Immune subtyping of extranodal NK/T-cell lymphoma: a new biomarker and an immune shift during disease progression.
结外 NK/T 细胞淋巴瘤的免疫分型: 一种新的生物标志物和疾病进展过程中的免疫转移。
- 作者列表："Cho J","Kim SJ","Park WY","Kim J","Woo J","Kim G","Yoon SE","Ko YH","Kim WS
:Extranodal NK/T-cell lymphoma is an aggressive lymphoma that is strongly associated with Epstein-Barr virus infection. Although some extranodal NK/T-cell lymphoma patients have shown responses to immune checkpoint blockade, biomarkers for predicting extranodal NK/T-cell lymphoma patient response to immunotherapy have not yet been defined. To understand the tumor immune microenvironment, we analyzed the expression of 579 immune-related genes and characterized the immune cells using immunohistochemistries and in situ hybridization for EBER. Based on comprehensive analyses, we developed an immune subtyping model that classifies extranodal NK/T-cell lymphoma patients into four tumor immune microenvironment subgroups using three immunohistochemical markers (FoxP3, PD-L1, and CD68). The four tumor immune microenvironment subgroups were named immune tolerance, immune evasion-A, immune evasion-B, and immune silenced. The immune tolerance group was characterized by high-Treg counts and was frequently observed in early stage, and nasal extranodal NK/T-cell lymphoma. The immune evasion group showed high cytotoxic T-cell counts and high PD-L1 expression but low Treg counts. In the immune-silenced group, almost all immune responses were exhausted, most patients were at an advanced stage, and had the poorest disease prognosis among the tumor immune microenvironment subgroups. In some patients (n = 3), a shift in the tumor immune microenvironment subgroup classification was observed in sequential biopsies. The response rate to pembrolizumab, an anti-PD-1 antibody, was 100% (1/1) in the immune tolerance group, 60% (3/5) in the immune evasion group, and 0% (0/5) in the immune-silenced group. We classified extranodal NK/T-cell lymphoma into four tumor immune microenvironment subgroups using a new classification system. In conclusion, we propose that the tumor immune microenvironment of extranodal NK/T-cell lymphoma may change during disease progression and may serve as a useful biomarker for immunotherapy.
: 结外 NK/T 细胞淋巴瘤是一种侵袭性淋巴瘤，与 eb病毒感染密切相关。尽管一些结外 NK/T 细胞淋巴瘤患者已经显示出对免疫检查点阻断的反应，但预测结外 NK/T 细胞淋巴瘤患者对免疫治疗反应的生物标志物尚未确定。为了了解肿瘤免疫微环境，我们分析了 579 个免疫相关基因的表达，并使用免疫组化和 EBER 原位杂交对免疫细胞进行了表征。基于综合分析，我们开发了一个免疫分型模型，使用三种免疫组化标记 (FoxP3 、 PD-L1 和 CD68) 将结外 NK/T 细胞淋巴瘤患者分为四个肿瘤免疫微环境亚组。四个肿瘤免疫微环境亚组分别命名为免疫耐受、免疫逃避-A 、免疫逃避-B 和免疫沉默。免疫耐受组以 Treg 计数高为特征，早期常观察到，鼻部结外 NK/T 细胞淋巴瘤。免疫逃避组表现出高细胞毒性 T 细胞计数和高 PD-L1 表达，但 Treg 计数低。在免疫沉默组中，几乎所有的免疫反应都耗尽，大多数患者处于晚期，在肿瘤免疫微环境亚组中疾病预后最差。在一些患者 (n = 3) 中，在序贯活检中观察到肿瘤免疫微环境亚组分类的转变。免疫耐受组对 anti-PD-1 抗体 pembrolizumab 的应答率为 100% (1/1)，免疫逃避组为 60% (3/5)，0% (0/5) 在免疫沉默组。我们使用新的分类系统将结外 NK/T 细胞淋巴瘤分为 4 个肿瘤免疫微环境亚组。总之，我们提出结外 NK/T 细胞淋巴瘤的肿瘤免疫微环境可能在疾病进展过程中发生变化，并可能作为免疫治疗的有用生物标志物。
METHODS::Macrophages (MΦ) play a critical role in tumor growth, immunosuppression and inhibition of adaptive immune responses in cancer. Hence, targeting signaling pathways in MΦs that promote tumor immunosuppression will provide therapeutic benefit. PI3Kγ has been recently established by our group and others as a novel immuno-oncology target. Herein, we report that a macrophage Syk-PI3K axis drives polarization of immunosuppressive MΦs which establish an immunosuppressive tumor microenvironment in in vivo syngeneic tumor models. Genetic or pharmacological inhibition of Syk and/or PI3Kγ in MΦs promotes a pro-inflammatory MΦphenotype, restores CD8+ T cell activity, destabilizes HIF under hypoxia, and stimulates an antitumor immune response. Assay for Transposase-accessible Chromatin using Sequencing (ATAC-seq) analyses on the bone marrow derived macrophages (BMDMs) show that inhibition of Syk kinase promotes activation and binding of NF-κB motif in SykMC-KO BMDMs, thus stimulating immunostimulatory transcriptional programming in MΦs to suppress tumor growth. Finally, we have developed in silico the "first in class" dual Syk/PI3K inhibitor, SRX3207, for the combinatorial inhibition of Syk and PI3K in one small molecule. This chemotype demonstrates efficacy in multiple tumor models and represents a novel combinatorial approach to activate antitumor immunity.
METHODS::Dysfunction of invariant natural killer T (iNKT) cells in tumor microenvironment hinders their anti-tumor efficacy, and the underlying mechanisms remain unclear. Here we report that iNKT cells increase lipid biosynthesis after activation, and that is promoted by PPARγ and PLZF synergically through enhancing transcription of Srebf1. Among those lipids, cholesterol is required for the optimal IFN-γ production from iNKT cells. Lactic acid in tumor microenvironment reduces expression of PPARγ in intratumoral iNKT cells and consequently diminishes their cholesterol synthesis and IFN-γ production. Importantly, PPARγ agonist pioglitazone, a thiazolidinedione drug for type 2 diabetes, successfully restores IFN-γ production in tumor-infiltrating iNKT cells from both human patients and mouse models. Combination of pioglitazone and alpha-galactosylceramide treatments significantly enhances iNKT cell-mediated anti-tumor immune responses and prolongs survival of tumor-bearing mice. Our studies provide a strategy to augment the anti-tumor efficacy of iNKT cell-based immunotherapies via promoting their lipid biosynthesis.
METHODS::Near-infrared photoimmunotherapy (NIR-PIT) is a newly developed and selective cancer treatment that induces necrotic and immunogenic cell death and utilizes a monoclonal antibody conjugated to a photo-absorber dye, IR700DX, activated by NIR light. Although CD44 is surface cancer marker associated with drug resistance, anti-CD44-IR700 NIR-PIT results in inhibited cell growth and prolonged survival in multiple tumor types. Meanwhile, anti-CD25-IR700-targeted NIR-PIT has been reported to achieve selective and local depletion of FOXP3+CD25+CD4+ regulatory T cells (Tregs), which are primary immunosuppressive cells in the tumor microenvironment (TME), resulting in activation of local antitumor immunity. Combined NIR-PIT with CD44- and CD25-targeted agents has the potential to directly eliminate tumor cells and also amplify the immune response by removing FOXP3+CD25+CD4+ Tregs from the TME. We investigated the difference in therapeutic effects of CD44-targeted NIR-PIT alone, CD25-targeted NIR-PIT alone, and the combination of CD44- and CD25-targeted NIR-PIT in several syngeneic tumor models, including MC38-luc, LL/2, and MOC1. The combined NIR-PIT showed significant tumor growth inhibition and prolonged survival compared with CD44-targeted NIR-PIT alone in all tumor models and showed prolonged survival compared with CD25-targeted NIR-PIT alone in MC38-luc and LL/2 tumors. Combined CD44/CD25 NIR-PIT also resulted in some complete remissions, whereas this was not achieved with either type of NIR-PIT alone. Therefore, combined NIR-PIT simultaneously targeting cancer antigens and immunosuppressive cells in the TME may be more effective than either type of NIR-PIT alone and may have potential to induce prolonged immune responses in treated tumors.