Perforin and Granzyme Levels in CXCR5⁺CD8⁺ Follicular Cytotoxic T Cells, CD8⁺ T Cells, and NK Cells in Chronic Lymphocytic Leukemia Patients

Year 2025, Volume: 8 Issue: 3, 308 - 313, 22.10.2025

Metin Yusuf Gelmez , Fatma Betul Oktelık , Murat Özbalak , İpek Yönal Hindilerden , Günnur Deniz

https://doi.org/10.53446/actamednicomedia.1691789

Abstract

Objective: Chronic lymphocytic leukemia (CLL) is a type of leukemia characterized by the accumulation of CD19+ malignant B cells in lymph nodes, blood, and bone marrow, which aberrantly express the T-cell marker CD5. Although several studies have investigated the frequencies and functions of CD8+ T, NK, and follicular cytotoxic T (TFC) cells in CLL patients, no study has specifically compared the perforin and granzyme contents of these cells. This study aimed to analyse the levels of perforin and granzyme in CD8+ T, NK, and TFC cells in CLL patients.
Methods: A total of 14 untreated CLL patients and 9 age-matched healthy individuals were enrolled in this study. The proportions of CD8+ T, NK, and TFC cells, as well as their perforin and granzyme levels, were determined using flow cytometry.
Results: Decreased NK cells and increased TFC and CD8+ T cells were observed in CLL patients. Compared to healthy individuals, granzyme B levels were lower in NK cells but higher in TFC cells of CLL patients, while perforin levels were elevated in all three cell groups. When comparing perforin and granzyme content among CD8+ T, NK, and TFC cells in CLL patients, NK and TFC cells exhibited higher perforin levels than CD8+ T cells.
Conclusion: The reduced number and low perforin content of NK cells in CLL patients suggest that their role in CLL pathogenesis may be more limited compared to other cytotoxic cells.
Our findings indicate that TFC cells may be more effective in targeting malignant B cells in CLL pathogenesis and could be considered as a potential target for immunotherapy.
with high perforin and granzyme levels, along with their higher perforin content compared to CD8+ T cells, suggests that TFC cells might be more effective in the lysis of malign B-CLL cells in CLL and could represent a potential new cell population for use in immunotherapies.

Keywords

Chronic lymphocytic leukemia , CD8+ T , NK , Follicular cytotoxic T cells , TFC

Kronik Lenfositik Lösemi Hastalarında CXCR5+CD8+ Foliküler Sitotoksik T hücreleri, CD8+ T hücreleri ve NK Hücrelerinde Perforin ve Granzim Seviyeleri

Abstract

Amaç: Kronik lenfositik lösemi (KLL), bir T hücre belirteci olan CD5 proteinini aberan olarak ifade eden CD19+ malign B hücrelerin lenf nodu, kan ve kemik iliğinde biriktiği lösemi tipidir. KLL hastalarında CD8+ T, NK ve foliküler sitotoksik T (TFC) hücre oranları ve fonksiyonlarını inceleyen farklı çalışmalar bulunmakla birlikte, bu hücrelerin perforin ve granzim içeriklerini karşılaştıran bir çalışma bulunmamaktadır. Bu çalışmada KLL hastalarında CD8+ T, NK ve TFC hücrelerin perforin ve granzim seviyeleri incelenmiştir.
Yöntem: Tedavi almayan KLL tanılı 14 hasta ve 9 sağlıklı birey çalışmaya dahil edilmiştir. CD8+ T, NK ve TFC hücre oranları ve bu hücrelerde perforin ve granzim seviyeleri akan hücre ölçer sistemi ile saptanmıştır.
Bulgular: KLL hastalarında NK hücrelerinin azaldığı, buna karşılık CD8+ T ve TFC hücrelerinin arttığı tespit edilmiştir. Sağlıklı bireyler ile karşılaştırıldığında, KLL hastalarının NK hücrelerinde granzim B seviyesinin düşük, TFC hücrelerinde ise yüksek olduğu gözlenirken, perforin seviyesinin her üç hücre grubunda da arttığı belirlenmiştir. KLL hastalarının CD8+ T, NK ve TFC hücrelerindeki perforin ve granzim içerikleri kendi aralarında değerlendirildiğinde, NK ve TFC hücrelerinde CD8+ T hücrelerine göre daha yüksek perforin seviyelerine sahip olduğu gözlenmiştir.
Sonuç: KLL hastalarında NK hücrelerinin sayıca azalması ve düşük perforin içerikleri, NK hücrelerinin KLL patogenezindeki rolünün diğer sitotoksik hücrelere göre daha sınırlı olabileceğini düşündürmektedir. Bulgularımız TFC hücrelerin KLL patogenezinde malign B hücrelerine karşı daha etkili olabileceğini ve immünoterapilerde hedeflenebilecek potansiyel bir hücre grubu olarak değerlendirilebileceğini göstermektedir.

Keywords

Kronik lenfositik lösemi , CD8+ T , NK , Foliküler sitotoksik T hücreler , TFC

Ethical Statement

Çalışmaya katılan tüm gönüllülerden bilgilendirilmiş onam formları alınmıştır (İ.Ü. İTF Etik Kurulu, no:1302, tarih:06.11.2017).

Supporting Institution

Bu çalışma İstanbul Üniversitesi Bilimsel Araştırma Projeleri Koordinasyon Birimi (Proje no: TDP-2019-32394) tarafından desteklenmiştir.

Thanks

Bu çalışmaya sağladığı katkılar için merhum Prof. Dr. Melih AKTAN’a en içten teşekkürlerimizi sunarız.

References

• Cao J, Yan Q. Cancer Epigenetics, Tumor Immunity, and Immunotherapy. Trends Cancer. Jul 2020;6(7):580-592. doi:10.1016/j.trecan.2020.02.003
• Rosenberg J, Huang J. CD8(+) T Cells and NK Cells: Parallel and Complementary Soldiers of Immunotherapy. Curr Opin Chem Eng. Mar 2018;19:9-20. doi:10.1016/j.coche.2017.11.006
• Lv Y, Ricard L, Gaugler B, Huang H, Ye Y. Biology and clinical relevance of follicular cytotoxic T cells. Front Immunol. 2022;13:1036616. doi:10.3389/fimmu.2022.1036616
• Yu D, Ye L. A Portrait of CXCR5(+) Follicular Cytotoxic CD8(+) T cells. Trends Immunol. Dec 2018;39(12):965-979. doi:10.1016/j.it.2018.10.002
• Kikushige Y. Pathogenesis of chronic lymphocytic leukemia and the development of novel therapeutic strategies. J Clin Exp Hematop. Dec 15 2020;60(4):146-158. doi:10.3960/jslrt.20036
• Braish J, Cerchione C, Ferrajoli A. An overview of prognostic markers in patients with CLL. Front Oncol. 2024;14:1371057. doi:10.3389/fonc.2024.1371057
• Brown JR. Clinical Risks for Chronic Lymphocytic Leukemia. J Natl Compr Canc Netw. Apr 2024;22(3)doi:10.6004/jnccn.2024.7020
• Metin Yusuf Gelmez SÇ, Aynur Dağlar-Aday , Gülce Özçit-Gürel , İpek Yönal-Hindilerden,Günnur Deniz, Melih Aktan. Evaluation of T Lymphocyte Subgroups in Patients with Chronic Lymphocytic Leukemia. Turkish Journal of Immunology. 2020;8(1):6. doi:https://doi.org/10.25002/tji.2020.1240
• Yano M, Byrd JC, Muthusamy N. Natural Killer Cells in Chronic Lymphocytic Leukemia: Functional Impairment and Therapeutic Potential. Cancers (Basel). Nov 24 2022;14(23)doi:10.3390/cancers14235787
• Gelmez MY, Oktelik FB, Cinar S, et al. High expression of OX-40, ICOS, and low expression PD-L1 of follicular helper and follicular cytotoxic T cells in chronic lymphocytic leukemia. J Hematop. Sep 2022;15(3):117-129. doi:10.1007/s12308-022-00497-5
• Taghiloo S, Asgarian-Omran H. Cross-talk between leukemic and immune cells at the tumor microenvironment in chronic lymphocytic leukemia: An update review. Eur J Haematol. Jul 2024;113(1):4-15. doi:10.1111/ejh.14224
• Tang J, Zha J, Guo X, Shi P, Xu B. CXCR5(+)CD8(+) T cells present elevated capacity in mediating cytotoxicity toward autologous tumor cells through interleukin 10 in diffuse large B-cell lymphoma. Int Immunopharmacol. Sep 2017;50:146-151. doi:10.1016/j.intimp.2017.06.020
• Gelmez MY, Çınar, S., Dağlar-Aday, A., Özçit-Gürel, G., Yönal-Hindilerden, İ., Deniz, G., Aktan, M. Evaluation of T Lymphocyte Subgroups in Patients with Chronic Lymphocytic Leukemia. Turkish Journal of Immunology. 2020;8(1):1-7. doi:/10.25002/tji.2020.1240
• Riches JC, Davies JK, McClanahan F, et al. T cells from CLL patients exhibit features of T-cell exhaustion but retain capacity for cytokine production. Blood. Feb 28 2013;121(9):1612-21. doi:10.1182/blood-2012-09-457531
• Philip M, Schietinger A. CD8(+) T cell differentiation and dysfunction in cancer. Nat Rev Immunol. Apr 2022;22(4):209-223. doi:10.1038/s41577-021-00574-3
• Peters FS, Strefford JC, Eldering E, Kater AP. T-cell dysfunction in chronic lymphocytic leukemia from an epigenetic perspective. Haematologica. May 1 2021;106(5):1234-1243. doi:10.3324/haematol.2020.267914
• Rubino V, Carriero F, Palatucci AT, et al. Adaptive and Innate Cytotoxic Effectors in Chronic Lymphocytic Leukaemia (CLL) Subjects with Stable Disease. Int J Mol Sci. May 31 2023;24(11)doi:10.3390/ijms24119596
• MacFarlane AWt, Jillab M, Smith MR, et al. NK cell dysfunction in chronic lymphocytic leukemia is associated with loss of the mature cells expressing inhibitory killer cell Ig-like receptors. Oncoimmunology. 2017;6(7):e1330235. doi:10.1080/2162402X.2017.1330235
• Akboga F, Hindilerden F, Gulturk E, et al. Increased Perforin- and IL-21-Expressing NK Cells in Patients with Early-Stage Chronic Lymphocytic Leukemia. Experimed. 2022;12(3):225-231.
• Pan K, Farrukh H, Chittepu V, Xu H, Pan CX, Zhu Z. CAR race to cancer immunotherapy: from CAR T, CAR NK to CAR macrophage therapy. J Exp Clin Cancer Res. Mar 31 2022;41(1):119. doi:10.1186/s13046-022-02327-z
• Todorovic Z, Todorovic D, Markovic V, et al. CAR T Cell Therapy for Chronic Lymphocytic Leukemia: Successes and Shortcomings. Curr Oncol. May 18 2022;29(5):3647-3657. doi:10.3390/curroncol29050293
• Herrera L, Santos S, Vesga MA, et al. The Race of CAR Therapies: CAR-NK Cells for Fighting B-Cell Hematological Cancers. Cancers (Basel). Oct 28 2021;13(21)doi:10.3390/cancers13215418