Preparation and characterization of a novel nanobody against T-cell immunoglobulin and mucin-3 (TIM-3) | ||
| Iranian Journal of Basic Medical Sciences | ||
| مقاله 8، دوره 19، شماره 11، بهمن 2016، صفحه 1201-1208 اصل مقاله (1.09 M) | ||
| نوع مقاله: Original Article | ||
| شناسه دیجیتال (DOI): 10.22038/ijbms.2016.7820 | ||
| نویسندگان | ||
| Vida Homayouni1؛ Mazdak Ganjalikhani-hakemi* 1؛ Abbas Rezaei1؛ Hossein Khanahmad2؛ Mahdi Behdani3؛ Fatemeh Kazemi Lomedasht3 | ||
| 1Immunology Department, Faculty of Medicine, Isfahan University of Medical Sciences, Isfahan, Iran | ||
| 2Genetic Department, Faculty of Medicine, Isfahan University of Medical Sciences, Isfahan, Iran | ||
| 3Biotechnology Research Center, Biotechnology Department, Venom & Bio-therapeutics Molecules Lab, Pasteur Institute of Iran, Tehran, Iran | ||
| چکیده | ||
| Objective(s): As T-cell immunoglobulin and mucin domain 3 (TIM-3) is an immune regulatory molecule; its blocking or stimulating could alter the pattern of immune response towards a desired condition. Based on the unique features of nanobodies, we aimed to construct an anti-TIM-3 nanobody as an appropriate tool for manipulating immune responses for future therapeutic purposes. Materials and Methods:We immunized a camel with TIM-3 antigen and then, synthesized a VHH phagemid library from its B cell’s transcriptome using nested PCR. Library selection against TIM-3antigen was performed in three rounds of panning. Using phage-ELISA, the most reactive colonies were selected for sub-cloning in soluble protein expression vectors. The Nanobody was purified and confirmed with a nickel-nitrilotriacetic acid (Ni-NTA) column, SDS-PAGE and Western blotting. A flowcytometric analysis was performed to analyze the binding and biologic activities of theTIM-3 specific nanobody with TIM-3 expressing HL-60 and HEK cell lines. Results:Specific 15kD band representing for nanobody was observed on the gel and confirmed with Western blotting. The nanobody showed significant specific immune-reactivity against TIM-3 with a relatively high binding affinity. The nanobody significantly suppressed the proliferation of TIM-3 expressing HL-60 cell line. Conclusion: Finally, we successfully prepared a functional anti-humanTIM-3 specific nanobody with a high affinity and an anti-proliferative activity on an AML cell line in vitro. | ||
| کلیدواژهها | ||
| antibody؛ Heavy chain antibody؛ Nanobody؛ Phage display؛ T-cell immunoglobulin and mucin domain 3 | ||
| مراجع | ||
|
1. Kane LP. T cell Ig and mucin domain proteins and immunity. J Immunol 2010; 184:2743-2749.
2. Umetsu DT, Umetsu SE, Freeman GJ, DeKruyff RH.TIM gene family and their role in atopic diseases. Curr Top Microbiol Immunol 2008; 321:201-215.
3. Tong D, Zhou Y, Chen W, Deng Y, Li L, Jia Z, et al. T cell immunoglobulin- and mucin-domain-containing molecule 3 gene polymorphisms and susceptibility to pancreatic cancer. Mol Biol Rep 2012; 39:9941-9946.
4. Monney L, Sabatos CA, Gaglia JL, Ryu A, Waldner H, Chernova T, et al. Th1-specific cell surface protein Tim-3 regulates macrophage activation and severity of an autoimmune disease. Nature 2002; 415:536-541.
5. Tang ZH, Liang S, Potter J, Jiang X, Mao HQ, Li Z. Tim-3/galectin-9 regulate the homeostasis of hepatic NKT cells in a murine model of nonalcoholic fatty liver disease. J Immunol 2013; 190:1788-1796.
6. Anderson AC, Anderson DE, Bregoli L, Hastings WD, Kassam N, Lei C, et al. Promotion of tissue inflammation by the immune receptor Tim-3 expressed on innate immune cells. Science 2007; 318:1141-1143.
7. Nagahara K, Arikawa T, Oomizu S, Kontani K, Nobumoto A, Tateno H, et al. Galectin-9 increases Tim-3+ dendritic cells and CD8+ T cells and enhances antitumor immunity via galectin-9-Tim-3 interactions. J Immunol 2008; 181:7660-7669.
8. Wherry EJ. T cell exhaustion. Nat Immunol 2011; 12:492-499.
9. Le Mercier I, Lines JL, Noelle RJ. Beyond CTLA-4 and PD-1, the Generation Z of Negative Checkpoint Regulators. Front Immunol 2015; 6:418.
10. Ngiow SF, von Scheidt B, Akiba H, Yagita H, Teng MW, Smyth MJ. Anti-TIM3 antibody promotes T Cell IFN-g-mediated antitumor immunity and suppresses established tumors. Cancer Res 2011; 71:3540-3551.
11. Baghdadi M, Takeuchi S, Wada H, Seino K. Blocking monoclonal antibodies of TIM proteins as orchestrators of anti-tumor immune response. MAbs 2014; 6:1124-1132.
12. Jin HT, Anderson AC, Tan WG, West EE, Ha SJ, Araki K, et al. Cooperation of Tim-3 and PD-1 in CD8 T-cell exhaustion during chronic viral infection. Proc Natl Acad of Sci U S A 2010; 107:14733-1478.
13. Dietze KK, Zelinskyy G, Liu J, Kretzmer F, Schimmer S, Dittmer U. Combining regulatory T cell depletion and inhibitory receptor blockade improves reactivation of exhausted virus-specific CD8 (+) T cells and efficiently reduces chronic retroviral loads. PLoS Pathog 2013; 9:e1003798.
14. Sakuishi K, , Apetoh L, Sullivan JM, Blazar BR, Kuchroo VK, Anderson AC. Targeting Tim-3 and PD-1 pathways to reverse T cell exhaustion and restore anti-tumor immunity. J Exp Med 2010; 207:2187- 2194.
15. Sakuishi K, Ngiow SF, Sullivan JM, Teng MW, Kuchroo VK, Smyth MJ, Anderson AC. TIM3FOXP3 regulatory T cells are tissue-specific promoters of T-cell dysfunction in cancer. Oncoimmunology 2013; 2:e23849.
16. Gonçalves Silva I, Gibbs BF, Bardelli M, Varani L, Sumbayev VV. Differential expression and biochemical activity of the immune receptor Tim-3 in healthy and malignant human myeloid cells. Oncotarget 2015; 6:33823-33833.
17. Anderson AC. Tim-3, a negative regulator of anti-tumor immunity. Curr Opin Immunol 2012; 24:213-216.
18. Freeman GJ, Casasnovas JM, Umetsu DT, DeKruyff RH. TIM genes:a family of cell surface phosphatidylserine receptors that regulate innate and adaptive immunity. Immunol Rev 2010; 235:172-189.
19. Siontorou CG. Nanobodies as novel agents for disease diagnosis and therapy. Int J Nanomed 2013; 8:4215–4227. .
20. Bell A, Wang ZJ, Arbabi-Ghahroudi M, Chang TA, Durocher Y, Trojahn U, et al. Differential tumor-targeting abilities of three single-domain antibody formats. Cancer Lett 2010; 289:81–90.
21. Hamers-Casterman C, Atarhouch T, Muyldermans S, Robinson G, Hamers C, Songa EB, et al. Naturally occuring antibodies devoid of light chains. Nature 1993; 363:446-448.
22. Yardehnavi N, Behdani M, Bagheri KP, Mahmoodzadeh A, Khanahmad H, Shahbazzadeh D, et al. A camelid antibody candidate for development of a therapeutic agent against Hemiscorpius lepturus envenomation. FASEB J 2014; 28:4004-4014.
23. Muyldermans S. Single domain camel antibodies: current status. J Biotechnol 2001; 74:277-302.
24. DeKruyff RH, Bu X, Ballesteros A, Santiago C, Chim YL, Lee HH, et al. T cell/transmembrane, Ig, and mucin-3 allelic variants differentially recognize phosphatidylserine and mediate phagocytosis of apoptotic cells. J Immunol 2010; 184:1918-1930.
25. Cao E, Zang X, Ramagopal UA, Mukhopadhaya A, Fedorov A, Fedorov E, et al. T cell immunoglobulin mucin-3 crystal structure reveals a galectin-9-independent ligand-binding surface. Immunity 2007; 26:311-321.
26. Chiba S, Chiba S, Baghdadi M, Akiba H, Yoshiyama H, Kinoshita I, Dosaka-Akita H, et al. Tumor-infiltrating DCs suppress nucleic acid-mediated innate immune responses through interactions between the receptor TIM-3 and the alarmin HMGB1. Nat Immunol 2012; 13:832-842.
27. Clayton KL, Haaland MS, Douglas-Vail MB, Mujib S, Chew GM, Ndhlovu LC, et al. T cell Ig and mucin domain-containing protein 3 is recruited to the immune synapse, disrupts stable synapse formation, and associates with receptor phosphatases. . J Immunol 2014; 192:782-791.
28. Sada-Ovalle I, Ocaña-Guzman R, Pérez-Patrigeón S, Chávez-Galán L, Sierra-Madero J, Torre-Bouscoulet L, et al. Tim-3 blocking rescue macrophage and T cell function against Mycobacterium tuberculosis infection in HIV+ patients. J Int AIDS Soc 2015; 19:20078.
29. Sakuishi K, Ngiow SF, Sullivan JM, Teng MW, Kuchroo VK, Smyth MJ, et al. TIM3FOXP3 regulatory T cells are tissue-specific promoters of T-cell dysfunction in cancer. Oncoimmunology 2013; 2:e23849.
30. Baghdadi M, Takeuchi S, Wada H, Seino K.Blocking monoclonal antibodies of TIM proteins as orchestrators of anti-tumor immune response.mAbs 2014; 6:1124-1132.
31. Chiba S, Baghdadi M, Akiba H, Yoshiyama H, Kinoshita I, Dosaka-Akita H, et al. Tumor-infiltrating DCs suppress nucleic acid-mediated innate immune responses through interactions between the receptor TIM-3 and the alarmin HMGB1. Nat Immunol 2012; 13:832-842.
32. Ngiow SF, von Scheidt B,Akiba H, Yagita H, Teng MWL, Smyth MJ. Anti-TIM3 antibody promotes T cell IFN-g-mediated antitumor immunity and sup- presses established tumors. Cancer Res 2011; 71:3540- 3551.
33. Leone RD, Lo YC, Powell JD. A2aR antagonists: Next generation checkpoint blockade for cancer immunotherapy. Comput Struct Biotechnol J 2015; 13:265-272.
34. Chames P, Van Regenmortel M, Weiss E, Baty D.Therapeutic antibodies: successes, limitations and hopes for the future. Br J Pharmacol 2009; 157:220-233.
35. Omidfar K, Rasaee MJ, Modjtahedi H, Forouzandeh M, Taghikhani M, Bakhtiari A, et al. Production and characterization of a new antibody specific for the mutant EGF receptor,EGFRvIII, in Camelus Bacterianus. Tumor Biol 2004; 25:179-187.
36. Behdani M, Zeinali S, Khanahmad H, Karimipour M, Asadzadeh N, Azadmanesh K,et al. Generation and characterization of a functional Nanobody against the vascular endothelial growth factor receptor-2; angiogenesis cell receptor. Mol Immunol 2012; 50:35-41.
37. Ahmadvand D, Rasaee MJ,Rahbarizadeh F, Kontermann RE, Sheikholislami F. Cell selection and characterization of a novel human endothelial cell specific nanobody. Mol Immunol 2009; 46:1814-1823.
38. Abul Abbas AHL, Shiv Pillai. Cellular and Molecular Immunology. 2015; 8th edition.
39. Roovers RC, Laeremans T, Huang L, De Taeye S, Verkleij AJ, Revets H, et al. Efficient inhibition of EGFR signaling and of tumour growth by antagonistic anti-EFGR Nanobodies. Cancer Immunol Immunother 2007; 56:303-317. | ||
|
آمار تعداد مشاهده مقاله: 1,193 تعداد دریافت فایل اصل مقاله: 1,142 |
||