TREM2 inhibition triggers anti-tumor cell activity of myeloid cells in glioblastoma [scRNA]. TREM2 inhibition triggers anti-tumor cell activity of myeloid cells in glioblastoma [scRNA]

The prevailing view is that myeloid cells in the tumor microenvironment (TME) are immunosuppressive and promote glioblastoma (GBM) progression. However, myeloid cells have the functional plasticity to restrict or support tumor cell growth. TREM2 plays important roles in brain microglial function in neurodegenerative diseases, but the role of TREM2 in the GBM TME has not been examined. We found TREM2 is highly expressed in myeloid subsets, including macrophages and microglia in human and mouse GBM tumors and that high TREM2 expression correlates with poor prognosis in GBM patients. TREM2 loss of function in human macrophages and mouse myeloid cells increased tumoricidal capacity. TREM2 in myeloid cells restricts IFNγ-induced immunoactivation and proinflammatory polarization. In orthotopic mouse GBM models, Trem2-/- mice and mice with acute brain Trem2 reduction demonstrate survival benefit. Trem2 inhibition reprograms myeloid phenotypes and increases PD-1+CD8+ T cells in the TME. Trem2 deficiency enhances the effectiveness of anti-PD-1 treatment and may represent a therapeutic strategy for GBM patients. Overall design: Bone marrow-derived macrophages (BMDMs) were isolated and differentiated from trem2 transgenic mice, Trem2+/+ (n=2), Trem2-/- (n=2). Cells were treated with PBS (n=2), 10 ng/mL IFNγ (n=2), 100 ng/mL LPS (n=2), or dead SB28 tumor cells (n=2) for 6 hours. Total RNA from each sample/genotype was isolated and sequenced. Primary microglia were enriched from brain cortex of postnatal day 2 Trem2+/+ mouse pups (n=12). Primary microglia were treated with Trem2-lowering antisense oligonucleotide (ASO) or inactive ASO for 72 hours. Trem2 ASO (n=2), inactive (n=2). Cells were then treated with PBS (n=2),10 ng/mL IFNγ (n=2) or dead SB28 tumor cells (n=2) for 6 hours. Total RNA from each sample/treatment was isolated and sequenced. Trem2+/+ mice were intracerebroventricularly injected with 400 μg (10ul) Trem2 ASO (n=5) or inactive ASO (n=5). 7 days later, all mice were intracerebrally injected with SB28 cells. 30 days later, SB28 glioblastoma were dissected from ASO-treated mice brain and dissociated into single-cell suspensions for RNA-seq. Trem2 ASO (n=2), inactive ASO (n=2).

当前主流观点认为，肿瘤微环境（tumor microenvironment, TME）中的髓系细胞具有免疫抑制功能，并可促进胶质母细胞瘤（glioblastoma, GBM）的进展。然而，髓系细胞具备功能可塑性，既可以抑制也可以促进肿瘤细胞的生长。髓系细胞触发受体2（TREM2）在神经退行性疾病的脑内小胶质细胞功能调控中发挥重要作用，但目前尚未有研究探讨其在GBM TME中的功能。本研究发现，TREM2在人源及小鼠GBM肿瘤的髓系细胞亚群（包括巨噬细胞与小胶质细胞）中高表达，且TREM2高表达与GBM患者的不良预后显著相关。在人源巨噬细胞及小鼠髓系细胞中，TREM2功能缺失可提升其肿瘤杀伤活性。髓系细胞中的TREM2可抑制干扰素γ（IFNγ）诱导的免疫激活与促炎极化过程。在小鼠原位GBM模型中，Trem2基因敲除（Trem2-/-）小鼠以及急性脑内Trem2表达降低的小鼠均展现出生存获益。Trem2抑制可重编程髓系细胞表型，并增加TME中PD-1阳性CD8阳性T细胞的浸润。Trem2功能缺失可增强抗PD-1治疗的疗效，或可成为GBM患者的潜在治疗策略。研究设计概述：从Trem2转基因小鼠（Trem2+/+，n=2；Trem2-/-，n=2）中分离并诱导分化骨髓来源巨噬细胞（bone marrow-derived macrophages, BMDMs）。将细胞分别用磷酸盐缓冲液（PBS，n=2）、10 ng/mL 干扰素γ（IFNγ，n=2）、100 ng/mL 脂多糖（LPS，n=2）或灭活的SB28肿瘤细胞（n=2）处理6小时。提取每个样本/基因型的总RNA并进行测序。从出生后2天的Trem2+/+幼鼠大脑皮层中富集原代小胶质细胞（n=12）。将原代小胶质细胞分别用靶向Trem2的反义寡核苷酸（antisense oligonucleotide, ASO，n=2）或无效对照ASO（n=2）处理72小时。随后将细胞分别用PBS（n=2）、10 ng/mL IFNγ（n=2）或灭活的SB28肿瘤细胞（n=2）处理6小时，提取每个样本/处理组的总RNA并进行测序。向Trem2+/+小鼠脑室内注射400 μg（10 μL）靶向Trem2的ASO（n=5）或无效对照ASO（n=5）。7天后，所有小鼠均脑内接种SB28肿瘤细胞。30天后，从经ASO处理的小鼠脑中分离SB28胶质母细胞瘤组织，解离为单细胞悬液后进行RNA测序（RNA-seq），其中Trem2 ASO组n=2，无效对照ASO组n=2。