HDAC6 selective inhibition of melanoma patient T-cells augments anti-tumor characteristics
HDAC6 selective inhibition of melanoma patient T-cells augments anti-tumor characteristics
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DOI：
10.1186/s40425-019-0517-0
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发表时间：
2019-02-06
影响因子：
10.9
通讯作者：
Woods, David M.
中科院分区：
文献类型：
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作者：
Laino, Andressa S.;Betts, B. C.;Woods, David M.
靶向抗肿瘤T细胞应答的疗法已被证明在治疗各种恶性肿瘤中是成功的。然而，由于大多数患者仍然没有反应，因此需要增加免疫功效的方法。在这里，我们研究了组蛋白去乙酰化酶6（HDAC 6）选择性抑制剂的能力，以减少免疫抑制和增强免疫功能的黑色素瘤患者T细胞在离体culture. T细胞从外周血或转移性黑色素瘤患者的肿瘤活检和培养的存在下，泛，类特异性或类选择性组蛋白去乙酰化酶（HDAC）抑制剂。通过Luminex和细胞内流式细胞术染色评价细胞因子产生的变化。通过流式细胞术评估表面标志物、转录因子、蛋白磷酸化和细胞活力的表达。通过ATAC-seq测定染色质结构的变化。低剂量的泛HDAC抑制剂损害T细胞活力，但特异性或选择性HDAC抑制剂不损害T细胞活力。HDAC 6选择性抑制剂ACY-1215（ricolinostat）和ACY-241（citarinostat）降低了Th 2细胞因子（即IL-4、IL-5、IL-6、IL-10和IL-13）的产生。在存在这些抑制剂的情况下，来自黑素瘤患者的外周血T细胞的扩增导致Th 2转录因子GATA 3的下调，Th 1转录因子T-BET的上调，中枢记忆表型T细胞的积累，在混合淋巴细胞反应中，减少的细胞因子（CD 45 RA-CD 45 RO + CD 62 L + CCR 7+）、减少的耗竭相关表型（即TIM 3 + LAG 3 + PD 1+和EOMES+ PD 1+）和增强的杀伤。在暴露于ACY-1215或ACY-241后，调节性T细胞（TCFs）的频率、FOXP 3表达和抑制功能降低。在黑色素瘤肿瘤浸润淋巴细胞（TIL）中观察到表达CD 107 a + IFN γ +和中枢记忆标志物的T细胞频率较高，在药物去除和进一步扩增后仍持续存在。ACY-1215处理后，在与T细胞效应子功能和记忆表型相关的区域中观察到染色质可及性增加，而在编码mTOR下游分子AKT、SGK 1和S6 K的区域中发现浓缩的染色质。在ACY-1215和ACY-241处理的T细胞中观察到这些蛋白的磷酸化降低。AKT和SGK 1特异性抑制分别重现了中枢记忆频率的增加和IL-4产生的减少，与HDAC 6选择性抑制剂观察到的效果相似。HDAC 6选择性抑制剂增强了黑色素瘤患者的T细胞免疫特性，为评估其潜在临床疗效的翻译研究提供了理论基础。
Therapies targeting anti-tumor T-cell responses have proven successful in the treatment of a variety of malignancies. However, as most patients still fail to respond, approaches to augment immunotherapeutic efficacy are needed. Here, we investigated the ability of histone deacetylase 6 (HDAC6)-selective inhibitors to decrease immunosuppression and enhance immune function of melanoma patient T-cells in ex vivo cultures.T-cells were harvested from peripheral blood or tumor biopsies of metastatic melanoma patients and cultured in the presence of pan-, class-specific or class-selective histone deacetylase (HDAC) inhibitors. Changes in cytokine production were evaluated by Luminex and intracellular flow cytometry staining. Expression of surface markers, transcription factors, protein phosphorylation, and cell viability were assessed by flow cytometry. Changes in chromatin structure were determined by ATAC-seq.T-cell viability was impaired with low doses of pan-HDAC inhibitors but not with specific or selective HDAC inhibitors. The HDAC6-selective inhibitors ACY-1215 (ricolinostat) and ACY-241 (citarinostat) decreased Th2 cytokine production (i.e. IL-4, IL-5, IL-6, IL-10 and IL-13). Expansion of peripheral blood T-cells from melanoma patients in the presence of these inhibitors resulted in downregulation of the Th2 transcription factor GATA3, upregulation of the Th1 transcription factor T-BET, accumulation of central memory phenotype T-cells (CD45RA-CD45RO + CD62L + CCR7+), reduced exhaustion-associated phenotypes (i.e. TIM3 + LAG3 + PD1+ and EOMES+PD1+), and enhanced killing in mixed lymphocyte reactions. The frequency, FOXP3 expression, and suppressive function of T regulatory cells (Tregs) were decreased after exposure to ACY-1215 or ACY-241. Higher frequencies of T-cells expressing CD107a + IFN gamma+ and central memory markers were observed in melanoma tumor-infiltrating lymphocytes (TIL), which persisted after drug removal and further expansion. After ACY-1215 treatment, increased chromatin accessibility was observed in regions associated with T-cell effector function and memory phenotypes, while condensed chromatin was found in regions encoding the mTOR downstream molecules AKT, SGK1 and S6K. Decreased phosphorylation of these proteins was observed in ACY-1215 and ACY-241-treated T-cells. AKT- and SGK1-specific inhibition recapitulated the increase in central memory frequency and decrease in IL-4 production, respectively, similar to the observed effects of HDAC6-selective inhibition.HDAC6-selective inhibitors augmented melanoma patient T-cell immune properties, providing a rationale for translational investigation assessing their potential clinical efficacy.