LBPO.ET02 · 实验与分子治疗 · Late-Breaking
以CTC/cfDNA生物标志物为指导的精准医学克服肝细胞癌免疫检查点抑制剂耐药
CTC/cfDNA biomarker-guided precision medicine to overcome immune checkpoint inhibitor resistance in hepatocellular carcinoma
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摘要 Abstract
中文摘要
背景/目的：免疫检查点抑制剂(ICs)已改善晚期肝细胞癌(HCC)的生存结局，但大多数HCC患者会出现治疗耐药。因此，我们旨在对HCC患者进行分层，以靶向治疗耐药的TICs，并通过干性mRNA谱分析识别应答者。我们的第二个目的是评估新型CTC特征，以预测对标准抗VEGFA+抗PD-L1抗体免疫治疗联合癌症驱动基因靶向化疗的疗效。
方法：采用全基因组CRISPR-Cas9敲除(GeCKO)方法鉴定杀伤表达TP53、CTNNB1和ARID2的肿瘤所需的基因。为确认药物疗效，我们采用CRISPR/Cas9介导的驱动基因敲入和敲除，并结合酒精西式饮食喂养，以在携带ARID1A突变或TP53突变的人源化FNRG(Fah-/-;Nod;Rag1-/-;Il2rgc-/-)小鼠PDX模型中促进HCC发生。在遗传学明确的PDX及这些人源化FNRG小鼠中测试了与ICI配伍的药物用于HCC的治疗。
结果：在ARID1A突变型HCC中的GeCKO筛选显示，多梳抑制复合物2(PRC2)抑制剂、抗PD-L1和抗VEGFA的联合治疗可缩小具有人源化免疫系统的ARID1A突变型PDX小鼠的肿瘤体积并延长生存。这些方法中最成功的组合——针对ARID1A突变型HCC的抗EZH1/2抑制剂+抗PD-L1+抗VEGFA，或针对TP53突变型HCC的全反式维甲酸(ATRA)和HDAC抑制剂联合抗PD-L1——根除了在小鼠体内所生成肿瘤离体培养的人患者来源肝TICs。对患者HCC的单细胞RNA-seq分析鉴定出16个具有统计学显著性的上调基因特征。主成分分析清晰地区分了巴塞罗那临床肝癌(BCLC)分期类别A(良性)与B(恶性)。酒精西式饮食相关的TLR4-AKT信号导致PRC2复合物中EZH2的磷酸化，使其从组蛋白抑制活性转为转录激活。对PRC2组分的最有效靶向消除了在小鼠体内离体培养的人患者来源肝TICs。
结论：我们这一创新方法运用精准医学识别最可能对靶向TICs及癌症驱动突变的特定药物组合产生应答的患者，将通过解决HCC治疗耐药的核心环节而革新癌症治疗。
查看英文原文 English abstract
Background/Aim: Immune checkpoint inhibitors (ICs) have improved survival outcomes in advanced hepatocellular carcinoma (HCC), but most HCC patients experience treatment resistance. Thus, we aimed to stratify HCC patients to target therapy-resistant TICs and identify responders through stemness mRNA profiling. Our second aim was to evaluate novel CTC signatures to predict treatment efficacy in response to standard anti-VEGFA+anti-PD-L1 antibody immunotherapy with cancer-driver-targeted chemotherapies.
Methods: The Genome-wide CRISPR-Cas9 knockout (GeCKO) method was used to identify genes required for killing tumors expressing TP53, CTNNB1, and ARID2. To confirm the drug efficacy, we used CRISPR/Cas9-mediated driver gene knock-ins and knockouts combined with alcohol Western diet feeding to promote HCC development in a humanized FNRG ( Fah - / - ;Nod;Rag1 - / - ;Il2rgc - / - ) mouse harboring ARID1A mutation or TP53 mutation in a PDX Model. Drugs paired with ICI were tested for treatment of HCC in genetically defined PDX and these humanized FNRG mice.
Results: The GeCKO screening in ARID1A-mutant HCC showed that a combination of a Polycomb repressive complex 2 (PRC2) inhibitor, anti-PD-L1, and anti-VEGFA treatment reduced tumor size and extended survival in ARID1A-mutant PDX mice with humanized immune systems. The most successful combination from these approaches, comprising anti-EZH1/2 inhibitor + anti-PD-L1 + anti-VEGFA for ARID1A mutant HCC or all-trans retinoic acid (ATRA) and an HDAC inhibitor, along with anti-PD-L1, for TP53 mutant HCC-eradicated human patient-derived liver TICs grown ex vivo from tumors generated in mice. Single-cell RNA-seq analyses of patient HCCs identified 16 upregulated gene signatures with statistical significance. Principal component analyses provide a clear separation between Barcelona Clinic Liver Cancer (BCLC) stage categories A (benign) and B (malignant). Alcohol Western diet-associated TLR4-AKT signal leads to EZH2 phosphorylation of PRC2 complex to switch from histone suppressive activity to transactivation. The most effective targeting of PRC2 components, which eliminated human patient-derived liver TICs grown ex vivo in mice.
Conclusions: Our innovative approach, using precision medicine to identify patients most likely to respond to specific drug combinations targeting TICs and cancer-driver mutations, will revolutionize cancer treatment by addressing a core component of therapeutic resistance in HCC.
利益披露 Disclosure
Y. Cui, None..
A. Borges, None..
M. Li, None..
L. Sher, None..
B. Stiles, None..
D. G. Duda, None..
A. El-Khoueiry, None..
K. Machida, None.