Antibodies

25 pages, 749 KB

Open AccessReview

by Yehan Xiao, Yaru Zou, Mingming Yang, Wendong Gu, Zizhen Ye, Jiaxin Deng, Kyoko Ohno-Matsui and Koju Kamoi

Antibodies 2026, 15(5), 94; https://doi.org/10.3390/antib15050094 - 8 Oct 2026

Abstract

Antibodies in ocular disease have heterogeneous biological meanings: they may act as direct pathogenic effectors, systemic diagnostic biomarkers, indicators of B-cell clonality, or therapeutic agents. This distinction is particularly important in the eye, where immune privilege and the blood–ocular barrier restrict tissue access, [...] Read more.

Antibodies in ocular disease have heterogeneous biological meanings: they may act as direct pathogenic effectors, systemic diagnostic biomarkers, indicators of B-cell clonality, or therapeutic agents. This distinction is particularly important in the eye, where immune privilege and the blood–ocular barrier restrict tissue access, whereas inflammation may permit entry of immunoglobulins, complement, and immune cells or support local humoral responses. This narrative review integrates antibody and B-cell biology across autoimmune and paraneoplastic retinopathies, aquaporin-4-immunoglobulin G-positive neuromyelitis optica spectrum disorder, myelin oligodendrocyte glycoprotein antibody-associated disease, ocular mucous membrane pemphigoid, ocular myasthenia gravis, thyroid eye disease, systemic autoimmune and fibroinflammatory disorders with ocular involvement, and B-cell malignancies including vitreoretinal and ocular adnexal lymphoma. We distinguish the evidence for direct antibody pathogenicity from disease association and emphasize that a positive result is interpretable only in relation to the ocular phenotype, assay platform, titer, specimen compartment, and orthogonal confirmation. Serum testing, ocular-fluid analysis, electrophysiology, multimodal imaging, cytology, histopathology, immunophenotyping, immunoglobulin clonality, and molecular testing have complementary roles. Therapeutic strategies include B-cell or B-lineage depletion, complement inhibition, cytokine- or receptor-directed monoclonal antibodies, insulin-like growth factor-1 receptor blockade, and emerging FcRn- or plasma-cell-directed approaches; however, regulatory status and ocular-specific evidence vary substantially among diseases. A mechanistically stratified framework may reduce overdiagnosis, improve recognition of malignant masquerade syndromes, and support more precise selection of targeted therapy. Full article

(This article belongs to the Section Humoral Immunity)

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18 pages, 29781 KB

Open AccessArticle

Head-to-Head Comparison of FAP Antibody Clones for Tissue Expression Analysis

by Elena Shmakova, Anna Kalinchuk, Elisaveta Tsarenkova, Pavel Bazhenov, Vlada Vasilchuk, Dmitry Loos and Liubov Tashireva

Antibodies 2026, 15(5), 93; https://doi.org/10.3390/antib15050093 - 8 Oct 2026

Abstract

Background: Fibroblast activation protein (FAP) is a promising target for cancer diagnosis and therapy, but the specificity and sensitivity of anti-FAP antibodies used for immunohistochemistry (IHC) vary considerably. A head-to-head comparison of commercially available clones is needed to guide antibody selection, thereby standardizing [...] Read more.

Background: Fibroblast activation protein (FAP) is a promising target for cancer diagnosis and therapy, but the specificity and sensitivity of anti-FAP antibodies used for immunohistochemistry (IHC) vary considerably. A head-to-head comparison of commercially available clones is needed to guide antibody selection, thereby standardizing FAP IHC protocols and ensuring reliable patient stratification for FAP-directed therapies. Methods: We evaluated three anti-FAP clones—FAP/4851 (mouse monoclonal), JA56-11 (recombinant rabbit), and EPR20021 (rabbit monoclonal)—in formalin-fixed, paraffin-embedded tissues from 137 patients with breast (n = 29), colorectal (n = 27), lung (n = 42), and head and neck squamous cell carcinoma (n = 39). FAP expression was assessed by manual semiquantitative scoring (H-score and percentage of positive cells) and, for a validation subset, by QuPath-based digital image analysis. The study included negative (normal liver/skeletal muscle) and positive (WI-38 cell block) controls. Results: Clones JA56-11 and EPR20021 showed specific stromal staining, whereas FAP/4851 exhibited intense non-specific reactivity and was excluded from quantitative analysis. EPR20021 demonstrated significantly higher H-scores than JA56-11 in all four cancer types (Wilcoxon test, p ≤ 0.0179), resulting in a higher overall detection rate under the tested conditions. Aberrant tumor cell expression was detected almost exclusively by EPR20021 in head and neck (59% of cases) and colorectal (59%) cancers. QuPath validation showed good correlation with manual scoring for EPR20021 in the tumor compartment of lung cancer (Spearman’s ρ = 0.701, p = 0.024) with small bias (Bland–Altman). Conclusions: Under the conditions tested, EPR20021 (Abcam) demonstrated higher staining intensity and detection rates than JA56-11 across several tumor types. Clone FAP/4851 showed nonspecific staining under the conditions used in this study and was therefore not suitable for quantitative analysis in the present experimental setting. Consequently, careful selection of well-validated antibody clones is essential for ensuring reproducible and reliable FAP immunohistochemical analysis. Full article

(This article belongs to the Section Antibody-Based Diagnostics)

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20 pages, 4306 KB

Open AccessArticle

Targeting the Complement C1 Proteases Using Camelid-Derived Single Domain Antibodies: Production, Characterization and Possible Application for Tissue/Fluid Analysis

by Nolwenn Haut, Véronique Rossi, Julie Peliconi, Jean-Baptiste Reiser, Nicole M. Thielens, Lubka T. Roumenina and Christine Gaboriaud

Antibodies 2026, 15(5), 92; https://doi.org/10.3390/antib15050092 - 5 Oct 2026

Abstract

Background. The complement C1r and C1s proteases initiate the classical complement pathway within the C1 complex, but emerging evidence points to additional roles in tissues, including cell-intrinsic functions linked to cancer and connective tissue disorders. Few tools are available to explore these plasma [...] Read more.

Background. The complement C1r and C1s proteases initiate the classical complement pathway within the C1 complex, but emerging evidence points to additional roles in tissues, including cell-intrinsic functions linked to cancer and connective tissue disorders. Few tools are available to explore these plasma and tissue-relevant functions. We report the first development and characterization of anti-C1r and anti-C1s VHH single-domain antibodies (nanobodies). Methods. A lama was immunized with the purified human C1r2/C1s2 tetramer. After phage display selection, two anti-C1r VHHs (C1rCl1, C1rCl6) and one anti-C1s VHH (C1sCl7) were produced in E. coli and characterized by ELISA, Western blot, SPR and size-exclusion chromatography using recombinant protease fragments and proenzyme forms. Functional impact on C1s activation was assessed by cleavage assay, and tissue applicability by immunohistochemistry on human skin sections. Results. All three VHHs bound with high affinity to both active and proenzyme forms. Epitope mapping localized C1rCl1 and C1rCl6 to the CCP2 module of C1r, and C1sCl7 to the CCP1 module of C1s. The two anti-C1r VHHs recognize distinct, non-overlapping epitopes and can bind simultaneously to C1r. C1rCl1 mildly inhibited C1s activation by C1r. The VHH pair C1rCl1/C1rCl6 enabled sandwich ELISA detection of C1r in serum, plasma, and cell culture supernatant. Immunohistochemistry on human skin confirmed clean C1r staining in keratinocytes and fibroblasts, with a dissociated tissue distribution compared to C1s. Conclusions. These first anti-C1r and anti-C1s nanobodies constitute useful tools for detection in biological fluids and tissue staining, with potential applications in diseases where these proteases play a pathological role. Full article

(This article belongs to the Section Antibody Discovery and Engineering)

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18 pages, 3357 KB

Open AccessReview

Complement in HER2-Positive Breast Cancer: A Conditionally Protective Mechanism Under Attack

by Dobromira Petkova, Ivaila Kuleva, Zhasmin Stoyanova, Boryana Ilcheva, Albena Fakirova, Zhasmina Mihaylova and Maria Radanova

Antibodies 2026, 15(5), 91; https://doi.org/10.3390/antib15050091 - 5 Oct 2026

Abstract

The therapeutic efficacy of anti-HER2 monoclonal antibodies in breast cancer has traditionally been attributed to direct inhibition of signaling and antibody-dependent cellular cytotoxicity (ADCC). Recent evidence, however, has established complement activation through the classical pathway as a possible adjunct mechanism underlying the synergistic [...] Read more.

The therapeutic efficacy of anti-HER2 monoclonal antibodies in breast cancer has traditionally been attributed to direct inhibition of signaling and antibody-dependent cellular cytotoxicity (ADCC). Recent evidence, however, has established complement activation through the classical pathway as a possible adjunct mechanism underlying the synergistic activity of the trastuzumab and pertuzumab combination. This synergy is mediated by complement-dependent cytotoxicity (CDC) and complement-dependent cellular phagocytosis (CDCP), and is lost upon C1q deletion in preclinical models. Tumor cells evade complement-mediated killing through overexpression of membrane-bound complement regulatory proteins (mCRPs), particularly CD55 and CD59, which correlate with shorter disease-free survival in patients treated with trastuzumab. C1q itself plays a dual, context-dependent role: it acts as a complement-independent tumor suppressor in HER2-positive breast cancer while promoting tumor progression in most other malignancies. Emerging therapeutic strategies, including biparatopic antibodies such as zanidatamab, complement-redirecting immunoconjugates, and mCRP-targeting approaches, aim to overcome complement resistance and enhance antitumor immunity. On this basis, we propose that HER2-positive breast cancer represents a conditionally complement-protective tumor, in which the balance between antibody-directed complement activation and mCRP-mediated evasion determines treatment outcome. This review evaluates the current evidence linking complement to anti-HER2 therapy and highlights future directions for successful treatment. Full article

(This article belongs to the Section Antibody-Based Therapeutics)

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20 pages, 4944 KB

Open AccessReview

Antibody Discovery and Therapeutics in Spondyloarthritis: Recent Advances and Future Directions

by Kwai Wei Chan, Ghee Chong Koo and Che Kang Lim

Antibodies 2026, 15(5), 90; https://doi.org/10.3390/antib15050090 - 5 Oct 2026

Abstract

Spondyloarthritis is a multi-faceted condition housing a group of inter-related chronic inflammatory rheumatic diseases such as ankylosing spondylitis, IBD-associated arthritis, psoriatic arthritis and juvenile idiopathic arthritis. While decades of research and technological advancements have significantly improved our understanding of the disease, there remain [...] Read more.

Spondyloarthritis is a multi-faceted condition housing a group of inter-related chronic inflammatory rheumatic diseases such as ankylosing spondylitis, IBD-associated arthritis, psoriatic arthritis and juvenile idiopathic arthritis. While decades of research and technological advancements have significantly improved our understanding of the disease, there remain many underlying genetic and physiological mechanisms which are unsolved but essential for the development of the right diagnosis and treatment. This mini-review examines our current foundational knowledge of spondyloarthritis, the main concepts related to the disease pathogenesis and the various lines of therapies developed thus far. We hope to provide a perspective on the current and potential antibody discovery platforms for the next decade as computational methods and artificial intelligence become increasingly mature and relevant in clinical research. Full article

(This article belongs to the Section Antibody-Based Therapeutics)

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25 pages, 4389 KB

Open AccessArticle

Development and Validation of Novel VHHs Targeting Astrocyte-Enriched Glutamate Transporters GLT-1 and GLAST-1

by Noelia Gesteira-Pérez, Pieterjan Van Maele, Benedetta Frizzi, Ludo Van Den Bosch, Laura Rué and Maarten Dewilde

Antibodies 2026, 15(5), 89; https://doi.org/10.3390/antib15050089 - 29 Sep 2026

Abstract

Background/Objectives. VHHs (Nanobodies^®) are camelid single-domain antibody fragments with broad applicability in central nervous system (CNS) research, including blood–brain barrier transport, modulation of pathological proteins and molecular detection. However, few VHHs have been developed for cell-type-specific targeting within the brain. Astrocytes [...] Read more.

Background/Objectives. VHHs (Nanobodies^®) are camelid single-domain antibody fragments with broad applicability in central nervous system (CNS) research, including blood–brain barrier transport, modulation of pathological proteins and molecular detection. However, few VHHs have been developed for cell-type-specific targeting within the brain. Astrocytes play essential roles in CNS homeostasis and contribute to many neurological disorders, making them an appealing target for diagnostic and therapeutic approaches. Here, we report the development and validation of the first VHHs directed against the astrocyte-enriched glutamate transporters GLT-1 and GLAST-1. Methods. VHHs were generated from a camelid immune library using phage display. Target specificity and affinity were evaluated in cells overexpressing the mouse and human transporters, as well as in iPSC-derived astrocytes. Target engagement and astrocyte specificity was further assessed ex vivo on brain sections using immunohistochemistry and in dissociated mouse brain cell suspensions by flow cytometry. VHH in vivo binding was evaluated by stereotactic injections within the brain striatum. Results. One anti-GLT-1 VHH and four anti-GLAST-1 VHHs demonstrated high specificity and apparent nanomolar affinity when binding tocells overexpressing the mouse transporters. The anti-GLT-1 VHH was additionally cross-reactive with the human ortholog, as demonstrated in overexpressing cells and iPSC-derived astrocytes. All VHHs showed target engagement on mouse brain sections and astrocyte specificity in dissociated mouse brain cell suspensions. VHH direct brain delivery resulted in localized transporter-specific signal around the injection site, whereas the non-binding isotype control VHH produced no detectable signal. Conclusions. Here we present the first set of VHHs capable of selectively recognizing astrocyte-enriched glutamate transporters GLT-1 and GLAST-1 in vitro, ex vivo, and in vivo. These VHHs constitute promising tools for astrocyte detection in research applications and the cross-reactive anti-GLT-1 lead might hold potential for future diagnostic or therapeutic strategies, including cell-targeted payload delivery. Full article

(This article belongs to the Section Antibody Discovery and Engineering)

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20 pages, 1497 KB

Open AccessReview

Targeted Therapy for Antihistamine-Refractory Chronic Spontaneous Urticaria: A Critical Narrative Review and an Evidence-Graded Approach to Agent Selection

by Luca Guarino, Giuseppe Rizzuto, Vincenzo Coppolelli, Alessia Scipione, Mirella Scattone, Luca Gargano, Pietro Scribani Rossi, Ester Del Duca, Annunziata Dattola, Giovanni Pellacani, Steven Nisticò and Camilla Chello

Antibodies 2026, 15(5), 88; https://doi.org/10.3390/antib15050088 - 17 Sep 2026

Abstract

Background/Objectives: Chronic spontaneous urticaria (CSU) is a mast-cell-driven disease with autoimmune mechanisms and an incompletely clarified aetiology, in which a substantial proportion of patients remain symptomatic despite second-generation H1-antihistamines; until recently, omalizumab was effectively the only targeted option, leaving a considerable non-responder fraction. [...] Read more.

Background/Objectives: Chronic spontaneous urticaria (CSU) is a mast-cell-driven disease with autoimmune mechanisms and an incompletely clarified aetiology, in which a substantial proportion of patients remain symptomatic despite second-generation H1-antihistamines; until recently, omalizumab was effectively the only targeted option, leaving a considerable non-responder fraction. Between February 2024 and September 2025 the landscape changed: dupilumab (anti-IL-4Rα) and the first oral Bruton tyrosine kinase (BTK) inhibitor, remibrutinib, were approved for CSU; omalizumab biosimilars reached the market; the higher-affinity anti-IgE antibody ligelizumab was discontinued; and the investigational anti-KIT antibody barzolvolimab advanced to phase 3—creating new therapeutic opportunities for non-responders while leaving the choice among these second-line options unresolved by current guidelines. We provide a critical synthesis and propose an evidence-graded process for agent selection, together with a proposed repositioning of the available agents, taking the 2022 EAACI/GA²LEN/EuroGuiDerm/APAAACI guideline as the acknowledged reference. Methods: Critical narrative review with a structured but non-systematic search (PubMed, ClinicalTrials.gov) and a pre-specified evidence hierarchy (guideline to expert opinion); each clinical claim is annotated by its level of evidence. Results: Four agents are appraised in turn—omalizumab (anti-IgE, first-line), dupilumab (anti-IL-4Rα), remibrutinib (oral BTK inhibitor, small-molecule comparator) and barzolvolimab (anti-KIT, investigational). Current evidence appears to support omalizumab as first-line, with mechanism-guided second-line choices conditioned on endotype, comorbidity, prior biologic exposure and safety. Conclusions: The complexity of CSU and the incomplete response to currently available therapies support a patient-tailored approach to treatment selection. Full article

(This article belongs to the Section Antibody-Based Therapeutics)

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31 pages, 14913 KB

Open AccessReview

B Cells and Tumor Immunometabolism: Emerging Insights into Immune Regulation and Therapeutic Resistance

by Swati Gupta, Sandip Rath, Surathi Maiti, Tapas Das and Farhat Afrin

Antibodies 2026, 15(5), 87; https://doi.org/10.3390/antib15050087 - 17 Sep 2026

Abstract

Cancer progression and therapeutic resistance are shaped by reciprocal dynamic interactions between malignant cells and the metabolically altered tumor microenvironment (TME). Tumor-associated hypoxia, glucose and amino acid competition, extracellular acidity, lactate accumulation, adenosine, prostaglandin E2 (PGE2) and other metabolic signals can remodel immune [...] Read more.

Cancer progression and therapeutic resistance are shaped by reciprocal dynamic interactions between malignant cells and the metabolically altered tumor microenvironment (TME). Tumor-associated hypoxia, glucose and amino acid competition, extracellular acidity, lactate accumulation, adenosine, prostaglandin E2 (PGE2) and other metabolic signals can remodel immune cell function and determine whether inflammation is tumoricidal or tumor-supportive. Although B lymphocytes have traditionally been viewed as antibody-producing cells, tumor-infiltrating B (TIL-B) cells comprise functionally heterogeneous populations that can act as antigen-presenting cells, cytokine and chemokine producers, antibody-secreting cells, cytotoxic effectors, regulatory B cells (Bregs) and organizers of tertiary lymphoid structures (TLSs). Their functional state is strongly influenced by the metabolic and spatial context in which they reside. This review focuses on the intersection of B cell biology and tumor immunometabolism, emphasizing how hypoxia, lactate, nutrient limitation, adenosine, PGE2, kynurenine and B cell-derived γ-aminobutyric acid (GABA) may shape B cell states and their interactions with myeloid and lymphoid cells. We discuss how metabolically conditioned Bregs and immunoglobulin (Ig)A-skewed humoral responses can contribute to immune suppression, whereas metabolically competent antigen-presenting, IgG-biased and TLS-associated B cell responses may support effective anti-tumor immunity. Importantly, the effects are tumor type- and context-dependent: B cell/TLS signatures are associated with favorable outcomes in several breast, lung and other solid tumors, whereas B cell-centered immune landscapes can be suppressed or neutral in pancreatic cancer and IgA-dominated responses may be unfavorable in selected malignancies. We further examine how these states may influence sensitivity or resistance to immune checkpoint blockade, chemotherapy, radiotherapy and cellular therapies. Finally, we highlight B cell metabolic pathways as potential therapeutic entry points and identify priorities for spatial metabolomics and prospective interventional studies. Full article

(This article belongs to the Special Issue Unravelling Effector Functions of B cells in Infectious Diseases and Cancer)

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25 pages, 3960 KB

Open AccessReview

Antibody-Based Biologics for CNS Disorders

by Eric Ka-Wai Hui and Ruben J. Boado

Antibodies 2026, 15(5), 86; https://doi.org/10.3390/antib15050086 - 16 Sep 2026

Abstract

The blood-brain barrier (BBB) is permeable only to small lipophilic molecules, whereas hydrophilic nutrients, such as glucose and amino acids, cross the BBB via carrier-mediated transport systems. In contrast, circulating proteins do not cross the BBB. However, the BBB expresses receptors able to [...] Read more.

The blood-brain barrier (BBB) is permeable only to small lipophilic molecules, whereas hydrophilic nutrients, such as glucose and amino acids, cross the BBB via carrier-mediated transport systems. In contrast, circulating proteins do not cross the BBB. However, the BBB expresses receptors able to induce receptor-mediated transcytosis (RMT) of selective proteins across the BBB, as in the case of insulin, transferrin, and insulin-like growth factor 1 (IGF1). This led to the hypothesis that the transport of protein-based biologics to the brain may be possible by targeting BBB receptors that induce RMT. Monoclonal antibodies targeting these receptors penetrate the BBB via RMT and are distributed throughout the brain, acting as molecular Trojan horses and/or shuttle systems. A generation of brain-penetrating fusion proteins was produced targeting BBB insulin, transferrin, and IGF1 receptors, respectively, with full antibodies and/or antibody fragments acting as transport domains. These antibody-based fusion proteins were validated in various experimental models, including lysosomal storage disorders (LSDs), stroke, Parkinson’s disease, and Alzheimer’s disease (AD), respectively. Clinical trials with brain-penetrating antibody-based biologicals targeting BBB transferrin and insulin receptors have been completed or are in progress in LSDs and in AD. The aim of this review is to examine the progress made in antibody-based biologicals for CNS disorders from genetic engineering to clinical trials. Full article

(This article belongs to the Section Antibody-Based Therapeutics)

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16 pages, 840 KB

Open AccessArticle

Prevalence and Clinical Associations of Systemic Sclerosis-Related Autoantibodies: A Nationwide Reuma.pt Cohort Study

by Carolina Mazeda, Eduardo Dourado, Raquel Freitas, Patrícia Martins, Liliana Saraiva, Tânia Santiago, Francisca Guimarães, Emanuel Costa, Diogo Esperança Almeida, Sara Dinis, Ana Sofia Pinto, Alexandra Daniel, Inês Genrinho, Maura Couto, Marília Rodrigues, Maria João Salvador, Ana Catarina Duarte, Ana Cordeiro, Maria José Santos, João Eurico Fonseca, Catarina Resende and Inês Cordeiro Show full author list Hide full author list

Antibodies 2026, 15(5), 85; https://doi.org/10.3390/antib15050085 - 15 Sep 2026

Abstract

Background: Autoantibodies are central to the diagnosis and risk stratification of systemic sclerosis (SSc), but their prevalence and clinical associations may vary across populations. This study aimed to evaluate the prevalence and immuno-clinical associations of different autoantibodies in the Rheumatic Diseases Portuguese Register [...] Read more.

Background: Autoantibodies are central to the diagnosis and risk stratification of systemic sclerosis (SSc), but their prevalence and clinical associations may vary across populations. This study aimed to evaluate the prevalence and immuno-clinical associations of different autoantibodies in the Rheumatic Diseases Portuguese Register systemic sclerosis cohort. Methods: This was a multicentre observational registry-based study that used data from the Reuma.pt SSc module. Patients were divided according to autoantibody status, and associations between autoantibody expression and clinical data were assessed using appropriate statistical tests, with Bonferroni correction applied for multiple comparisons. Multivariable binary logistic regression was performed for clinically relevant outcomes, with adjustment for sex, age at diagnosis and disease duration. Results: A total of 1080 patients were included, 87.5% female, with a mean age at last evaluation of 60.2 ± 14.6 years and a mean disease duration of 12.4 ± 10.0 years. Limited cutaneous SSc was the most frequent clinical category (57.4%), followed by diffuse cutaneous SSc (17.7%), very early diagnosis of systemic sclerosis (VEDOSS; 12.3%), overlap syndromes (9.8%) and SSc sine scleroderma (2.8%). Antinuclear antibodies were present in 93.4% of patients. Anti-centromere antibodies (ACAs) were the most frequent SSc-specific autoantibodies (54.6%), followed by topoisomerase I antibodies (ATAs; 21.8%). ACA positivity was associated with limited cutaneous disease; older age at diagnosis; and lower frequency of interstitial lung disease (ILD), myositis and flexion contractures. ATA positivity was associated with diffuse cutaneous disease, male sex, higher modified Rodnan skin score (mRSS), digital ulcers, flexion contractures, oesophageal involvement and ILD. Among less frequent autoantibodies, anti-Pm/Scl antibodies were associated with myositis, joint involvement, calcinosis and ILD; anti-U1RNP antibodies with younger age, MCTD overlap, myositis and joint involvement; anti-RNA polymerase III antibodies (ARAs) with scleroderma renal crisis and higher mRSS; anti-U3RNP antibodies with diffuse cutaneous disease and renal involvement; and anti-Ku antibodies with overlap syndromes. Conclusions: In this large real-world SSc cohort, autoantibody status was strongly associated with distinct clinical phenotypes, confirming its value for disease stratification. While most established immuno-clinical associations were reproduced, some differences from international cohorts were observed, supporting the need for population-specific validation of autoantibody associations in SSc. Full article

(This article belongs to the Section Antibody-Based Diagnostics)

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66 pages, 184831 KB

Open AccessReview

Molecular Architecture and Clinical Landscape of Immune Checkpoint Receptors and Ligands

by Milena Czosnek, Agata Sowa, Łucja Rolek, Ewelina Grywalska, Sebastian Mertowski and Paulina Mertowska

Antibodies 2026, 15(5), 84; https://doi.org/10.3390/antib15050084 - 9 Sep 2026

Abstract

Immune checkpoints (ICPs) are essential regulators of immune homeostasis, maintaining the balance between effective immune responses and tolerance to self-antigens. Dysregulation of ICP signaling may contribute to impaired immune surveillance, immune evasion, chronic inflammation, autoimmunity, persistent infections, and tumor progression. Consequently, ICP molecules [...] Read more.

Immune checkpoints (ICPs) are essential regulators of immune homeostasis, maintaining the balance between effective immune responses and tolerance to self-antigens. Dysregulation of ICP signaling may contribute to impaired immune surveillance, immune evasion, chronic inflammation, autoimmunity, persistent infections, and tumor progression. Consequently, ICP molecules are increasingly recognized not only as therapeutic targets but also as potential diagnostic, prognostic, predictive, and treatment-monitoring biomarkers. This review provides a comprehensive overview of the biological functions, signaling mechanisms, and clinical significance of major co-inhibitory and co-stimulatory ICP pathways, including PD-1/PD-L1/PD-L2, CTLA-4/CD28/CD80/CD86, LAG-3, TIM-3, TIGIT, BTLA, VISTA, ICOS, OX40, 4-1BB, GITR, CD27, CD40, and CD2, together with their corresponding ligands. Particular emphasis is placed on their biomarker potential in cancer and immune-mediated diseases. In addition, the review presents a bioinformatic characterization of ICP receptors and ligands based primarily on data available in UniProtKB and complementary bioinformatic resources. The analysis includes protein sequence length, molecular weight, theoretical isoelectric point, amino acid composition, subcellular localization, conserved and functional domains, protein family classification, post-translational modifications, isoforms, and selected structural features. Collectively, the available evidence indicates that ICPs constitute a structurally and functionally diverse group of immunoregulatory molecules with substantial biomarker potential. Integrating their molecular, structural, functional, and bioinformatic characteristics may improve disease classification, prognosis, patient stratification, treatment selection, and therapeutic monitoring. Such an integrated approach may also support the identification of novel biomarkers and therapeutic targets and contribute to the further development of precision and personalized medicine. Full article

(This article belongs to the Special Issue Overcoming Resistance to Immune Checkpoint Blockade: Mechanisms and Novel Therapeutics)

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18 pages, 17386 KB

Open AccessArticle

Ca₁₆Mab-56, a Novel Anti-Cadherin-16/Ksp-Cadherin Monoclonal Antibody for Multiple Applications

by Yukari Ogura, Hiroyuki Suzuki, Mika K. Kaneko and Yukinari Kato

Antibodies 2026, 15(5), 83; https://doi.org/10.3390/antib15050083 - 7 Sep 2026

Abstract

Background/Objectives: Cadherin-16 (CDH16, Ksp-cadherin) possesses unique seven extracellular cadherin repeats, and its expression is restricted to normal kidney epithelium. CDH16 is downregulated in renal cell carcinoma (RCC) and is associated with poor prognosis. Therefore, developing mAbs that specifically recognize cell-surface CDH16 is essential [...] Read more.

Background/Objectives: Cadherin-16 (CDH16, Ksp-cadherin) possesses unique seven extracellular cadherin repeats, and its expression is restricted to normal kidney epithelium. CDH16 is downregulated in renal cell carcinoma (RCC) and is associated with poor prognosis. Therefore, developing mAbs that specifically recognize cell-surface CDH16 is essential for tumor diagnosis and for isolating CDH16-positive renal epithelial cells. Methods: Anti-human CDH16 mAbs (designated as Ca₁₆Mabs) were developed by immunizing mice with CDH16-overexpressed tumor cells, followed by a high-throughput flow cytometry-based screening. Results: Among the 58 established Ca₁₆Mabs, a clone, Ca₁₆Mab-56 (IgG₁, κ), specifically recognized CDH16-overexpressed Chinese hamster ovary-K1 (CHO/CDH16) cells with no detectable cross-reactivity to 21 other CDHs in flow cytometry. Ca₁₆Mab-56 also detected endogenous CDH16 in human RCC cell lines (OS-RC-2 and KMRC-20) and normal kidney epithelial cell lines. The dissociation constant (K_D) values of Ca₁₆Mab-56 for CHO/CDH16 and OS-RC-2 were determined as 6.5 × 10⁻⁹ M and 1.2 × 10⁻⁹ M, respectively. Furthermore, Ca₁₆Mab-56 detected endogenous CDH16 by Western blotting and showed potent staining in normal kidney tubular epithelium and clear membranous staining in renal cell carcinoma in immunohistochemistry. Conclusions: Ca₁₆Mab-56 is a versatile tool for detecting CDH16 and has potential for tumor diagnosis. Full article

(This article belongs to the Section Antibody Discovery and Engineering)

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14 pages, 1480 KB

Open AccessArticle

Seroprevalence of IgG Antibodies to Measles, Mumps, Rubella and Varicella in Adolescents with Documented Two-Dose MMR Vaccination: Cross-Sectional Study

by Mikhail P. Kostinov, Ivan S. Samolygo, Aristitsa M. Kostinova, Natalya L. Starikova, Marina A. Manina, Valentina B. Polishchuk, Albina S. Pestova, Pavel I. Zhuravlev, Anton S. Antishin and Svetlana I. Erdes

Antibodies 2026, 15(5), 82; https://doi.org/10.3390/antib15050082 - 4 Sep 2026

Abstract

Background: Measles, mumps, and rubella outbreaks continue to occur despite high vaccination coverage, raising concerns about long-term vaccine-induced immunity. This study assessed serological protection against measles, mumps, rubella, and varicella in fully vaccinated Russian adolescents to identify immunity gaps and inform revaccination [...] Read more.

Background: Measles, mumps, and rubella outbreaks continue to occur despite high vaccination coverage, raising concerns about long-term vaccine-induced immunity. This study assessed serological protection against measles, mumps, rubella, and varicella in fully vaccinated Russian adolescents to identify immunity gaps and inform revaccination strategies. Methods: Serum IgG was quantified by commercial ELISA. Analytical cut-offs were: measles ≥ 0.18 IU/mL, rubella ≥ 30 IU/mL, mumps index ≥ 1.0 (index = OD/0.467), and varicella ≥ 1.0 IU/mL; equivocal results were classified as negative in the primary analysis and as positive in a sensitivity analysis. Geometric mean concentrations (GMC) were computed on the natural-logarithm scale. Results: Seropositivity differed markedly between infections (χ² 55.1; p < 0.001): rubella 81.4% (95% CI 74.6–86.7), varicella 57.7% (95% CI 49.8–65.2), mumps 53.2% (95% CI 45.4–60.9) and measles 41.0% (95% CI 33.6–48.9). GMC was lowest for measles (0.132; 95% CI 0.111–0.158) and highest for rubella (61.9; 95% CI 55.3–69.2). The measles estimate was sensitive to the treatment of equivocal results, rising to 53.2% (95% CI 45.4–60.9) at the manufacturer’s cut-off of ≥0.12 IU/mL; rubella rose to 94.9% at ≥16 IU/mL. Conclusions: Lower seropositivity rates for measles and mumps were identified among fully vaccinated adolescents, which fall below standard epidemiological benchmarks. In contrast, rubella seropositivity was robust at 81.4%. Because the measles estimate depended strongly on the assay cut-off applied, and because enzyme immunoassays are known to underestimate seroprevalence relative to neutralization testing, these findings describe the distribution of circulating antibody rather than the prevalence of susceptibility and warrant confirmation in larger representative samples with neutralization testing. Full article

(This article belongs to the Special Issue Antiviral Antibody Immune Responses in the Context of Vaccination and Infection)

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34 pages, 511 KB

Open AccessReview

The Evolution of Artificial Intelligence in Antibody Design: From Structure-Based Engineering to Generative Models

by Ida Szataniak and Kacper Packi

Antibodies 2026, 15(5), 81; https://doi.org/10.3390/antib15050081 - 2 Sep 2026

Abstract

Background/Objectives: Artificial intelligence (AI) has transformed computational antibody engineering by enabling accurate prediction of antibody structures, rational optimization of therapeutic properties, and de novo antibody design. Recent advances in deep learning, protein language models, and generative AI have fundamentally changed the way [...] Read more.

Background/Objectives: Artificial intelligence (AI) has transformed computational antibody engineering by enabling accurate prediction of antibody structures, rational optimization of therapeutic properties, and de novo antibody design. Recent advances in deep learning, protein language models, and generative AI have fundamentally changed the way antibodies are discovered and engineered. This review aims to present the historical evolution of computational antibody engineering, from early structure-based design strategies to modern AI-driven approaches, while highlighting the major computational tools, publicly available databases, current limitations, and future directions of the field. Methods: A comprehensive narrative review of the literature was conducted using PubMed, Scopus, Web of Science, and Google Scholar. Original research articles, methodological studies, and review papers published between 1985 and 2026 were evaluated. Publications were selected according to their scientific relevance, methodological quality, and contribution to the historical development of computational antibody engineering. Results: The review describes the progression of antibody engineering from phage display and structure-based computational methods to machine learning, deep learning, protein language models, and generative artificial intelligence. It summarizes key public databases supporting antibody research, discusses advances in antibody structure prediction and developability assessment, and reviews recent generative models capable of designing antibody sequences and structures. Current challenges, including limited experimental validation, dataset bias, prediction of highly flexible regions, model interpretability, and clinical translation, are also discussed. Conclusions: Artificial intelligence has fundamentally reshaped computational antibody engineering by integrating sequence, structural, and functional information into increasingly accurate predictive and generative frameworks. Although important challenges remain, recent developments indicate that AI-driven approaches will play an increasingly central role in the discovery and optimization of next-generation therapeutic antibodies. Full article

(This article belongs to the Section Antibody Discovery and Engineering)

28 pages, 2228 KB

Open AccessReview

Advances in Bispecific Antibodies and Antibody–Drug Conjugates for Colorectal Cancer Treatment

by Maya G. Cappellino, Sean P. Sullivan, Peyton C. High, Tiffani A. Blackburn and Kendra S. Carmon

Antibodies 2026, 15(5), 80; https://doi.org/10.3390/antib15050080 - 1 Sep 2026

Abstract

Bispecific antibodies (bsAbs) and bispecific antibody–drug conjugates (bsADCs) represent promising classes of emerging targeted therapeutics with the potential to overcome tumor heterogeneity and resistance in colorectal cancer (CRC). BsAbs can simultaneously engage multiple tumor antigens or immune cells or bind two distinct epitopes [...] Read more.

Bispecific antibodies (bsAbs) and bispecific antibody–drug conjugates (bsADCs) represent promising classes of emerging targeted therapeutics with the potential to overcome tumor heterogeneity and resistance in colorectal cancer (CRC). BsAbs can simultaneously engage multiple tumor antigens or immune cells or bind two distinct epitopes within a single target, enabling mechanisms of action beyond the capabilities of monoclonal antibodies. Bispecific T cell engagers facilitate targeted destruction of tumors through immune cell recruitment, while dual immune checkpoint inhibitors enhance immune activation by blocking T cell inhibitory signals. Furthermore, bsAbs can mediate dual signaling pathway inhibition through binding multiple receptor tyrosine kinase receptors or other tumor cell surface proteins. BsADCs integrate the dual-antigen recognition of bsAbs with targeted payload delivery, utilizing receptor-mediated endocytosis to deliver potent cytotoxic payloads selectively to CRC cells, while minimizing systemic toxicity. Recent advances in bsAb engineering, linker chemistry, site-specific conjugation, and payload design have accelerated the development of bsADCs for solid tumors, including CRC. BsAbs and bsADCs provide opportunities to improve tumor selectivity, enhance internalization, overcome antigen escape, and expand the population of CRC patients eligible for targeted therapy. Emerging preclinical studies demonstrate encouraging anti-tumor activity for bispecific modalities in CRC, while early clinical trials are beginning to establish their translational potential. This review summarizes the current landscape of bsAbs and bsADCs in therapeutic development for CRC, highlighting key biological targets, engineering strategies, mechanisms of action, and clinical status. We also discuss the major challenges facing clinical translation and provide perspectives on future directions for bispecific therapies in CRC. Full article

(This article belongs to the Section Antibody-Based Therapeutics)

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17 pages, 674 KB

Open AccessArticle

Clinical Characteristics and Risk Factors for Severe Immune-Related Liver Injury Following Nivolumab and Ipilimumab Combination Therapy

by Yuichi Yamazaki, Satoru Kakizaki, Yuki Tamura, Yumeo Tateyama, Tatsuma Murakami, Takuya Adachi, Hiroki Tojima and Toshio Uraoka

Antibodies 2026, 15(5), 79; https://doi.org/10.3390/antib15050079 - 1 Sep 2026

Abstract

Background: Combination therapy with nivolumab and ipilimumab (Nivo-Ipi) has substantially improved outcomes in several advanced malignancies but is frequently associated with early-onset, severe immune-related liver injury (irAE-LI). This study aimed to identify pretreatment predictors of severe irAE-LI and to characterize its clinical [...] Read more.

Background: Combination therapy with nivolumab and ipilimumab (Nivo-Ipi) has substantially improved outcomes in several advanced malignancies but is frequently associated with early-onset, severe immune-related liver injury (irAE-LI). This study aimed to identify pretreatment predictors of severe irAE-LI and to characterize its clinical course. Methods: We retrospectively analyzed 81 patients who received Nivo-Ipi combination therapy between 2018 and 2025. Severe irAE-LI was defined as liver injury requiring systemic corticosteroid therapy. Results: Severe irAE-LI developed in 11 patients (13.6%), with a median onset of 38 days after treatment initiation. More than half of the cases occurred after the first treatment cycle. This exploratory multivariable analysis identified baseline serum Krebs von den Lungen-6 (KL-6) levels of ≥400 U/mL as a potential biomarker associated with a strong preliminary risk signal for severe irAE-LI (odds ratio, 12.60; 95% confidence interval, 2.32–68.60; p = 0.003), whereas conventional liver function tests and inflammatory markers showed no significant association. The robustness of this signal was further confirmed by a sensitivity analysis excluding patients with lung adenocarcinoma (odds ratio 13.21, p = 0.005). Clinically, 54.5% of affected patients experienced steroid-refractory disease or relapse during corticosteroid tapering despite guideline-recommended high-dose corticosteroid therapy. Cytomegalovirus (CMV) reactivation was detected in 50.0% of patients tested on the basis of clinical indicators such as cytopenia or gastrointestinal symptoms, which may have contributed to a pseudo-refractory clinical course in some cases. Conclusions: Pretreatment KL-6 measurement may facilitate risk stratification before Nivo-Ipi therapy. In patients with steroid-refractory or relapsing severe irAE-LI, early introduction of mycophenolate mofetil together with CMV screening may improve clinical management. Full article

(This article belongs to the Topic Antibody-Mediated Therapy and Other Emerging Therapies in Cancer Treatment)

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4 pages, 207 KB

Open AccessEditorial

A Festschrift Celebrating Dr. Dimiter Stanchev Dimitrov: Antibodies, Innovation, and Impact on Infectious Disease and Cancer Research

by Ponraj Prabakaran, Tianlei Ying and Wei Li

Antibodies 2026, 15(5), 78; https://doi.org/10.3390/antib15050078 - 1 Sep 2026

Abstract

Antibody therapeutics now span across oncology, infectious disease, and autoimmunity, and the molecules entering development are increasingly being engineered rather than merely selected [...] Full article

(This article belongs to the Special Issue A Festschrift Celebrating Dr. Dimiter Stanchev Dimitrov: Antibodies, Innovation, and Impact on Infectious Disease and Cancer Research)

19 pages, 1690 KB

Open AccessArticle

An Open-Label, Comparative, Parallel Clinical Trial of the Safety, Pharmacokinetics and Immunogenicity of the Ustekinumab Biosimilar GNR-068 and the Originally Developed Ustekinumab as a Reference Drug After a Single Subcutaneous Administration in Healthy Male Volunteers

by Ivan Lyagoskin, Alena Agafonova, Ivan Shevchenko, Nina Akhtyamova-Givirovskaya, Oksana Markova, Marina Pantyushenko, Rakhim Shukurov and Ravil Khamitov

Antibodies 2026, 15(4), 77; https://doi.org/10.3390/antib15040077 - 12 Aug 2026

Abstract

GNR-068 is a proposed biosimilar to the ustekinumab reference product (RP), which works through the antagonism of interleukin 12 and interleukin 23. Ustekinumab RP is used for the treatment of chronic inflammatory conditions, including certain forms of plaque psoriasis, psoriatic arthritis, Crohn’s disease, [...] Read more.

GNR-068 is a proposed biosimilar to the ustekinumab reference product (RP), which works through the antagonism of interleukin 12 and interleukin 23. Ustekinumab RP is used for the treatment of chronic inflammatory conditions, including certain forms of plaque psoriasis, psoriatic arthritis, Crohn’s disease, and ulcerative colitis. Objectives: The purpose of the study is to study the safety, pharmacokinetics, and immunogenicity of the ustekinumab biosimilar GNR-068 and the reference drug Stelara^® after a single subcutaneous administration in healthy male volunteers. Methods: This was an open-label, randomized, comparative, parallel-group clinical trial of the safety, pharmacokinetics (PK), and immunogenicity of GNR-068 (JSC GENERIUM) and Stelara^® (Manufacturer: Silag AG, Switzerland; RU Holder: JOHNSON & JOHNSON) after a single subcutaneous administration of 45 mg in healthy volunteers. During the trial, 146 volunteers were screened, and 122 were randomized. Results: The results showed that PK similarity was established based on 90% confidence intervals (CIs) for the ratios of geometric means of the primary endpoints of area under the concentration-time curve from time 0 extrapolated to infinity (AUC_0–∞) and maximum observed serum concentration (Cmax) being contained within the pre-specified margin of 80.00–125.00%. The incidence of total ADA was lower in the GNR-068 group compared with the reference product group. Adverse events were similar between treatment groups and consistent with the safety profile of the ustekinumab RP. Conclusions: These results indicate that GNR-068 and the ustekinumab RP share similar PK and safety profiles. Full article

(This article belongs to the Special Issue Immune Phenomena in Autoimmune Skin Disorders)

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18 pages, 10673 KB

Open AccessArticle

A CSF-1R Ig4-5 Domain-Targeting Antibody for the Treatment of Idiopathic Pulmonary Fibrosis

by Wusong Luo, Zhe Shao, Tao Wang, Kenghoe Lok, Rongjing Zhang and Yao Li

Antibodies 2026, 15(4), 76; https://doi.org/10.3390/antib15040076 - 11 Aug 2026

Abstract

Background: Idiopathic pulmonary fibrosis (IPF) is closely associated with fibroblast proliferation, macrophage polarization, and the accumulation of extracellular matrix (ECM). Targeting CSF-1R can rebalance macrophages, reduce ECM deposition, and block pro-fibrotic signaling. Methods: A Fab fragment targeting CSF-1R was screened via phage display. [...] Read more.

Background: Idiopathic pulmonary fibrosis (IPF) is closely associated with fibroblast proliferation, macrophage polarization, and the accumulation of extracellular matrix (ECM). Targeting CSF-1R can rebalance macrophages, reduce ECM deposition, and block pro-fibrotic signaling. Methods: A Fab fragment targeting CSF-1R was screened via phage display. After sequence optimization, it was constructed into an IgG1 antibody (BC006). In vitro activity, domain binding, and signaling blockade were investigated, and in vitro safety indicators were evaluated. Efficacy was assessed using an induced IPF organoid-on-a-chip and bleomycin-induced mouse model. In vivo safety evaluation was conducted in cynomolgus monkeys. Results: BC006 showed an EC₅₀ value of 226 ± 57 nM and a KD of 30 ± 2 nM. It specifically bound to the Ig4-5 domain of CSF-1R by inhibiting receptor dimerization without blocking ligand binding. It could dose-dependently inhibit the differentiation of monocytes into M2 macrophages and exert anti-IPF effects. In the induced IPF organoid-on-a-chip model, BC006 maintained lung barrier function and decreased α-SMA and Collagen I. It also improved lung function and attenuated the degree of fibrosis in the mouse model. Moreover, BC006 had no ADCC, CDC, cytokine release, or hemagglutination, and demonstrated favorable safety profiles in cynomolgus monkeys. Conclusions: BC006 is a novel anti-fibrosis antibody specifically targeting the CSF-1R Ig4-5 domain and offers a new therapeutic strategy for IPF. Full article

(This article belongs to the Section Antibody-Based Therapeutics)

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