeutic potential of targeting distinct functional domains of CD73 to overcome adenosine-mediated immunosuppression in cancer.
Copyright © 2026 Zhang, Yuan, Pan, Li, Wang, Zhang, Gu, Hu, Qu, Wang, Gu, Wang, Cao, Mu, Kang, de Marco, Zhou and Huang.

Conventional tumor-infiltrating lymphocyte (TIL) therapy for solid tumors relies on high-dose interleukin-2 (IL-2) during expansion and post-infusion, and promotes T-cell exhaustion and toxicity. Herein, we developed a feeder-free, low-dose IL-2 TIL expansion protocol and evaluated whether hydroxychloroquine (HCQ) or programmed cell death protein 1 (PD-1) blockade might enhance therapeutic efficacy and decrease IL-2 dependence.
TILs from multiple solid tumors were expanded ex vivo with decreased-dose IL-2, IL-7, and IL-15 plus CD3/CD28 co-stimulation, without feeder cells. TIL products were assessed via quality control, T-cell phenotypes, and exhaustion markers. Cytotoxic activity was measured in vitro through interferon-gamma (IFN-γ) release and real-time cell analysis (RTCA). HCQ-induced changes in major histocompatibility complex class I (MHC-I) and programmed death-ligand 1 (PD-L1) expression were assessed in tumor cell lines, and RTCA-based cytotoxicity was evaluated using T-cell receptor-engineered T cells (TCR-T cells). The in vivo efficacy of HCQ and PD-1 blockade separately combined with TIL therapy was examined in a colorectal cancer patient-derived xenograft (PDX) model.
The protocol consistently produced viable TILs of favorable quality across tumor types, with variable CD8+ and memory T-cell profiles. Expanded TILs showed effector-to-target (E:T) ratio-dependent tumor cell killing in RTCA and secreted IFN-γ across multiple tumor types. HCQ significantly upregulated MHC-I expression in vitro (P < 0.05) without affecting PD-L1 expression or impairing TIL proliferation, and enhanced early TCR-T-mediated killing. In the PDX model, TIL plus HCQ, compared with TIL, showed less tumor growth and greater MHC-I expression, although these differences were not significant, given the small sample size. TIL plus low-dose PD-1 blockade significantly reduced tumor volume versus the control group (P = 0.002) and maintained higher body weights than the TIL-only and control groups.
The feasibility of a feeder-free, low-dose IL-2 TIL expansion system was demonstrated. PD-1 blockade significantly enhanced antitumor activity and treatment tolerability, thus supporting its promise as an alternative to high-dose IL-2. HCQ demonstrated potential immunomodulatory effects, although its in vivo benefit was minimal. This strategy warrants further clinical evaluation in solid tumors.
Copyright © 2026 The Authors.

The pro-inflammatory cytokine, interleukin-18 (IL-18), plays an instrumental role in bolstering anti-tumor immunity. However, the therapeutic application of IL-18 has been limited due to its susceptibility to neutralization by IL-18 binding protein (IL-18BP), short in vivo half-life, and unfavorable physicochemical properties.
In order to overcome the poor drug-like properties of IL-18, we installed an artificial disulfide bond, removed the native, unpaired cysteines, and fused the stabilized cytokine to an IgG Fc domain. The stability, potency, pharmacokinetic and pharmacodynamic properties as well as efficacy of disulfide-stabilized IL-18 Fc-fusion (dsIL-18-Fc) were assessed via in vitro and in vivo studies.
The stability and mammalian host cell production yields of dsIL-18-Fc were improved, compared to the wild-type (WT) cytokine, while maintaining its biological potency and interactions with IL-18 receptor α (IL-18Rα) and IL-18BP. Recombinant fusion of the cytokine to an IgG Fc domain provided extended half-life. Notably, despite maintaining sensitivity to IL-18BP, dsIL-18-Fc was effective at activating both T and natural killer (NK) cells, and elicited a strong anti-tumor response, either as a single agent, or in conjunction with anti-programmed cell death-ligand 1 (anti-PD-L1) therapy.
We engineered IL-18 for reinforced stability, extended half-life, and improved manufacturability. The therapeutic benefit of dsIL-18-Fc, coupled with a more favorable manufacturability profile and enhanced drug-like properties, underscores the potential utility of this engineered cytokine in cancer immunotherapy.
© Author(s) (or their employer(s)) 2025. Re-use permitted under CC BY-NC. No commercial re-use. See rights and permissions. Published by BMJ Group.

Glucocorticoid-induced tumor necrosis factor receptor (GITR) is a co-stimulatory receptor and an important target for cancer immunotherapy. We herein present a potent FcγR-independent GITR agonist IBI37G5 that can effectively activate effector T cells and synergize with anti-programmed death 1 (PD1) antibody to eradicate established tumors. IBI37G5 depends on both antibody bivalency and GITR homo-dimerization for efficient receptor cross-linking. Functional analyses reveal bell-shaped dose responses due to the unique 2:2 antibody-receptor stoichiometry required for GITR activation. Antibody self-competition is observed after concentration exceeded that of 100% receptor occupancy (RO), which leads to antibody monovalent binding and loss of activity. Retrospective pharmacokinetics/pharmacodynamics analysis demonstrates that the maximal efficacy is achieved at medium doses with drug exposure near saturating GITR occupancy during the dosing cycle. Finally, we propose an alternative dose-finding strategy that does not rely on the traditional maximal tolerated dose (MTD)-based paradigm but instead on utilizing the RO-function relations as biomarker to guide the clinical translation of GITR and similar co-stimulatory agonists.
Copyright © 2022 The Author(s). Published by Elsevier Inc. All rights reserved.

B-cell maturation antigen (BCMA) is an ideal target for the treatment of multiple myeloma (MM), and a bispecific antibody (bsAb) of BCMA × CD3 has entered clinical trials with great potential. In this study, we characterized GR1803, a novel bsAb for MM treatment. We reported that GR1803 bound to recombinant BCMA, CD3, BCMA-positive (BCMA + ) MM cells, and human T cells; that it induced T cell activation and cytokine release dependent on the presence of BCMA + cells; and that it effectively killed MM cells with EC 50 values less than 10 ng/mL. We found that GR1803 induced MM cells to release various cytokines, including interleukin (IL)-2, IL-4, IL-6, IL-8, IL-10, interferon-g (IFN-γ), GM-CSF, and TNF-α. Finally, we found that GR1803 significantly and dose-dependently inhibited the growth of tumors specifically expressing human BCMA in vivo . Taken together, these results demonstrate that GR1803, a novel BCMA × CD3 bsAb, efficiently and selectively kills MM cells and represents a novel immunotherapy for treating MM.