Comparative In Vitro Apoptotic Activity of Two Chimeric Anti-CD99 Antibodies Recognizing Distinct CD99 Regions in T-ALL Models

Juengsamretkarn, Phasinee; Sampaoloi, Phakhwan; Ruangkul, Kadkanok; Chinakarapong, Manatchanok; Pliensak, Praweekorn; Thu, Myint Myat; Chokepaichitkool, Tawan; Pata, Supansa; Laopajon, Witida; Kasinrerk, Watchara; Takheaw, Nuchjira

doi:10.3390/biomedicines14091913

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Comparative In Vitro Apoptotic Activity of Two Chimeric Anti-CD99 Antibodies Recognizing Distinct CD99 Regions in T-ALL Models

by

Phasinee Juengsamretkarn

^1,2,

Phakhwan Sampaoloi

¹,

Kadkanok Ruangkul

^1,3,

Manatchanok Chinakarapong

²

,

Praweekorn Pliensak

²,

Myint Myat Thu

²,

Tawan Chokepaichitkool

⁴,

Supansa Pata

^1,2

,

Witida Laopajon

^1,2

,

Watchara Kasinrerk

² and

Nuchjira Takheaw

^1,2,*

¹

Division of Clinical Immunology, Department of Medical Technology, Faculty of Associated Medical Sciences, Chiang Mai University, Chiang Mai 50200, Thailand

²

Biomedical Technology Research Center, Faculty of Associated Medical Sciences, Chiang Mai University, Chiang Mai 50200, Thailand

³

Department of Clinical Pathology, Central Chest Institute of Thailand, Nonthaburi 11000, Thailand

⁴

Thailand Excellence Center for Tissue Engineering and Stem Cells, Department of Biochemistry, Faculty of Medicine, Chiang Mai University, Chiang Mai 50200, Thailand

^*

Author to whom correspondence should be addressed.

Biomedicines 2026, 14(9), 1913; https://doi.org/10.3390/biomedicines14091913

Submission received: 24 June 2026 / Revised: 20 August 2026 / Accepted: 25 August 2026 / Published: 26 August 2026

(This article belongs to the Special Issue Emerging Trends in Cancer: Biomarkers, Therapeutic Strategies and Future Directions)

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Abstract

Background/Objectives: Therapeutic antibodies have become an important modality for cancer treatment; however, effective targeted antibody therapies for T-cell acute lymphoblastic leukemia (T-ALL) remain limited. CD99 is a promising therapeutic target due to its high expression in leukemic T cells and its capacity to transmit apoptotic signals. However, the efficacy of antibody-mediated apoptosis may depend on the CD99 region recognized by the antibody. This study compared the in vitro apoptotic activity of two chimeric anti-CD99 antibodies, ChAbMT99/1 and ChAbMT99/3, which recognize distinct regions of CD99 in T-ALL models. Methods: ChAbMT99/1 and ChAbMT99/3 were generated as human IgG1 antibodies. Antibody reactivity and binding site specificity were characterized using peptide-based ELISA. Binding to native CD99, apoptosis induction in two-dimensional (2D) Jurkat E6.1 and MOLT-4 suspension cultures and three-dimensional (3D) Jurkat E6.1-derived spheroid models, and cytotoxic effects on peripheral blood mononuclear cells (PBMCs) were assessed by flow cytometry. In vitro hematologic effects were assessed using hemagglutination and platelet aggregation assays. Results: ChAbMT99/1 and ChAbMT99/3 specifically recognized peptides corresponding to distinct CD99 regions, with peptide-binding EC₅₀ values of 0.813 and 0.819 μg/mL, respectively. Both antibodies exhibited binding reactivity to native CD99 on the tested cells. Functionally, both antibodies induced Annexin V/7-AAD-defined cell death in both 2D and 3D T-ALL models compared with controls in a crosslinking-dependent manner. In contrast, both antibodies induced low levels of cell death (<10%) in bulk PBMCs, with no visible hemagglutination or platelet aggregation observed in samples from five selected donors under the stated assay conditions. Conclusions: Both chimeric anti-CD99 antibodies demonstrated in vitro apoptotic activity against T-ALL models despite recognizing distinct regions of CD99. These findings provide a rationale for further investigation of their other mechanisms of action to support the future development of CD99-targeted antibody therapy for T-ALL.

Keywords: CD99; therapeutic antibody; apoptosis; T-ALL; chimeric antibody

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MDPI and ACS Style

Juengsamretkarn, P.; Sampaoloi, P.; Ruangkul, K.; Chinakarapong, M.; Pliensak, P.; Thu, M.M.; Chokepaichitkool, T.; Pata, S.; Laopajon, W.; Kasinrerk, W.; et al. Comparative In Vitro Apoptotic Activity of Two Chimeric Anti-CD99 Antibodies Recognizing Distinct CD99 Regions in T-ALL Models. Biomedicines 2026, 14, 1913. https://doi.org/10.3390/biomedicines14091913

AMA Style

Juengsamretkarn P, Sampaoloi P, Ruangkul K, Chinakarapong M, Pliensak P, Thu MM, Chokepaichitkool T, Pata S, Laopajon W, Kasinrerk W, et al. Comparative In Vitro Apoptotic Activity of Two Chimeric Anti-CD99 Antibodies Recognizing Distinct CD99 Regions in T-ALL Models. Biomedicines. 2026; 14(9):1913. https://doi.org/10.3390/biomedicines14091913

Chicago/Turabian Style

Juengsamretkarn, Phasinee, Phakhwan Sampaoloi, Kadkanok Ruangkul, Manatchanok Chinakarapong, Praweekorn Pliensak, Myint Myat Thu, Tawan Chokepaichitkool, Supansa Pata, Witida Laopajon, Watchara Kasinrerk, and et al. 2026. "Comparative In Vitro Apoptotic Activity of Two Chimeric Anti-CD99 Antibodies Recognizing Distinct CD99 Regions in T-ALL Models" Biomedicines 14, no. 9: 1913. https://doi.org/10.3390/biomedicines14091913

APA Style

Juengsamretkarn, P., Sampaoloi, P., Ruangkul, K., Chinakarapong, M., Pliensak, P., Thu, M. M., Chokepaichitkool, T., Pata, S., Laopajon, W., Kasinrerk, W., & Takheaw, N. (2026). Comparative In Vitro Apoptotic Activity of Two Chimeric Anti-CD99 Antibodies Recognizing Distinct CD99 Regions in T-ALL Models. Biomedicines, 14(9), 1913. https://doi.org/10.3390/biomedicines14091913

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Comparative In Vitro Apoptotic Activity of Two Chimeric Anti-CD99 Antibodies Recognizing Distinct CD99 Regions in T-ALL Models

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