Abstract：
BACKGROUND: Chronic immune activation plays a significant role in the pathogenesis and disease progression of human immunodeficiency virus (HIV), and the existing interventions to address this issue are limited. In a phase II clinical trial, (5R)-5-hydroxytriptolide (LLDT-8) demonstrated promising potential in enhancing CD4+ T cell recovery. However, the therapeutical effects of LLDT-8 remained to be systemic explored.
METHODS: To assess the treatment effects of LLDT-8, we conducted flow cytometry and RNA-seq analyses on eight Chinese rhesus monkeys infected with simian immunodeficiency virus (SIV). Additionally, we performed comprehensive transcriptomic analyses, including cross-sectional and longitudinal differentially expressed gene (DEG) analysis, gene set enrichment analysis (GSEA), weighted gene co-expression network analysis (WGCNA), and deconvolution analysis using peripheral blood mononuclear cell (PBMC) samples from 14-time points. These findings were further validated with RNA-seq analysis on patients who received LLDT-8 treatment, along with in vitro cellular experiments using human PBMCs.
RESULTS: Flow cytometry analysis revealed that LLDT-8 treatment significantly reduced the percentage of HLA-DR+CD38+CD8+ T cells in SIV-infected rhesus monkeys (P < 0.001). The cross-sectional and longitudinal analysis identified 2531 and 1809 DEGs, respectively. GSEA analysis indicated that LLDT-8 treatment led to significant downregulation of proliferation-related pathways, such as E2F targets, G2M checkpoint, and mitotic spindle pathways. WGCNA analysis identified two modules and 202 hub genes associated with CD8 activation levels. Deconvolution analysis showed a significant decrease in the proportion of CD8+ T cells and activated CD4+ T cells during LLDT-8 treatment. Gene ontology results demonstrated that the common DEGs between LLDT-8-treated patients and rhesus monkeys were primarily enriched in cell activation and cell cycle progression. Furthermore, in vitro cellular experiments validated the consistent impact of LLDT-8 in inhibiting proliferation, activation (HLA-DR and CD38 expression), exhaustion (PD-1 expression), and IFN-γ production in human CD4+ and CD8+ T cells.
CONCLUSIONS: LLDT-8 exhibited notable efficacy in alleviating immune activation in both an in vivo animal model and in vitro human cell experiments. These findings suggest that LLDT-8 may hold potential as a drug for managing systemic immune activation associated with SIV/HIV infection, warranting further prospective clinical exploration.
METHODS: To assess the treatment effects of LLDT-8, we conducted flow cytometry and RNA-seq analyses on eight Chinese rhesus monkeys infected with simian immunodeficiency virus (SIV). Additionally, we performed comprehensive transcriptomic analyses, including cross-sectional and longitudinal differentially expressed gene (DEG) analysis, gene set enrichment analysis (GSEA), weighted gene co-expression network analysis (WGCNA), and deconvolution analysis using peripheral blood mononuclear cell (PBMC) samples from 14-time points. These findings were further validated with RNA-seq analysis on patients who received LLDT-8 treatment, along with in vitro cellular experiments using human PBMCs.
RESULTS: Flow cytometry analysis revealed that LLDT-8 treatment significantly reduced the percentage of HLA-DR+CD38+CD8+ T cells in SIV-infected rhesus monkeys (P < 0.001). The cross-sectional and longitudinal analysis identified 2531 and 1809 DEGs, respectively. GSEA analysis indicated that LLDT-8 treatment led to significant downregulation of proliferation-related pathways, such as E2F targets, G2M checkpoint, and mitotic spindle pathways. WGCNA analysis identified two modules and 202 hub genes associated with CD8 activation levels. Deconvolution analysis showed a significant decrease in the proportion of CD8+ T cells and activated CD4+ T cells during LLDT-8 treatment. Gene ontology results demonstrated that the common DEGs between LLDT-8-treated patients and rhesus monkeys were primarily enriched in cell activation and cell cycle progression. Furthermore, in vitro cellular experiments validated the consistent impact of LLDT-8 in inhibiting proliferation, activation (HLA-DR and CD38 expression), exhaustion (PD-1 expression), and IFN-γ production in human CD4+ and CD8+ T cells.
CONCLUSIONS: LLDT-8 exhibited notable efficacy in alleviating immune activation in both an in vivo animal model and in vitro human cell experiments. These findings suggest that LLDT-8 may hold potential as a drug for managing systemic immune activation associated with SIV/HIV infection, warranting further prospective clinical exploration.
摘要：
背景：慢性免疫激活在人类免疫缺陷病毒（HIV）的发病机理和疾病进展中起着重要作用，和现有的干预措施，以解决这一问题是有限的。在一项II期临床试验中,（5R）-5-羟基雷公藤甲素（LLDT-8）在增强CD4T细胞恢复方面显示出有希望的潜力。然而,LLDT-8的治疗作用仍有待系统探索.
方法：为了评估LLDT-8的治疗效果，我们对8只感染猿猴免疫缺陷病毒（SIV）的中国猕猴进行了流式细胞术和RNA-seq分析。此外,我们进行了全面的转录组学分析，包括横截面和纵向差异表达基因(DEG)分析，基因集富集分析(GSEA)，加权基因共表达网络分析(WGCNA)，使用来自14个时间点的外周血单核细胞（PBMC）样品进行去卷积分析。这些发现通过对接受LLDT-8治疗的患者的RNA-seq分析得到了进一步验证。以及使用人PBMC的体外细胞实验。
结果：流式细胞术分析显示，LLDT-8治疗可显著降低SIV感染的恒河猴中HLA-DR+CD38+CD8+T细胞的百分比（P<0.001）。横截面和纵向分析确定了2531和1809DEG，分别。GSEA分析表明，LLDT-8治疗导致增殖相关途径的显著下调，例如E2F目标，G2M检查点,和有丝分裂纺锤体途径。WGCNA分析鉴定了与CD8活化水平相关的两个模块和202个hub基因。去卷积分析显示在LLDT-8处理期间CD8+T细胞和活化的CD4+T细胞的比例显著降低。基因本体论结果表明，LLDT-8治疗的患者和恒河猴之间的常见DEG主要富集在细胞活化和细胞周期进程中。此外，体外细胞实验验证了LLDT-8在抑制增殖方面的一致影响，激活（HLA-DR和CD38表达），耗尽（PD-1表达），和人CD4+和CD8+T细胞中IFN-γ的产生。
结论：LLDT-8在体内动物模型和体外人细胞实验中均表现出显著的减轻免疫活化的功效。这些发现表明，LLDT-8可能具有作为管理与SIV/HIV感染相关的全身免疫激活的药物的潜力。需要进一步的前瞻性临床探索。
方法：为了评估LLDT-8的治疗效果，我们对8只感染猿猴免疫缺陷病毒（SIV）的中国猕猴进行了流式细胞术和RNA-seq分析。此外,我们进行了全面的转录组学分析，包括横截面和纵向差异表达基因(DEG)分析，基因集富集分析(GSEA)，加权基因共表达网络分析(WGCNA)，使用来自14个时间点的外周血单核细胞（PBMC）样品进行去卷积分析。这些发现通过对接受LLDT-8治疗的患者的RNA-seq分析得到了进一步验证。以及使用人PBMC的体外细胞实验。
结果：流式细胞术分析显示，LLDT-8治疗可显著降低SIV感染的恒河猴中HLA-DR+CD38+CD8+T细胞的百分比（P<0.001）。横截面和纵向分析确定了2531和1809DEG，分别。GSEA分析表明，LLDT-8治疗导致增殖相关途径的显著下调，例如E2F目标，G2M检查点,和有丝分裂纺锤体途径。WGCNA分析鉴定了与CD8活化水平相关的两个模块和202个hub基因。去卷积分析显示在LLDT-8处理期间CD8+T细胞和活化的CD4+T细胞的比例显著降低。基因本体论结果表明，LLDT-8治疗的患者和恒河猴之间的常见DEG主要富集在细胞活化和细胞周期进程中。此外，体外细胞实验验证了LLDT-8在抑制增殖方面的一致影响，激活（HLA-DR和CD38表达），耗尽（PD-1表达），和人CD4+和CD8+T细胞中IFN-γ的产生。
结论：LLDT-8在体内动物模型和体外人细胞实验中均表现出显著的减轻免疫活化的功效。这些发现表明，LLDT-8可能具有作为管理与SIV/HIV感染相关的全身免疫激活的药物的潜力。需要进一步的前瞻性临床探索。
