Shraga Segal Department of Microbiology, Immunology, and Genetics, Ben-Gurion University of the Negev, 8410501, Beer-Sheva, Israel.
Department of Health Policy and Management, School of Public Health, Faculty of Health Sciences, Ben-Gurion University of the Negev, 8410501, Beer-Sheva, Israel.
J Exp Clin Cancer Res. 2023 May 1;42(1):107. doi: 10.1186/s13046-023-02680-7.
Ovarian cancer (OC) is known for exhibiting low response rates to immune checkpoint inhibitors that activate T cells. However, immunotherapies that activate B cells have not yet been extensively explored and may be a potential target, as B cells that secrete immunoglobulins have been associated with better outcomes in OC. Although the secretion of immunoglobulins is often mediated by the microbiome, it is still unclear what role they play in limiting the progression of OC.
We conducted an in-vivo CRISPR screen of immunodeficient (NSG) and immune-intact wild type (WT) C57/BL6 mice to identify tumor-derived immune-escape mechanisms in a BRAC1- and TP53-deficient murine ID8 OC cell line (designated ITB1). To confirm gene expression and signaling pathway activation in ITB1 cells, we employed western blot, qPCR, immunofluorescent staining, and flow cytometry. Flow cytometry was also used to identify immune cell populations in the peritoneum of ITB1-bearing mice. To determine the presence of IgA-coated bacteria in the peritoneum of ITB1-bearing mice and the ascites of OC patients, we employed 16S sequencing. Testing for differences was done by using Deseq2 test and two-way ANOVA test. Sequence variants (ASVs) were produced in Qiime2 and analyzed by microeco and phyloseq R packages.
We identified tumor necrosis factor receptor-associated factor 3 (TRAF3) as a tumor-derived immune suppressive mediator in ITB1 cells. Knockout of TRAF3 (TRAF3KO) activated the type-I interferon pathway and increased MHC-I expression. TRAF3KO tumors exhibited a growth delay in WT mice vs. NSG mice, which was correlated with increased B cell infiltration and activation compared to ITB1 tumors. B cells were found to be involved in the progression of TRAF3KO tumors, and B-cell surface-bound and secreted IgA levels were significantly higher in the ascites of TRAF3KO tumors compared to ITB1. The presence of commensal microbiota was necessary for B-cell activation and for delaying the progression of TRAF3KO tumors in WT mice. Lastly, we observed unique profiles of IgA-coated bacteria in the ascites of OC-bearing mice or the ascites of OC patients.
TRAF3 is a tumor-derived immune-suppressive modulator that influences B-cell infiltration and activation, making it a potential target for enhancing anti-tumor B-cell responses in OC.
卵巢癌 (OC) 的特点是对激活 T 细胞的免疫检查点抑制剂反应率低。然而，激活 B 细胞的免疫疗法尚未得到广泛探索，可能是一个潜在的目标，因为分泌免疫球蛋白的 B 细胞与 OC 的更好结果相关。尽管免疫球蛋白的分泌通常由微生物组介导，但它们在限制 OC 进展中的作用仍不清楚。
我们对免疫缺陷型 (NSG) 和免疫完整型野生型 (WT) C57/BL6 小鼠进行了体内 CRISPR 筛选，以鉴定 BRAC1 和 TP53 缺陷型鼠 ID8 OC 细胞系 (命名为 ITB1) 中的肿瘤衍生免疫逃逸机制。为了确认 ITB1 细胞中的基因表达和信号通路激活，我们采用了 Western blot、qPCR、免疫荧光染色和流式细胞术。流式细胞术还用于鉴定 ITB1 荷瘤小鼠腹腔中的免疫细胞群。为了确定 ITB1 荷瘤小鼠腹腔和 OC 患者腹水内是否存在 IgA 包被细菌，我们采用了 16S 测序。使用 Deseq2 测试和双因素方差分析测试来检测差异。Qiime2 中生成序列变体 (ASVs)，并使用 microeco 和 phyloseq R 包进行分析。
我们发现肿瘤坏死因子受体相关因子 3 (TRAF3) 是 ITB1 细胞中肿瘤衍生的免疫抑制介质。TRAF3 敲除 (TRAF3KO) 激活了 I 型干扰素途径并增加了 MHC-I 表达。与 ITB1 肿瘤相比，TRAF3KO 肿瘤在 WT 小鼠中表现出生长延迟，这与与 ITB1 肿瘤相比，B 细胞浸润和激活增加有关。发现 B 细胞参与了 TRAF3KO 肿瘤的进展，并且与 ITB1 相比，TRAF3KO 肿瘤的腹水内 B 细胞表面结合和分泌的 IgA 水平显着升高。共生微生物群的存在对于 B 细胞的激活以及在 WT 小鼠中延迟 TRAF3KO 肿瘤的进展是必要的。最后，我们在 OC 荷瘤小鼠或 OC 患者的腹水中观察到独特的 IgA 包被细菌谱。
TRAF3 是一种肿瘤衍生的免疫抑制调节剂，可影响 B 细胞浸润和激活，使其成为增强 OC 中抗肿瘤 B 细胞反应的潜在靶点。
