pSTAT3: a target biomarker to study the pharmacology of the anti-IL-21R antibody ATR-107 in human whole blood

Journal of Translational Medicine

Jaime L Masferrer²  John B Cheng³  David A Wunderlich³  Tsung H Lin¹  Susan Pleasic-Williams³  Ming Zhu² 

[1] Immunoscience, Pfizer Worldwide R&D, Cambridge, MA, USA;Precision Medicine, Pfizer BioTx Clinical R&D, 200 Cambridge Park Drive, Cambridge, MA 02140, USA;Regulated Bioanalysis and Biotherapeutics Clinical Research, Pfizer Worldwide R&D, Groton, CT, USA

关键词: IL-21R;    ATR-107;    pSTAT3;    IL-21;    Mechanism of action;    Target engagement;    Biomarker;   

Others  :  827952
DOI  :  10.1186/1479-5876-11-65

 received in 2012-12-18, accepted in 2013-02-19,  发布年份 2013

【 摘 要 】

Background

IL-21 has been shown to play an important role in autoimmune diseases. ATR-107 is an antibody which directly targets the IL-21 receptor (IL-21R). To aid the clinical development of ATR-107, there is a need for understanding the mechanism of action (MOA) of this antibody when assessing target engagement in human subjects.

Methods

To determine ATR-107 biological activity and potency in human blood, its inhibitory function against IL-21 induced STAT3 phosphorylation in human peripheral T and B cells was measured.

Results

The data show that IL-21 induces STAT3 phosphorylation in a concentration-dependent manner, consistent with its migration to the nuclear. Using a flow cytometry based functional whole blood assay, ATR-107 is demonstrated to be a potent IL-21 pathway inhibitor. It competes with IL-21 for receptor binding in a competitive manner, but once it binds to the receptor it behaves like a non-competitive inhibitor, most probably due to the long observed k_off. The concentration-dependent inhibition observed with ATR-107 correlates inversely with the levels of receptor occupancy, both in ex vivo whole blood assays and directly in human blood when ATR-107 was given to healthy volunteers.

Conclusions

IL-21 induced phosphorylation of STAT3 in T and B cells can be used as a biomarker to evaluate the target engagement of ATR-107 in human whole blood. The antibody behaves like a potent non-competitive inhibitor blocking IL-21 induced STAT3 phosphorylation for a long period of time. These results may help with the translation of preclinical information and dose selection towards ATR-107 clinical efficacy.

【 授权许可】

   
2013 Zhu et al.; licensee BioMed Central Ltd.

【 图 表 】

Figure 1.

Figure 2.

Figure 3.

Figure 4.

【 参考文献 】

• [1]Leonard WJ, Spolski R: Interleukin-21: a modulator of lymphoid proliferation, apoptosis and differentiation. Nat Rev Immunol 2005, 5:688-698.
• [2]Korn T, Bettelli E, Gao W, Awasthi A, Jager A, Strom TB, Oukka M, Kuchroo VK: IL-21 initiates an alternative pathway to induce proinflammatory T(H)17 cells. Nature 2007, 448:484-487.
• [3]Spolski R, Leonard WJ: Interleukin-21: basic biology and implications for cancer and autoimmunity. Annu Rev Immunol 2008, 26:57-79.
• [4]Coquet JM, Kyparissoudis K, Pellicci DG, Besra G, Berzins SP, Smyth MJ, Godfrey DI: IL-21 is produced by NKT cells and modulates NKT cell activation and cytokine production. J Immunol 2007, 178:2827-2834.
• [5]Ettinger R, Kuchen S, Lipsky PE: The role of IL-21 in regulating B-cell function in health and disease. Immunol Rev 2008, 223:60-86.
• [6]Asao H, Okuyama C, Kumaki S, Ishii N, Tsuchiya S, Foster D, Sugamura K: Cutting edge: the common gamma-chain is an indispensable subunit of the IL-21 receptor complex. J Immunol 2001, 167:1-5.
• [7]Habib T, Senadheera S, Weinberg K, Kaushansky K: The common gamma chain (gamma c) is a required signaling component of the IL-21 receptor and supports IL-21-induced cell proliferation via JAK3. Biochemistry 2002, 41:8725-8731.
• [8]Young DA, Hegen M, Ma HL, Whitters MJ, Albert LM, Lowe L, Senices M, Wu PW, Sibley B, Leathurby Y: Blockade of the interleukin-21/interleukin-21 receptor pathway ameliorates disease in animal models of rheumatoid arthritis. Arthritis Rheum 2007, 56:1152-1163.
• [9]Herber D, Brown TP, Liang S, Young DA, Collins M, Dunussi-Joannopoulos K: IL-21 has a pathogenic role in a lupus-prone mouse model and its blockade with IL-21R.Fc reduces disease progression. J Immunol 2007, 178:3822-3830.
• [10]McGuire HM, Walters S, Vogelzang A, Lee CM, Webster KE, Sprent J, Christ D, Grey S, King C: Interleukin-21 is critically required in autoimmune and allogeneic responses to islet tissue in murine models. Diabetes 2011, 60(3):867-875.
• [11]Wei L, Laurence A, Elias KM, O'Shea JJ: IL-21 is produced by Th17 cells and drives IL-17 production in a STAT3-dependent manner. J Biol Chem 2007, 282:34605-34610.
• [12]Avery DT, Deenick EK, Ma CS, Suryani S, Simpson N, Chew GY, Chan TD, Palendira U, Bustamante J, Boisson-Dupuis S: B cell-intrinsic signaling through IL-21 receptor and STAT3 is required for establishing long-lived antibody responses in humans. J Exp Med 2010, 207:155-171.
• [13]Goodman WA, Young AB, McCormick TS, Cooper KD, Levine AD: Stat3 phosphorylation mediates resistance of primary human T cells to regulatory T cell suppression. J Immunol 2011, 186:3336-3345.
• [14]Vugmeyster Y, Guay H, Szklut P, Qian MD, Jin M, Widom A, Spaulding V, Bennett F, Lowe L, Andreyeva T: In vitro potency, pharmacokinetic profiles, and pharmacological activity of optimized anti-IL-21R antibodies in a mouse model of lupus. MAbs 2010, 2:335-346.
• [15]Vugmeyster Y, Allen S, Szklut P, Bree A, Ryan M, Ma M, Spaulding V, Young D, Guay H, Bloom L: Correlation of pharmacodynamic activity, pharmacokinetics, and anti-product antibody responses to anti-IL-21R antibody therapeutics following IV administration to cynomolgus monkeys. J Transl Med 2010, 8:41. BioMed Central Full Text
• [16]Arai M, Jain S, Weaver AA, Hill AA, Guo Y, Bree AG, Smith MF Jr, Allen SW, LaVallie ER, Young D: Development and application of a biomarker assay for determining the pharmacodynamic activity of an antagonist candidate biotherapeutic antibody to IL21R in whole blood. J Transl Med 2010, 8:51. BioMed Central Full Text
• [17]Lin TH, Hegen M, Quadros E, Nickerson-Nutter CL, Appell KC, Cole AG, Shao Y, Tam S, Ohlmeyer M, Wang B: Selective functional inhibition of JAK-3 is sufficient for efficacy in collagen-induced arthritis in mice. Arthritis Rheum 2010, 62:2283-2293.
• [18]George TC, Fanning SL, Fitzgerald-Bocarsly P, Medeiros RB, Highfill S, Shimizu Y, Hall BE, Frost K, Basiji D, Ortyn WE: Quantitative measurement of nuclear translocation events using similarity analysis of multispectral cellular images obtained in flow. J Immunol Methods 2006, 311:117-129.
• [19]Pleasic-Williams S, Zhu M, Wunderlich D, Masferrer J, Hua FJC: Qualification of a human blood pSTAT3 assay for analysis of clinical samples: An involvement of its nuclear nuclear translocation in CD4+ cells. Clin Cytometry 2012, 82B(5):338.
• [20]Krutzik PO, Irish JM, Nolan GP, Perez OD: Analysis of protein phosphorylation and cellular signaling events by flow cytometry: techniques and clinical applications. Clin Immunol 2004, 110:206-221.
• [21]Dolff S, Abdulahad WH, Westra J, Doornbos-van Der Meer B, Limburg PC, Kallenberg CG, Bijl M: Increase in IL-21 producing T-cells in patients with systemic lupus erythematosus. Arthritis Res Ther 2011, 13:R157. BioMed Central Full Text
• [22]Terrier B, Costedoat-Chalumeau N, Garrido M, Geri G, Rosenzwajg M, Musset L, Klatzmann D, Saadoun D, Cacoub P: Interleukin 21 correlates with T cell and B cell subset alterations in systemic lupus erythematosus. J Rheumatol 2012, 39:1819-1828.
• [23]Mitoma H, Horiuchi T, Kimoto Y, Tsukamoto H, Uchino A, Tamimoto Y, Miyagi Y, Harada M: Decreased expression of interleukin-21 receptor on peripheral B lymphocytes in systemic lupus erythematosus. Int J Mol Med 2005, 16:609-615.
• [24]Vinh N, Horea R, Rus V: Increased IL-21R Expression and Signaling in B Cells From Patients with Systemic Lupus Erythematosus. 2011.