Dipartimento di Scienze Cliniche e Molecolari, Università Politecnica delle Marche, Ancona, Italy; Clinica di Ematologia, Azienda Ospedaliero Universitaria Ospedali Riuniti, Ancona, Italy.
Dipartimento di Scienze Cliniche e Molecolari, Università Politecnica delle Marche, Ancona, Italy; Servizio di Immunologia Clinica, Azienda Ospedaliero Universitaria Ospedali Riuniti, Ancona, Italy.
Biol Blood Marrow Transplant. 2018 Feb;24(2):267-275. doi: 10.1016/j.bbmt.2017.10.039. Epub 2017 Nov 8.
Pathogenesis of chronic graft-versus-host disease (cGVHD) is incompletely defined, involving donor-derived CD4 and CD8-positive T lymphocytes as well as B cells. Standard treatment is lacking for steroid-dependent/refractory cases; therefore, the potential usefulness of tyrosine kinase inhibitors (TKIs) has been suggested, based on their potent antifibrotic effect. However, TKIs seem to have pleiotropic activity. We sought to evaluate the in vitro and in vivo impact of different TKIs on lymphocyte phenotype and function. Peripheral blood mononuclear cells (PBMCs) from healthy donors were cultured in the presence of increasing concentrations of nilotinib, imatinib, dasatinib, and ponatinib; in parallel, 44 PBMC samples from 15 patients with steroid-dependent/refractory cGVHD treated with nilotinib in the setting of a phase I/II trial were analyzed at baseline, after 90, and after 180 days of therapy. Flow cytometry was performed after labeling lymphocytes with a panel of monoclonal antibodies (CD3, CD4, CD16, CD56, CD25, CD19, CD45RA, FoxP3, CD127, and 7-amino actinomycin D). Cytokine production was assessed in supernatants of purified CD3 T cells and in plasma samples from nilotinib-treated patients. Main T lymphocyte subpopulations were not significantly affected by therapeutic concentrations of TKIs in vitro, whereas proinflammatory cytokine (in particular, IL-2, IFN-γ, tumor necrosis factor-α, and IL-10) and IL-17 production showed a sharp decline. Frequency of T regulatory, B, and natural killer (NK) cells decreased progressively in presence of therapeutic concentrations of all TKIs tested in vitro, except for nilotinib, which showed little effect on these subsets. Of note, naive T regulatory cell (Treg) subset accumulated after exposure to TKIs. Results obtained in vivo on nilotinib-treated patients were largely comparable, both on lymphocyte subset kinetics and on cytokine production by CD3-positive cells. This study underlines the anti-inflammatory and immunomodulatory effects of TKIs and supports their potential usefulness as treatment for patients with steroid-dependent/refractory cGVHD. In addition, both in vitro and in vivo data point out that compared with other TKIs, nilotinib could better preserve the integrity of some important regulatory subsets, such as Treg and NK cells.
慢性移植物抗宿主病（cGVHD）的发病机制尚未完全明确，涉及供体来源的 CD4 和 CD8 阳性 T 淋巴细胞以及 B 细胞。对于依赖于激素/难治性病例，目前缺乏标准治疗方法；因此，基于其强大的抗纤维化作用，已提出使用酪氨酸激酶抑制剂（TKI）的可能性。然而，TKI 似乎具有多效性。我们试图评估不同 TKI 对淋巴细胞表型和功能的体外和体内影响。从健康供体的外周血单核细胞（PBMC）中培养在增加浓度的 nilotinib、imatinib、dasatinib 和 ponatinib；同时，分析了来自 15 例接受 nilotinib 治疗的依赖激素/难治性 cGVHD 患者的 44 例 PBMC 样本，在 I/II 期试验中作为基线、90 天后和 180 天后的治疗。在使用一组单克隆抗体（CD3、CD4、CD16、CD56、CD25、CD19、CD45RA、FoxP3、CD127 和 7-氨基放线菌素 D）标记淋巴细胞后进行流式细胞术。在来自 nilotinib 治疗患者的纯化 CD3 T 细胞上清液和血浆样本中评估细胞因子产生。主要 T 淋巴细胞亚群在体外治疗浓度的 TKI 作用下没有明显受到影响，而促炎细胞因子（特别是 IL-2、IFN-γ、肿瘤坏死因子-α和 IL-10）和 IL-17 的产生则急剧下降。在体外测试的所有 TKI 存在的治疗浓度下，T 调节细胞、B 和自然杀伤（NK）细胞的频率逐渐降低，除了 nilotinib，它对这些亚群几乎没有影响。值得注意的是，在接触 TKI 后，幼稚 T 调节细胞（Treg）亚群积累。在接受 nilotinib 治疗的患者体内获得的结果在很大程度上是可比的，无论是在淋巴细胞亚群动力学还是在 CD3 阳性细胞产生的细胞因子方面。这项研究强调了 TKI 的抗炎和免疫调节作用，并支持它们作为治疗依赖激素/难治性 cGVHD 患者的潜在用途。此外，体外和体内数据均表明，与其他 TKI 相比，nilotinib 可以更好地保留 Treg 和 NK 细胞等一些重要调节亚群的完整性。
