Deniz Gören ŞAHİN, Ayla KORKMAZ, Sema MERİÇ, Günhan GÜRMAN, Muhit ÖZCAN, Önder ARSLAN, Mutlu ARAT

Flow Cytometric Analysis of T Lymphocyte Activation in CML Patients Under Imatinib Therapy

To analyze T cell functions by flow cytometry and to evaluate the possible functional changes that might occur under imatinib therapy in CML patients. A total of 29 patients and 9 healthy control subjects were enrolled. Newly diagnosed patients having no treatment (group 1), patients receiving imatinib for 1 year (group 2) and patients receiving imatinib more than 1 year (group 3), healthy control subjects (group 4). IL-4 and IFN gamma expression on CD4+ cells; how much percentage of CD3+ T cells were activated (CD3+CD69+); CD8+ T cells and the ratio and grade of expression of HLA-ABC and HLA-DR on those cells were evaluated, respectively. There was no significant difference in terms of mean number of CD4+ cells between the groups. However, there was a tendency towards higher CD4+ cells in control group. IL-4 and IFN gamma were found not to be statistically significant between the groups. Control group has lower IL-4 and IFN gamma expression values. Mean number of CD4+ cells, which did not express IL-4 and IFN gamma, were statistically higher in control group when compared to other groups. In control group, % activation was decreased when compared to that of other groups. CD8+ cell ratio was found to be statistically lower in all patient groups (p=0.001). The expression of HLA-ABC and HLA-DR on CD8+ cells were similar between the groups. We could not show any inhibitory effect of imatinib on T cell functions in concordance with clinical experience and safety profile.

I matinib Tedavisi Alan Hastalarda T Lenfosit Aktivasyonunun Akım Sitometrik Analizi

İmatinibin in vivo ortamda T lenfosit fonksiyonları üzerine olan etkileri akım sitometrik olarak değerlendirilmiştir. Çalışmaya toplam 29 KML hastası ve 9 sağlıklı birey dahil edilmiştir. KML hastaları kendi aralarında sırasıyla, yeni tanı almış ve tedavi görmeyen, 1 yıldır imatinib alan ve 1 yıldan uzun süredir imatinib alan hastalar olarak 3 gruba ayrılmıştır. CD4+ hücreler ve bu hücrelerdeki IL-4 ve IFNg ifadeleri, CD3+ T lenfositlerin % kaçının aktive edilmiş olduğu (CD3+CD69+), CD8+ hücreler ve bu hücrelerde HLA-ABC ve HLA-DR ifade eden hücrelerin oranı ve HLA-ABC ve HLA-DR ifadelerinin şiddeti değerlendirilmiştir. Gruplar arasında ortalama CD4 + hücre sayısı açısından anlamlı fark yoktu. Bununla birlikte, kontrol grubunda daha yüksek CD4 + hücrelerine doğru bir eğilim vardı. IL-4 ve IFN gama, gruplar arasında istatistiksel olarak anlamlı bulunmadı. Kontrol grubu daha düşük IL-4 ve IFN gama ifade değerlerine sahipti. IL-4 ve IFN gama ifade etmeyen ortalama CD4 + hücre sayısı kontrol grubunda diğer gruplara göre istatistiksel olarak daha yüksekti. Kontrol grubunda ise % aktivasyon diğer gruplara göre azdı. Tüm hasta gruplarında CD8 + hücre oranı istatistiksel olarak daha düşük bulundu (p = 0,001). HLA-ABC ve HLA-DR'nin CD8 + hücrelerinde ekspresyonu gruplar arasında benzerdi. İmatinibin sitokin sentezi üzerine herhangi bir etkisi gözlenmemiştir. İmatinib tedavisi altındaki hastalarda T lenfosit fonksiyonlarının etkilendiğine dair klinik belirgin gözlemler olmamakla birlikte, subklinik etki varlığı araştırma konusudur. Yapılan in vitro çalışmaların in-vivo korele olup olmadığının aydınlatılabilmesi için daha ileri çalışmalara gereksinim vardır.

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1. Shtivelman E, Lifshitz B, Gale RP, Canaani E. Fused transcript of abl and bcr genes in chronic myelogenous leukaemia. Nature. 1985;315:550-4.

2. Kurzrock R, Gutterman JU, Talpaz M. The molecular genetics of Philadelphia chromosome-positive leukemias. N Engl J Med. 1988;319:990-8.

3. Lugo TG, Pendergast AM, Muller AJ, Witte ON. Tyrosine kinase activity and transformation potency of bcr-abl oncogene products. Science. 1990;247:1079-82.

4. Druker BJ, Tamura S, Buchdunger E, et al. Effects of a selective inhibitor of the Abl tyrosine kinase on the growth of Bcr-Abl positive cells. Nat Med. 1996;2:561-6.

5. Kantarjian HM, O'Brien S, Cortes J, et al. Imatinib mesylate therapy improves survival in patients with newly diagnosed Philadelphia chromosome-positive chronic myelogenous leukemia in the chronic phase: comparison with historic data. Cancer. 2003;98:2636-42.

6. Druker BJ, Guilhot F, O'Brien SG, Gathmann I, Kantarjian H, Gattermann N, et al. Five-year follow-up of patients receiving imatinib for chronic myeloid leukemia. N Engl J Med. 2006;355:2408-17.

7. Barrett AJ. Mechanisms of the graft-versusleukemia reaction. Stem Cells. 1997;15:248- 58.

8. Cwynarski K, Laylor R, Macchiarulo E, Goldman J, Lombardi G, Melo JV, et al. Imatinib inhibits the activation and proliferation of normal T lymphocytes in vitro. Leukemia. 2004;18:1332-9.

9. Seggewiss R, Lore K, Greiner E, Magnusson MK, Price DA, Douek DC, et al. Imatinib inhibits T-cell receptor-mediated T-cell proliferation and activation in a dosedependent manner. Blood. 2005;105:2473-9.

10. Aswald JM, Lipton JH, Aswald S, Messner HA. Increased IFN-gamma synthesis by T cells from patients on imatinib therapy for chronic myeloid leukemia. Cytokines, cellular & molecular therapy. 2002;7:143-9.

11. Pendergast AM, Muller AJ, Havlik MH, Maru Y, Witte ON. BCR sequences essential for transformation by the BCR-ABL oncogene bind to the ABL SH2 regulatory domain in a non-phosphotyrosine-dependent manner. Cell. 1991;66:161-71.

12. Lim SH, Coleman S. Chronic myeloid leukemia as an immunological target. Am J Hematol. 1997;54:61-7.

13. Clark RE, Christmas SE. BCR-ABL fusion peptides and cytotoxic T cells in chronic myeloid leukaemia. Leuk Lymphoma. 2001;42:871-80.

14. Garicochea B, Chase A, Lazaridou A, Goldman JM. T lymphocytes in chronic myelogenous leukaemia (CML): no evidence of the BCR/ABL fusion gene detected by fluorescence in situ hybridization in 14 patients. Leukemia. 1994;8:1197-201.

15. Gao H, Lee BN, Talpaz M, Donato NJ, Cortes JE, Kantarjian HM, et al. Imatinib mesylate suppresses cytokine synthesis by activated CD4 T cells of patients with chronic myelogenous leukemia. Leukemia. 2005;19:1905-11.

16. Tsuda H, Yamasaki H. Type I and type II T cell profiles in chronic myelogenous leukemia. Acta haematologica. 2000;103:96-101.

17. Leder C, Ortler S, Seggewiss R, Einsele H, Wiendl H. Modulation of T-effector function by imatinib at the level of cytokine secretion. Exp Hematol. 2007;35:1266-71.

18. Dietz AB, Souan L, Knutson GJ, et al. Imatinib mesylate inhibits T-cell proliferation in vitro and delayed-type hypersensitivity in vivo. Blood. 2004;104:1094-9.

19. Appel S, Balabanov S, Brummendorf TH, Brossart P. Effects of imatinib on normal hematopoiesis and immune activation. Stem cells (Dayton, Ohio). 2005;23:1082-8.

20. Chen J, Schmitt A, Chen B, Rojewski M, Ringhoffer M, von Harsdorf S, et al. Imatinib impairs CD8+ T lymphocytes specifically directed against the leukemia-associated antigen RHAMM/CD168 in vitro. Cancer Immunol Immunother. 2007;56:849-61.

21. Brauer KM, Werth D, von Schwarzenberg K, Bringmann A, Kanz L, Grunebach F, et al. BCR-ABL activity is critical for the immunogenicity of chronic myelogenous leukemia cells. Cancer research. 2007;67:5489-97.

22. Mumprecht S, Matter M, Pavelic V, Ochsenbein AF. Imatinib mesylate selectively impairs expansion of memory cytotoxic T cells without affecting the control of primary viral infections. Blood. 2006;108:3406-13.