Division of Cancer Sciences, University of Manchester, Manchester M13 9PL, UK.
Computational Biology Support, CRUK Manchester Institute, Alderley Park SK10 4TG, UK.
Int J Mol Sci. 2023 May 18;24(10):8956. doi: 10.3390/ijms24108956.
Hypoxia and a suppressive tumour microenvironment (TME) are both independent negative prognostic factors for muscle-invasive bladder cancer (MIBC) that contribute to treatment resistance. Hypoxia has been shown to induce an immune suppressive TME by recruiting myeloid cells that inhibit anti-tumour T cell responses. Recent transcriptomic analyses show hypoxia increases suppressive and anti-tumour immune signalling and infiltrates in bladder cancer. This study sought to investigate the relationship between hypoxia-inducible factor (HIF)-1 and -2, hypoxia, and immune signalling and infiltrates in MIBC. ChIP-seq was performed to identify HIF1α, HIF2α, and HIF1β binding in the genome of the MIBC cell line T24 cultured in 1% and 0.1% oxygen for 24 h. Microarray data from four MIBC cell lines (T24, J82, UMUC3, and HT1376) cultured under 1%, 0.2%, and 0.1% oxygen for 24 h were used. Differences in the immune contexture between high- and low-hypoxia tumours were investigated using in silico analyses of two bladder cancer cohorts (BCON and TCGA) filtered to only include MIBC cases. GO and GSEA were used with the R packages "limma" and "fgsea". Immune deconvolution was performed using ImSig and TIMER algorithms. RStudio was used for all analyses. Under hypoxia, HIF1α and HIF2α bound to ~11.5-13.5% and ~4.5-7.5% of immune-related genes, respectively (1-0.1% O). HIF1α and HIF2α both bound to genes associated with T cell activation and differentiation signalling pathways. HIF1α and HIF2α had distinct roles in immune-related signalling. HIF1 was associated with interferon production specifically, whilst HIF2 was associated with generic cytokine signalling as well as humoral and toll-like receptor immune responses. Neutrophil and myeloid cell signalling was enriched under hypoxia, alongside hallmark pathways associated with Tregs and macrophages. High-hypoxia MIBC tumours had increased expression of both suppressive and anti-tumour immune gene signatures and were associated with increased immune infiltrates. Overall, hypoxia is associated with increased inflammation for both suppressive and anti-tumour-related immune signalling and immune infiltrates, as seen in vitro and in situ using MIBC patient tumours.
缺氧和抑制性肿瘤微环境（TME）都是浸润性膀胱癌（MIBC）的独立负预后因素，导致治疗耐药。缺氧已被证明通过招募抑制抗肿瘤 T 细胞反应的髓样细胞来诱导免疫抑制性 TME。最近的转录组分析显示，缺氧增加了膀胱癌中的抑制性和抗肿瘤免疫信号和浸润。本研究旨在探讨 MIBC 中缺氧诱导因子（HIF）-1 和 -2、缺氧与免疫信号和浸润之间的关系。ChIP-seq 用于鉴定 MIBC 细胞系 T24 在 1%和 0.1%氧气中培养 24 小时时 HIF1α、HIF2α 和 HIF1β 在基因组中的结合。使用在 1%、0.2%和 0.1%氧气中培养 24 小时的四个 MIBC 细胞系（T24、J82、UMUC3 和 HT1376）的微阵列数据。使用两个膀胱癌队列（BCON 和 TCGA）的计算分析研究高氧和低氧肿瘤之间免疫结构的差异，该队列经过过滤仅包括 MIBC 病例。使用 R 包“limma”和“fgsea”进行 GO 和 GSEA。使用 ImSig 和 TIMER 算法进行免疫去卷积。所有分析均使用 RStudio 进行。在缺氧条件下，HIF1α 和 HIF2α 分别结合约 11.5-13.5%和 4.5-7.5%的免疫相关基因（1-0.1%O）。HIF1α 和 HIF2α 都结合到与 T 细胞激活和分化信号通路相关的基因上。HIF1α 和 HIF2α 在免疫相关信号中具有不同的作用。HIF1 与干扰素的产生特异性相关，而 HIF2 与通用细胞因子信号以及体液和 Toll 样受体免疫反应相关。缺氧时，中性粒细胞和髓样细胞信号被富集，同时与 Treg 和巨噬细胞相关的标志性途径也被富集。高氧 MIBC 肿瘤中抑制性和抗肿瘤免疫基因特征的表达增加，与免疫浸润增加相关。总的来说，缺氧与抑制性和抗肿瘤相关免疫信号和免疫浸润的炎症增加有关，这在体外和 MIBC 患者肿瘤的原位观察中都可以看到。
