肝细胞癌 PD-1/PD-L1 耐药的细胞与微环境、代谢重编程与肠肝轴微生态机制、联合干预策略及多组学标志物

肝细胞癌是原发性肝癌中最常见的类型，进展迅速、侵袭力强，治疗方法多样。但因其发生、发展所涉及的基因较多、机制复杂，不可避免地出现了治疗耐药性，导致治疗效果不明显，而耐药相关机制尚未完全明确。lncRNA是一类具有多种生物学功能的新型非编码RNA，能通过基因突变等作用于肝癌的发展、转化和侵袭，同时可以通过影响肿瘤免疫微环境、调节肿瘤细胞生物学功能等参与肝癌耐药。本文对lncRNA在肝细胞癌放化疗、靶向和免疫治疗耐药中的作用机制及相关进展进行综述，以期为解决肝癌耐药提供新思路。

程序性死亡受体1（programmed cell death protein 1，PD-1）是一种重要的免疫抑制分子，其通过与细胞程序性死亡-配体1（programmed cell death 1 ligand 1，PD-L1）结合启动T细胞程序性死亡使肿瘤细胞获得免疫逃逸。免疫检查点抑制剂通过阻断PD-1与PD-L1结合，重新激活患者自身的免疫细胞来杀伤肿瘤，进而治疗多种癌症。随着美国FDA陆续批准抗PD-1单抗nivolumab、pembrolizumab和抗PD-L1单抗atezolizumab等免疫检查点抑制剂用于治疗黑色素瘤和进展期非小细胞肺癌等，癌症治疗迎来新曙光。但是只有约20%患者在接受治疗后获得了长期疗效，且多数患者后期耐药复发。因此，确定有效的生物标记物和开发新靶点是当前提高患者免疫治疗响应率的重要任务。本文就抗PD-1/PD-L1药物在肿瘤中的作用机制、预测PD1获得性耐药的潜在生物标志物以及联合治疗的发展进行综述。

免疫治疗被认为是继手术、放疗和化疗后的第四种肿瘤治疗方法。近年来随着对免疫治疗特别是免疫检查点抑制剂研究的深入，PD-1/PD-L1通路抑制剂被批准用于许多癌种的治疗，但由于肿瘤细胞通过多种耐药机制规避免疫应答，免疫检查点阻断存在整体应答率低、原发或继发性耐药等难题。本文阐述了肿瘤免疫耐药的机制，探讨了耐药后再治疗的策略，为提高免疫检查点抑制剂的应答率、降低免疫耐药发生的概率提供理论和临床依据。

肝细胞癌(HCC)是全球最常见的恶性肿瘤之一,尤其在中国。70%80%患者确诊时已届晚期,只能接受姑息治疗。10年来Sorafenib一直是肝癌治疗的标准用药,而系统化疗方案FOLFOX4也已作为我国HCC的指南推荐。尽管如此,系统治疗的疗效不高、生存时间不长的局面仍有待突破。免疫检查点抑制剂在诸多实体瘤中的应用开启了系统治疗的新局面,而免疫检查点通路分子在病毒相关肝炎、肝硬化、肝癌的发生发展过程起到重要的作用,同时也是Sorafenib治疗HCC失败的耐药机制之一。2017年9月23日,美国食品药品管理局(FDA)基于Ⅰ/Ⅱ期临床研究结果批准了Nivolumab在HCC中的应用,这标志着HCC免疫治疗新时代的到来。

肝细胞癌是原发性肝癌中最常见的类型，具有高度异质性，其肿瘤细胞的基因表达差异、细胞亚群的异常分布及肿瘤微环境中细胞的复杂交互作用是导致部分患者免疫和靶向治疗失败的关键因素。理解肝细胞癌异质性对于揭示治疗耐药机制及开发新的治疗策略至关重要。时空组学技术可以在连续的时间和空间维度下解析组织细胞水平的分子表达图谱。近年来，以空间转录组和空间蛋白质组为代表的时空组学技术快速发展，两者在揭示组织表达谱特征的同时保留了空间位置信息，能确定分子在细胞或组织中的分布、定位和相互作用，为解析肝细胞癌肿瘤微环境异质性和治疗反应的差异性提供了新的研究视角。本文综述了时空组学技术在揭示肝细胞癌异质性包括细胞亚群、肿瘤微环境及治疗反应异质性方面的研究进展。

肝细胞癌是常见且致死率较高的恶性肿瘤，近年来，肠道菌群介导的肠-肝轴在肝细胞癌中的作用受到广泛关注。研究表明，肠道菌群失调可通过诱发炎症反应、调控免疫反应及驱动代谢重编程等机制促进肝细胞癌的发生发展。特定菌群的组成变化在肝细胞癌早期筛查中展现出潜在的诊断价值。此外，肠道微生态可显著影响肝细胞癌患者术后恢复和综合治疗反应，提示肠道菌群可作为肝细胞癌治疗的潜在靶点。目前，饮食调控、益生菌/益生元、粪菌移植等干预手段在改善肠-肝微环境、增强治疗敏感性以及延缓疾病进展方面显示出良好前景。本文就肠道菌群在肝细胞癌发病机制、早期诊断与治疗干预中的研究进展进行系统阐述，旨在为肝细胞癌的综合防治提供理论基础与新策略。

原发性肝癌是全球发病率和病死率较高的恶性肿瘤之一，其中肝细胞癌占90%以上。肠道和肝脏通过门静脉、胆道和体循环的双向连接进行交流，肠肝轴成为肝细胞癌发病机制中的重要环节。肠道屏障功能障碍和肠道微生物失调是肝细胞癌发生的重要因素。中医药临床应用已有数千年的历史，基于肠肝轴理论，中医药防治肝细胞癌可有效阻断肠道-肝脏信号，对肝细胞癌的精准预防与治疗具有重要意义。本文在中医药脏腑理论和肠肝轴理论的基础上，总结中医药及其活性成分对肝细胞癌的调节及干预作用，希望为肝细胞癌的进一步临床防治提供新的策略和思路。

肠道菌群在生命健康以及疾病发生发展过程中起着举足轻重的作用，能够调节新陈代谢，并参与炎症 and 免疫的关键阶段。最近的研究已经表明肠道菌群在癌症的发生以及小鼠模型和患者的抗癌免疫反应中发挥重要作用。肝细胞癌（HCC）是全球第三位癌症死亡原因，大多数HCC发生在有慢性肝病的患者中，主要由病毒性肝炎、非酒精性脂肪性肝病（NAFLD）和酒精性肝病发展而来。本文回顾分析肠道菌群在HCC中的作用，探讨菌群失调、肠泄露及菌群代谢物通过肠-肝轴促进HCC发展的机制，并对近年来肠道菌群作为HCC的潜在诊断工具，以及在免疫治疗背景下肠道菌群作为新治疗靶点的进展做一综述。

原发性肝癌的潜在病因繁多且起病隐匿,病情的进展受多方面因素影响。免疫治疗与靶向药物治疗一直是作为肝癌常规的非根治性治疗方法,但疗效并不理想,耐药情况不容乐观。近年来16s高通量测序技术的广泛应用与微生物组学的不断深入研究,揭示了微生物在肝癌发生发展中的关键作用。肝脏与口腔、肠道菌群的联系逐渐明确,口腔与肠道的菌群调控为该病开辟了新的治疗方法。综述了口-肠-肝轴的微生物理论基础及其在原发性肝癌治疗上的应用和发展方向。

目的：探讨对于多线抗肿瘤治疗失败合并恶液质的恶性肿瘤患者，予以肠菌移植联合免疫检查点抑制剂（ICIs）的疗效及安全性，探索患者血清免疫及肠道微生物环境的变化。方法：入组5例多线抗肿瘤治疗后失败的恶液质肿瘤患者，予以特瑞普利单抗联合肠菌移植治疗，每2～3周期进行疗效评估，观察患者不良反应并动态检测粪便16srRNA基因测序、血清免疫学指标。结果：除1名患者因入组时肿瘤负荷过大在移植2.5个月后死亡，其余4名患者的总生存期延长（7.4、8.3、28.5、52.3个月），1名肺腺癌颅内多发转移患者在肠菌移植后颅内转移病灶明显缩小，几乎消失。患者血清IL-2、IL-10、TGF-β等指标随着移植时间的增长，先迅速升高后缓慢降低，最终高于移植前，有统计学差异。16srRNA基因测序发现移植后患者总体肠道菌群结构分布出现明显差异，并逐渐向健康移植的供体靠拢，普拉梭菌可能是该类患者免疫治疗的疗效预测标志物。所有患者未出现2级以上不良反应，安全性良好。结论：肠菌移植联合免疫治疗或可改善生活质量、血清免疫环境及肠道微生物构成，对生存预后带来积极影响，且安全可控，为终末期恶液质患者开拓新的治疗手段。

三级淋巴结构（tertiary lymphoid structures，TLSs）可发生于包括肿瘤在内的多种病理情况下，反映了非淋巴组织内的淋巴样新生，其功能和结构类似于淋巴结，在产生和调节肿瘤局部免疫反应强弱，抑制肿瘤细胞增殖、侵袭与转移中具有重要作用。TLSs与大多数实体瘤患者的长期生存率显著相关，且TLSs可评估接受免疫检查点抑制剂治疗患者的治疗有效性。多项动物学实验证实，通过靶向肿瘤组织中TLSs形成和成熟，可促进抗肿瘤免疫反应效能，有效抑制肿瘤的恶性生物学行为。因此，本文将就TLSs的结构特征及形成机制、功能特点以及在肿瘤患者预后和治疗疗效评估中的作用等方面作一综述。

原发性肝癌居我国癌症非成熟死亡原因的首位，其中肝细胞癌占93.0%，84.4%是乙肝病毒（HBV）慢性感染所致。在HBV致癌过程中，非可控性炎性反应通过IL-6上调胞苷脱氨酶AID/APOBEC3s家族成员以促病毒和宿主基因变异、下调尿嘧啶DNA糖基化酶（UNG）等降低变异修复，炎性微环境为HBV和人肝细胞的“变异-选择-适应”进化过程提供了必要条件并促进变异细胞逆向分化。该进化过程需满足体细胞驱动突变和免疫抑制微环境这两个前提条件。促癌基因组不稳定和HBV变异通过cGAS-STING抑制Ⅰ型干扰素信号，诱导免疫抑制性炎性反应，招募抑制性免疫细胞如TAMs、Treg、MDSC等构建肿瘤微环境。微环境中肿瘤细胞迅速生长所致的缺氧通过诱导免疫抑制性炎性反应因子激活肿瘤犬尿氨酸代谢，增强Treg的PD-1表达以加强免疫耐受、抑制CD8+T细胞和NK细胞毒性、促进新血管生成。以上“癌症进化发育学”理论为癌症的防治指明了方向。针对新血管生成的靶向治疗能显著遏制肝癌生长、提高抗肿瘤免疫，加强免疫治疗效果。靶向-免疫联合治疗应成为晚期肝癌的主要治疗措施。

肝细胞癌治疗已进入免疫联合治疗时代。双免疫疗法(如O+Y和STRIDE方案)基于关键Ⅲ期研究成为标准一线选择。其核心价值在于诱导深度且持久的免疫应答，带来显著的“拖尾效应”，使部分患者获得长期生存可能，并为存在抗血管生成治疗禁忌者提供“纯免疫”选项。然而，其疗效呈现异质性，且存在早期生存曲线交叉和与传统实体瘤疗效评价标准(Response Evaluation Criteria in Solid Tumours, RECIST)不匹配等挑战。临床决策需个体化权衡：对于需快速缩瘤(如伴门静脉癌栓)者，靶免联合治疗更具早期控制优势；对于追求长期生存、肝功能较差或存在特定禁忌者，双免疫疗法价值凸显。未来方向在于探索“双免+”联合策略和针对淋巴细胞活化基因3(lymphocyte activation gene-3, LAG-3)和T细胞免疫球蛋白黏蛋白分子3(T cell immunoglobulin and mucin domain-containing protein 3, TIM-3)等新靶点的药物，并利用生物标志物实现精准诊疗，推动治疗迈向“患者匹配”的新阶段。

肝癌是一种严重的全球健康问题，也是常见的癌症相关死亡原因。肝细胞癌（hepatocellular carcinoma, HCC）是肝癌常见的病理类型。早期HCC临床症状不明显，50%的HCC患者确诊时已处于晚期。系统全身治疗被推荐用于晚期HCC。随着分子靶向药物（索拉非尼、仑伐替尼）的发展，晚期HCC的系统全身治疗取得了一定进展，但对HCC患者生存获益仍然不大。近年来，免疫检查点抑制剂的出现改变了HCC治疗的格局，为HCC精准治疗提供了更多的可能性并展现出较好的效果。特别是阿替利珠单抗和贝伐珠单抗的联合疗法显著改善了HCC患者的生存预后，此外，过继性细胞疗法、肿瘤疫苗、溶瘤病毒和非特异性免疫治疗也已成为免疫治疗策略。本文就HCC免疫治疗的现状及发展进行概述。

肝细胞肝癌是全球第六大常见癌症，预后极差。现行肝癌治疗手段包括手术切除、肝移植、射频消融及经肝动脉栓塞化疗等，但这些治疗方案在晚期肝癌患者中收效甚微。目前，针对晚期肝癌患者，以细胞增殖、转移和血管生成为靶点的治疗策略也取得一定的疗效，但总体效果仍未能令人满意。值得注意的是，肝癌治疗中化疗、靶向治疗及免疫治疗的有效性与患者自身（原发性）或治疗后（继发性）的组织免疫微环境密切相关。总结肝癌免疫微环境塑造和功能的研究新进展，介绍新近肝癌临床治疗策略，重点关注现行治疗策略对局部肿瘤免疫微环境的影响，并进一步讨论目前肝癌临床前研究对后续新治疗策略产生的启发。

肝细胞癌（hepatocellular carcinoma，HCC）是全球范围内高发病率与死亡率的恶性肿瘤，其发生发展与代谢异常密切相关。线粒体作为细胞的能量代谢中心和信号枢纽，在肝细胞癌中经历复杂且多样的改变与重塑，深刻影响着肿瘤的起始、进展及治疗应答。然而，线粒体发生改变的具体机制及其在肝细胞癌中的作用尚未完全阐明，这在一定程度上阻碍了以线粒体为靶点的肝细胞癌治疗药物的研发与应用。从代谢重编程、氧化应激与线粒体DNA损伤以及线粒体动力学失调三个方面，系统总结了肝细胞癌中的线粒体改变，以及线粒体的改变在肝细胞癌发生发展和治疗应答中的作用机制，并对当前靶向线粒体的肝细胞癌治疗的相关研究进行了梳理与展望，以期为后续治疗肝细胞癌的基础研究与临床干预提供理论依据和新的思路。

肿瘤是世界级公共卫生问题，导致了全球每年约六分之一的死亡。在诊断、治疗和监测的每个阶段实现对患者的精准化管理，既是当前临床实践中的一大挑战，也可作为提升患者生存率的重要策略。液体活检相较于传统检测具有创伤性小、可反复采样的优势。通过对患者身体易于重复获取的生物样本进行检测，液体活检能够在疾病诊断、治疗及预后随访全阶段提供信息，成为临床上实现对肿瘤微创、实时、全面监控的极具前景的组织样本替代方法。本文系统综述了肿瘤领域中常见的液体活检标志物及其最新检测技术，涵盖脑脊液、粪便等非血液样本的检测方案，并提出了基于“多标志物联合+全样本覆盖”的肿瘤全程精准管理新框架。文中也探讨了当前将液体活检发展为成熟临床检测项目所面临的一系列挑战，包括样本检测流程标准化、标准化报告制度的建立，以及如何平衡检测方法的普及性与成本控制等问题，以推动液体活检在肿瘤早筛、治疗革新与广泛应用方面的发展。我们期望通过构建从基础研究到转化生产、临床应用的全链条体系，开发一体化检测平台，建立标准化报告流程与完善监管机制，为患者提供从诊断到康复的全周期精准管理，最终使肿瘤从“不治之症”转变为“可防可控”的慢性疾病。

The application of immune checkpoint inhibitors (ICIs) has markedly enhanced the treatment of hepatocellular carcinoma (HCC), and HCC patients who respond to ICIs have shown prolonged survival. However, only a subset of HCC patients benefit from ICIs, and those who initially respond to ICIs may develop resistance. ICI resistance is likely related to various factors, including the immunosuppressive tumor microenvironment (TME), the absence of antigen expression and impaired antigen presentation, tumor heterogeneity, and gut microbiota. Therefore, exploring the possible mechanisms of ICI resistance is crucial to improve the clinical benefit of ICIs further. Various combination therapies for HCC immunotherapy have prevented and reversed ICI resistance to a certain extent. In addition, many new combination therapies that can overcome resistance are being explored. This review seeks to characterize the complex TME in HCC, explore the possible mechanisms of immune resistance to ICIs in different resistance categories, and review the combination therapies currently being applied and those under investigation for immunotherapy.

Hepatocellular carcinoma is one of the leading causes of cancer mortality globally, and its incidence is increasing. Immune checkpoint therapy has revolutionized the treatment of hepatocellular carcinoma over the past few years. However, only a limited proportion of patients with hepatocellular carcinoma respond to immunotherapy. Despite the significant breakthroughs, the molecular mechanisms that drive immune responses and evasion are largely unresolved. Predicting tumor response and resistance to immune checkpoint inhibitors is a significant challenge. In this review, we focus on the current research progress of immune checkpoint inhibitors in hepatocellular carcinoma. Importantly, this review highlights the underlying mechanisms of resistance to immune checkpoint inhibitors and summarizes potential strategies to overcome the resistance to immune checkpoint inhibitors in hepatocellular carcinoma.

Hepatocellular carcinoma (HCC) is the most common primary liver malignancy worldwide and a leading cause of death worldwide. Its incidence continues to increase in the US due to hepatitis C infection and nonalcoholic steatohepatitis. Liver transplantation and resection remain the best therapeutic options for a cure, but these are limited by the shortage of available organs for transplantation, diagnosis at an advanced stage, and underlying chronic liver disease found in most patients with HCC. Immune checkpoint inhibitors (ICIs) have been shown to be an evolving novel treatment option in certain advanced solid tumors and have been recently approved for inoperable, advanced and metastatic HCC. Unfortunately, a large cohort of patients with HCC fail to respond to immunotherapy. In this review, we discuss the ICIs currently approved for HCC treatment and their various mechanisms of action. We will highlight the current understanding of the mechanism of resistance and limitations to ICIs. Finally, we will describe emerging biomarkers of response to ICIs and address future direction on overcoming resistance to immune checkpoint therapy.

… with intermediate and advanced HCC has become the focus of … drug resistance with immune combination regimens for HCC in … resistance to ICIs HCC and exploring sensitive/resistant …

Abstract Hepatocellular carcinoma (HCC) is a concerning liver cancer with rising incidence and mortality rates worldwide. The effectiveness of traditional therapies in managing advanced HCC is limited, necessitating the development of new therapeutic strategies. Immune checkpoint inhibitors (ICIs) have emerged as a promising strategy for HCC management. By preventing tumor cells from evading immune surveillance through immunological checkpoints, ICIs can restore the immune system’s ability to target and eliminate tumors. While ICIs show promise in enhancing the immune response against malignancies, challenges such as drug resistance and adverse reactions hinder their efficacy. To address these challenges, developing individualized ICI treatment strategies is critical. Combining targeted therapy and immunotherapy holds the potential for comprehensive therapeutic effects. Additionally, biomarker-based individualized ICI treatment strategies offer promise in predicting treatment response and guiding personalized patient care. Future research should explore emerging ICI treatment methods to optimize HCC immunotherapy. This review provides an overview of ICIs as a new treatment for HCC, demonstrating some success in promoting the tumor immune response. However, drug resistance and adverse reactions remain important considerations that must be addressed. As tailored treatment plans evolve, the prospect of immunotherapy for HCC is expected to grow, offering new opportunities for improved patient outcomes.

Simple Summary Immune checkpoint inhibitors (ICIs) are now a cornerstone of systemic treatment for hepatocellular carcinoma (HCC). However, their efficacy is blunted by mechanisms of tumour resistance in many patients. This review reports on the state of the art of resistance to ICIs, focusing on HCC, with the aim to provide clear and direct information to clinicians and researchers. Growing knowledge on the mechanisms of resistance to immunotherapy can indeed guide the choice of and expand the application of novel combined treatments. Abstract Hepatocellular carcinoma (HCC) is the most common liver cancer and a relevant global health problem. Immune checkpoint inhibitors (ICIs) represent the most effective systemic treatment for HCC. However, due to primary resistance, approximately 40% of HCC patients do not achieve a disease control with ICIs. Moreover, a similar proportion will experience disease progression after an initial response caused by secondary resistance. This review describes the mechanisms of primary and secondary resistance and reports the ongoing therapeutic strategies to overcome these obstacles.

Hepatocellular carcinoma (HCC) constitutes most primary liver cancers and is one of the most lethal and life-threatening malignancies globally. Unfortunately, a substantial proportion of HCC patients are identified at an advanced stage that is unavailable for curative surgery. Thus, palliative therapies represented by multi-tyrosine kinase inhibitors (TKIs) sorafenib remained the front-line treatment over the past decades. Recently, the application of immune checkpoint inhibitors (ICIs), especially targeting the PD-1/PD-L1/CTLA-4 axis, has achieved an inspiring clinical breakthrough for treating unresectable solid tumors. However, many HCC patients with poor responses lead to limited benefits in clinical applications, which has quickly drawn researchers’ attention to the regulatory mechanisms of immune checkpoints in HCC immune evasion. Evasion of immune surveillance by cancer is attributed to intricate reprogramming modulation in the tumor microenvironment. Currently, more and more studies have found that epigenetic modifications, such as chromatin structure remodeling, DNA methylation, histone post-translational modifications, and non-coding RNA levels, may contribute significantly to remodeling the tumor microenvironment to avoid immune clearance, affecting the efficacy of immunotherapy for HCC. This review summarizes the rapidly emerging progress of epigenetic-related changes during HCC resistance to ICIs and discusses the mechanisms of underlying epigenetic therapies available for surmounting immune resistance. Finally, we summarize the clinical advances in combining epigenetic therapies with immunotherapy, aiming to promote the formation of immune combination therapy strategies.

In recent years, immune checkpoint inhibitors (ICIs) have revolutionized the treatment of patients with hepatocellular carcinoma (HCC). Following the positive results of the IMbrave150 trial, the combination of atezolizumab (an anti-PD-L1 antibody) and bevacizumab (an anti-VEGF antibody) became the standard of care frontline treatment for patients with advanced stage HCC. Several other trials evaluated immunotherapy in HCC, demonstrating that ICIs-based regimens are currently the most effective treatment strategies and expanding the therapeutic possibilities. Despite the unprecedent rates of objective tumor response, not all patients benefit from treatment with ICIs. Therefore, in order to select the appropriate therapy as well as to correctly allocate medical resources and avoid unnecessary treatment-related toxicities, there is great interest in identifying the predictive biomarkers of response or resistance to immunotherapy-based regimens. Immune classes of HCC, genomic signatures, anti-drug antibodies, and patient-related factors (e.g., etiology of liver disease, gut microbiota diversity) have been associated to the response to ICIs, but none of the proposed biomarkers have been translated into clinical practice so far. Considering the crucial importance of this topic, in this review we aim to summarize the available data on tumor and clinical features associated with the response or resistance of HCC to immunotherapies.

Hepatocellular carcinoma (HCC) remains a critical global health concern, particularly in regions with high endemicity of hepatitis B, hepatitis C, and non-alcoholic fatty liver disease. Immunotherapy, particularly immune checkpoint inhibitors (ICIs), has emerged as a promising therapeutic strategy for advanced HCC. Despite encouraging results, primary and acquired resistance to ICIs continues to pose significant challenges in clinical practice. Recent research has identified tumor-associated macrophages (TAMs) as key contributors to immune evasion and ICI resistance in HCC, primarily through polarization to the M2 phenotype. M2-polarized TAMs secrete a range of immunosuppressive cytokines that inhibit T cell activation and promote tumor progression through processes such as angiogenesis and epithelial-mesenchymal transition. These mechanisms compromise the efficacy of ICIs and facilitate tumor expansion and metastasis. This review summarizes the role of TAM-related signaling pathways in driving immune evasion and ICI resistance in HCC, with particular emphasis on the contribution of TAM surface receptors and chemokines in immune suppression. Additionally, the review highlights emerging insights into TAM metabolic reprogramming and transcriptional regulation, which have been closely linked to ICI resistance. Furthermore, we explore promising therapeutic strategies targeting TAMs and their associated signaling pathways to enhance ICI efficacy in HCC. Integrating these novel approaches could potentially overcome TAM-driven immune evasion and ICI resistance, boosting the efficacy of immunotherapy and improving patient prognosis in HCC.

Hepatocellular carcinoma (HCC) is the typical inflammation-induced neoplasia. It often prospers where a chronic liver disease persists, thus leading a strong rationale for immune therapy. Several immune-based treatments, including immune checkpoint inhibitors (ICI), cytokines, adoptive cell transfer, and vaccines, have been tested in the treatment of HCC. In this review, we summarize the role of the ICI in HCC patients in various sets of treatment. As for advanced HCC, the anti-Programmed cell Death protein 1 (PD1) antibodies and the anti-Cytotoxic T-Lymphocyte Antigen 4 (CTLA-4) antibodies have been examined in patients with enthusiastic results in phase I-II-III studies. Overall, this led the Food and Drug Administration (FDA) to approve pembrolizumab, nivolumab, and nivolumab + ipilimumab in the second-line setting. The anti- Programmed Death-Ligand 1 (PDL-1) antibodies have also been evaluated. Thanks to the results obtained from phase III IMbrave study, atezolizumab + bevacizumab is now the standard of care in the first-line advanced setting of HCC. As for localized HCC, the putative immunological effect of locoregional therapies led to evaluate the combination strategy with ICI. This way, chemoembolization, ablation with radiofrequency, and radioembolization combined with ICI are currently under study. Likewise, the study of adjuvant immunotherapy following surgical resection is underway. In addition, the different ICI has been studied in combination with other ICI as well as with multikinase inhibitors and anti-angiogenesis monoclonal antibody. The evidence available suggests that combining systemic therapies and locoregional treatments with ICI may represent an effective strategy in this context.

… with advanced HCC treated with immune checkpoint inhibitors failed to … resistance to immune checkpoint inhibitors through upregulation of indoleamine 2,3-dioxygenase (IDO) in HCC. …

Hepatocellular carcinoma (HCC) poses a significant clinical burden due to its aggressive nature, profound tumor heterogeneity, and limited therapeutic efficacy. While immune checkpoint inhibitors (ICIs) have revolutionized treatment paradigms and demonstrated considerable promise, the emergence of resistance mechanisms has posed a critical challenge in contemporary clinical oncology. The accelerated development of novel agents and innovative combination strategies has further complicated this resistance landscape. In this review, we present a unique and comprehensive analysis of ICI resistance mechanisms in HCC by integrating insights into primary resistance, acquired resistance, and host-related factors. Building upon this mechanistic framework, we explore emerging therapeutic strategies to overcome ICI resistance. Furthermore, we evaluate the dual role of ICIs in HCC management - serving as a neoadjuvant therapy for transplant candidates while simultaneously posing risks of post-transplant rejection. By bridging preclinical discoveries with clinical realities, this analysis aims to inform rational therapeutic design and optimize immuno-oncology trials for HCC patients.

This review focuses on the opportunities and challenges of immune checkpoint inhibitor therapies for hepatocellular carcinoma, especially hepatitis B virus‐related hepatocellular carcinoma.

HCC comprises ∼80% of primary liver cancer. HCC is the only major cancer for which death rates have not improved over the last 10 years. Most patients are diagnosed with advanced disease when surgical and locoregional treatments are not feasible or effective. Sorafenib, a multikinase inhibitor targeting cell growth and angiogenesis, was approved for advanced unresectable HCC in 2007. Since then, other multikinase inhibitors have been approved. Lenvatinib was found to be noninferior to sorafenib as a first-line agent. Regorafenib, cabozantinib, and ramucirumab were shown to prolong survival as second-line agents. Advances in immunotherapy for HCC have also added hope for patients, but their efficacy remains limited. A large proportion of patients with advanced HCC gain no long-term benefit from systemic therapy due to primary and acquired drug resistance, which, combined with its rising incidence, keeps HCC a highly fatal disease. This review summarizes mechanisms of primary and acquired resistance to therapy and includes methods for bypassing resistance. It addresses recent advancements in immunotherapy, provides new perspectives on the linkage between drug resistance and molecular etiology of HCC, and evaluates the role of the microbiome in drug resistance. It also discusses alterations in signaling pathways, dysregulation of apoptosis, modulations in the tumor microenvironment, involvement of cancer stem cells, changes in drug metabolism/transport, tumor hypoxia, DNA repair, and the role of microRNAs in drug resistance. Understanding the interplay among these factors will provide guidance on the development of new therapeutic strategies capable of improving patient outcomes.

Resistance limits the efficacy and durability of immune checkpoint inhibitors (ICIs) in hepatocellular carcinoma (HCC). Therefore, we conducted a retrospective cohort study to investigate the outcomes and characteristics of HCC patients with resistance to immunotherapy. Patients with HCC who have received ICIs at Eastern Hepatobiliary Surgery Hospital between 2016 and 2021 were retrospectively screened and divided into primary resistance, secondary resistance, and durable response group. Time to progression (TTP), overall survival (OS), subsequent management and post-progression survival (PPS) were analyzed. Of 496 patients included, 229 (46.2%) and 141 (28.4%) patients developed primary and secondary resistance, and 126 (25.4%) patients achieved a durable response, the median TTP was 2.83 [2.56–3.09] months, 11.93 [10.45–13.40] months, and not reached, respectively, whereas the median OS was 12.83 [10.36–15.30] months, 31.53 [28.09–34.97] and not reached, respectively. Multivariate logistic regression revealed that Child–Pugh score, BCLC stage, and combined systemic therapies (ICI plus bevacizumab or lenvatinib versus ICI monotherapy) were independently associated with primary resistance, and only combined systemic therapies (ICI plus bevacizumab versus ICI monotherapy) were independently associated with secondary resistance. AFP levels were independently associated with PPS in patients with primary resistance, while post-progression therapies (ICI-based therapies versus others) were independently associated with PPS in patients with resistance. The risk of resistance was notably lower in patients receiving the combination of ICI plus bevacizumab. High AFP levels were associated with the survival of patients with primary resistance. ICI-based maintenance therapy after resistance may provide a significant survival advantage for HCC patients.

Simple Summary Immune checkpoint inhibitors (ICIs) have become an important treatment for hepatocellular carcinoma (HCC), a common form of liver cancer. However, a significant number of patients do not respond well to this treatment, and many who initially benefit eventually develop resistance. This research aims to better understand the mechanisms behind resistance to ICIs in HCC, which could help improve patient outcomes. By identifying factors such as immune system dysfunction, changes in the tumor microenvironment, and genetic mutations, the study seeks to develop strategies to overcome these barriers. The findings may guide future treatment approaches, combining ICIs with other therapies to enhance effectiveness, offering hope for more patients with this challenging disease.

Simple Summary Hepatocellular carcinoma (HCC) is the most common primary hepatic malignancy, with increasing incidence over the past several decades. The majority of patients with HCC present with advanced unresectable disease, making treatment options with curative intent limited and survival outlooks dismal. Systemic therapy with sorafenib had been traditionally used, with marginal benefit. Immunotherapy, successfully used to treat other malignant tumors, has recently been shown to be safe and well tolerated and to have promising long-term outcomes in patients with advanced HCC. We herein review the outcomes of immune checkpoint inhibitors (ICI) from major clinical trials, summarize predictors of treatment response, and highlight adverse events related to ICI treatment. Abstract Hepatocellular carcinoma (HCC) is the most frequent primary liver tumor. As a result of advanced disease being often present at diagnosis, only a small percentage of patients are amenable to curative-intent treatment options such as surgical resection and liver transplantation. Systemic therapy consisting of tyrosine kinase inhibitors such as sorafenib had been used for over a decade with limited efficacy. More recently, treatment with immune checkpoint inhibitors has revolutionized the treatment landscape of various malignant tumors. With this shifting paradigm, recent data have demonstrated encouraging outcomes among patients with HCC. In particular, several trials have investigated the safety and efficacy of various immune checkpoint inhibitors (ICI) either as monotherapy or in the form of combined treatments. We sought to provide an overview of recent clinical trials among patients with advanced HCC as well as to highlight predictors of response and immune-related adverse events and to review the evidence on perioperative administration of ICI in patients with resectable HCC.

Abstract Primary liver cancer, represented mainly by hepatocellular carcinoma (HCC) and intrahepatic cholangiocarcinoma (CCA), is one of the most common and deadliest tumors worldwide. While surgical resection or liver transplantation are the best option in early disease stages, these tumors often present in advanced stages and systemic treatment is required to improve survival time. The emergence of immune checkpoint inhibitor (ICI) therapy has had a positive impact especially on the treatment of advanced cancers, thereby establishing immunotherapy as part of first-line treatment in HCC and CCA. Nevertheless, low response rates reflect on the usually cold or immunosuppressed tumor microenvironment of primary liver cancer. In this review, we aim to summarize mechanisms of resistance leading to tumor immune escape with a special focus on the composition of tumor microenvironment in both HCC and CCA, also reflecting on recent important developments in ICI combination therapy. Furthermore, we discuss how combination of ICIs with established primary liver cancer treatments (e.g. multikinase inhibitors and chemotherapy) as well as more complex combinations with state-of-the-art therapeutic concepts may reshape the tumor microenvironment, leading to higher response rates and long-lasting antitumor immunity for primary liver cancer patients.

Importance For more than a decade, sorafenib has been the only systemic treatment option for patients with advanced hepatocellular carcinoma (HCC). However, rapid progress over the past few years led to approval of other angiogenesis inhibitors and several immune checkpoint blockers (ICBs) that have been added to the treatment armamentarium for advanced HCC. Moreover, the recent success of a combination of bevacizumab with atezolizumab signals an important change in the front-line treatment of HCC. Observations This review summarizes rapidly emerging clinical data on the promise and challenges of implementing ICBs in HCC and discusses the unmet need of biomarkers to predict response or resistance to therapy. Two strategies to target immunosuppression in tumors are also discussed: one proven (vascular endothelial growth factor pathway inhibition) and one currently under investigation (transforming growth factor-β pathway inhibition). The rationale and preliminary evidence on how their inhibition may reprogram the immunosuppressive milieu and enhance the efficacy of ICBs in HCC are reviewed. Conclusion and Relevance The recent successes and failures of angiogenesis inhibitors and ICBs, alone and in combination, have provided important insights into how to implement this novel systemic therapy in HCC and led to new avenues to enhance immunotherapy efficacy in this disease.

Approximately 11–37% of hepatocellular carcinomas (HCC) have WNT/β-catenin mutations [1]. These HCCs are classified as either S3, according to the molecular classification of HCC proposed by Hoshida et al. [2], or G5-G6, according to the molecular classifications of Boyault et al. [3] and Zucman-Rossi et al. [4], and they have a more favorable prognosis than other subclasses [5, 6] (Fig. 1). The development of immune checkpoint inhibitor (ICI) therapies led to the classification of HCC into immune subclasses according to the tumor microenvironment (TME), which should affect the outcome of ICI therapy [7–10]. For example, Llovet et al. [10] proposed 3 Received: June 5, 2020 Accepted: June 18, 2020 Published online: August 20, 2020

As one of the emerging hallmarks of tumorigenesis and tumor progression, metabolic remodeling is common in the tumor microenvironment. Hepatocellular carcinoma (HCC) is the third leading cause of global tumor-related mortality, causing a series of metabolic alterations in response to nutrient availability and consumption to fulfill the demands of biosynthesis and carcinogenesis. Despite the efficacy of immunotherapy in treating HCC, the response rate remains unsatisfactory. Recently, research has focused on metabolic reprogramming and its effects on the immune state of the tumor microenvironment, and immune response rate. In this review, we delineate the metabolic reprogramming observed in HCC and its influence on the tumor immune microenvironment. We discuss strategies aimed at enhancing response rates and overcoming immune resistance through metabolic interventions, focusing on targeting glucose, lipid, or amino acid metabolism, as well as systemic regulation.

The liver is essential for metabolic homeostasis. The onset of liver cancer is often accompanied by dysregulated liver function, leading to metabolic rearrangements. Overwhelming evidence has illustrated that dysregulated cellular metabolism can, in turn, promote anabolic growth and tumor propagation in a hostile microenvironment. In addition to supporting continuous tumor growth and survival, disrupted metabolic process also creates obstacles for the anticancer immune response and restrains durable clinical remission following immunotherapy. In this review, we elucidate the metabolic communication between liver cancer cells and their surrounding immune cells and discuss how metabolic reprogramming of liver cancer impacts the immune microenvironment and the efficacy of anticancer immunotherapy. We also describe the crucial role of the gut–liver axis in remodeling the metabolic crosstalk of immune surveillance and escape, highlighting novel therapeutic opportunities.

Hepatocellular carcinoma is the most common primary liver cancer, ranking third among the leading causes of cancer-related mortality worldwide and whose incidence varies according to geographical area and ethnicity. Metabolic rewiring was recently introduced as an emerging hallmark able to affect tumor progression by modulating cancer cell behavior and immune responses. This review focuses on the recent studies examining HCC’s metabolic traits, with particular reference to the alterations of glucose, fatty acid and amino acid metabolism, the three major metabolic changes that have gained attention in the field of HCC. After delivering a panoramic picture of the peculiar immune landscape of HCC, this review will also discuss how the metabolic reprogramming of liver cancer cells can affect, directly or indirectly, the microenvironment and the function of the different immune cell populations, eventually favoring the tumor escape from immunosurveillance.

Hepatocellular carcinoma (HCC), the most common primary liver cancer, continues to rank among the leading causes of cancer-related death despite improvements in early detection and systemic therapies. Therapeutic advances, including immune checkpoint blockade, cancer vaccines, and adoptive cell therapies, have broadened treatment possibilities. However, their efficacy and durability are often limited by immune evasion within a metabolically challenging tumor microenvironment (TME). This review consolidates current knowledge on how metabolic reprogramming in immune cells influences HCC progression, therapy resistance, and clinical outcomes. We discuss the roles of glycolysis, oxidative phosphorylation, fatty acid oxidation, and amino acid metabolism kynurenine pathways—in regulating the differentiation and function of T cells, regulatory T cells, macrophages, dendritic cells, natural killer cells, and B cells. Environmental factors such as hypoxia, lactate accumulation, adenosine signaling, and lipid remodeling act as key TME cues that suppress antigen presentation, impair cytotoxic responses, and promote immunosuppressive myeloid phenotypes. Building on these mechanisms, current strategies focus on targeting metabolic checkpoints in immune cells, reshaping the TME, and integrating metabolic modulation with checkpoint inhibitors to enhance therapeutic efficacy. In addition, candidate biomarkers (including circulating metabolites, multi-omics profiles, and liquid-biopsy indicators of immune metabolism) offer opportunities for patient stratification and dynamic monitoring. Together, these insights provide a conceptual framework in which precise modulation of immune metabolism can potentiate existing immunotherapies and guide rational combination strategies, warranting further clinical investigation to achieve sustained benefit in HCC.

Metabolic reprogramming is one of the major biological features of malignant tumors, playing a crucial role in the initiation and progression of cancer. The tumor microenvironment consists of various non-cancer cells, such as hepatic stellate cells, cancer-associated fibroblasts (CAFs), immune cells, as well as extracellular matrix and soluble substances. In liver cancer, metabolic reprogramming not only affects its own growth and survival but also interacts with other non-cancer cells by influencing the expression and release of metabolites and cytokines (such as lactate, PGE2, arginine). This interaction leads to acidification of the microenvironment and restricts the uptake of nutrients by other non-cancer cells, resulting in metabolic competition and symbiosis. At the same time, metabolic reprogramming in neighboring cells during proliferation and differentiation processes also impacts tumor immunity. This article provides a comprehensive overview of the metabolic crosstalk between liver cancer cells and their tumor microenvironment, deepening our understanding of relevant findings and pathways. This contributes to further understanding the regulation of cancer development and immune evasion mechanisms while providing assistance in advancing personalized therapies targeting metabolic pathways for anti-cancer treatment.

Hepatocellular carcinoma (HCC) is a typical inflammation‐induced cancer and displays a complex interaction between the tumor microenvironment and tumor development. Immune cells in the HCC microenvironment play both pro‐ and anti‐tumoral roles in HCC progression. An increasing number of findings indicate that metabolic reprogramming is essential for immune cell differentiation and function. In this review, we discuss the metabolic changes of different immune cells and correlate these findings to HCC progression.

Tumor cells possess complex immune evasion mechanisms to evade immune system attacks, primarily through metabolic reprogramming, which significantly alters the tumor microenvironment (TME) to modulate immune cell functions. When a tumor is sufficiently immunogenic, it can activate cytotoxic T-cells to target and destroy it. However, tumors adapt by manipulating their metabolic pathways, particularly glucose, amino acid, and lipid metabolism, to create an immunosuppressive TME that promotes immune escape. These metabolic alterations impact the function and differentiation of non-tumor cells within the TME, such as inhibiting effector T-cell activity while expanding regulatory T-cells and myeloid-derived suppressor cells. Additionally, these changes lead to an imbalance in cytokine and chemokine secretion, further enhancing the immunosuppressive landscape. Emerging research is increasingly focusing on the regulatory roles of non-tumor cells within the TME, evaluating how their reprogrammed glucose, amino acid, and lipid metabolism influence their functional changes and ultimately aid in tumor immune evasion. Despite our incomplete understanding of the intricate metabolic interactions between tumor and non-tumor cells, the connection between these elements presents significant challenges for cancer immunotherapy. This review highlights the impact of altered glucose, amino acid, and lipid metabolism in the TME on the metabolism and function of non-tumor cells, providing new insights that could facilitate the development of novel cancer immunotherapies.

Hepatocellular carcinoma (HCC) is a leading cause of cancer-related deaths worldwide, with limited treatment options for advanced stages. Metabolic reprogramming is a hallmark of cancer, enabling tumor cells to adapt to the harsh tumor microenvironment (TME) and evade immune surveillance. This review involves the role of metabolic reprogramming in HCC, focusing on the dysregulation of glucose, lipid, and amino acid metabolism, and its impact on immune evasion. Key metabolic pathways, such as the Warburg effect, fatty acid synthesis, and glutaminolysis, are discussed, along with their influence on tumor-associated macrophages (TAMs) and immune cell function. Targeting these metabolic alterations presents a promising therapeutic approach to enhance immunotherapy efficacy and improve HCC patient outcomes.

Metabolic reprogramming is a fundamental mechanism through which tumor cells reshape their energy metabolism to sustain rapid proliferation. It facilitates malignant growth by reprogramming key pathways, including glycolysis and amino acid metabolism. The tumor microenvironment (TME) is composed of tumor cells, stromal cells, and immune cells. The characteristics of hypoxia, acidity, and nutrient deficiency are mainly driven by the metabolic products and cytokines secreted by tumor cells. This metabolic pressure not only inhibits the functions of immune cells, but also further enhances immune evasion through nutrient competition. Targeting metabolic reprogramming can reverse immunosuppression within the TME and enhance the response to immunotherapy. This article systematically reviews the regulatory mechanisms of metabolic reprogramming in hepatocellular carcinoma and its impact on the TME, while also exploring therapeutic strategies based on metabolic interventions.

Histidine metabolism is crucial in role in tumor biology, contributing to tumor progression, immune regulation, and metabolic reprogramming. In hepatocellular carcinoma (HCC), dysregulated histidine metabolism may promote tumor growth and immune evasion, although the specific mechanisms remain poorly understood. Using single-cell RNA sequencing, the expression patterns of histidine metabolism–related genes were evaluated across different cell types in HCC samples. In vivo and in vitro experiments were conducted to validate how histidine treatment affects macrophage and T-cell function. Furthermore, the TCGA database was utilized to construct a prognostic model to identify the key gene BUD23 and to examine its correlation with metabolism and immune infiltration. The proportion of parenchymal cells exhibiting high histidine metabolism was significantly increased, accompanied by a general reduction in immune and stromal cell infiltration. Notably, macrophages and T cells demonstrated impaired antitumor functions. In the high histidine metabolism group, multiple critical cell communication pathways (e.g., MIF, CLEC, MHC II) were downregulated, macrophages shifted toward immunosuppressive subpopulations, T cells exhibited an exhaustion phenotype, and CD8 + T-cell activation was diminished. Further in vivo and in vitro co-culture experiments confirmed that elevated histidine concentrations promoted M2 polarization in macrophages and weakened T-cell cytotoxicity, accelerating tumor proliferation. According to TCGA analyses, BUD23 was upregulated in the high histidine metabolism group and significantly negatively correlated with patient survival and immune cell infiltration. Silencing BUD23 boosted immune cell activation and cytotoxic effects, effectively reversing the immunosuppressive microenvironment. A multivariable Cox regression–based prognostic model indicated unfavorable outcomes in patients with high histidine metabolism. Histidine metabolism drives tumor cell metabolic reprogramming and reshapes the tumor immune microenvironment through intercellular communication, thereby promoting tumor progression. BUD23 shows promise as a biomarker for prognosis and immune response prediction in liver cancer. This study provides new therapeutic targets and theoretical support for liver cancer treatment by targeting histidine metabolism.

Cancer cells often encounter hypoxic and hypo-nutrient conditions, which force them to make adaptive changes to meet their high demands for energy and various biomaterials for biomass synthesis. As a result, enhanced catabolism (breakdown of macromolecules for energy production) and anabolism (macromolecule synthesis from bio-precursors) are induced in cancer. This phenomenon is called “metabolic reprogramming,” a cancer hallmark contributing to cancer development, metastasis, and drug resistance. HCC and cholangiocarcinoma (CCA) are 2 different liver cancers with high intertumoral heterogeneity in terms of etiologies, mutational landscapes, transcriptomes, and histological representations. In agreement, metabolism in HCC or CCA is remarkably heterogeneous, although changes in the glycolytic pathways and an increase in the generation of lactate (the Warburg effect) have been frequently detected in those tumors. For example, HCC tumors with activated β-catenin are addicted to fatty acid catabolism, whereas HCC tumors derived from fatty liver avoid using fatty acids. In this review, we describe common metabolic alterations in HCC and CCA as well as metabolic features unique for their subsets. We discuss metabolism of NAFLD as well, because NAFLD will likely become a leading etiology of liver cancer in the coming years due to the obesity epidemic in the Western world. Furthermore, we outline the clinical implication of liver cancer metabolism and highlight the computation and systems biology approaches, such as genome-wide metabolic models, as a valuable tool allowing us to identify therapeutic targets and develop personalized treatments for liver cancer patients.

Simple Summary Liver cancer is one of the most frequently occurring cancer types and one of the leading causes of cancer-related mortality globally. Despite of its constantly growing incidence, the efficacy of the current therapeutic interventions is limited. Metabolic and epigenetic aberrations are two distinct but mutually influential factors which contribute to the plasticity and adaptability of tumor cells to a hostile microenvironment, leading to high resilience and permissive conditions. This review offers the most up-to-date overview of the recently reported major metabolic dysregulations in liver cancer cells, the consequential epigenetic reprogramming, as well as the opportunity to explore the effect on metabolic competition and inhibition of immune cells in the tumor microenvironment. Finally, we discuss the potential therapeutic value of pharmaceutical inhibition of these essential pathways in combating liver cancer. Abstract Metabolic reprogramming and epigenetic changes have been characterized as hallmarks of liver cancer. Independently of etiology, oncogenic pathways as well as the availability of different energetic substrates critically influence cellular metabolism, and the resulting perturbations often cause aberrant epigenetic alterations, not only in cancer cells but also in the hepatic tumor microenvironment. Metabolic intermediates serve as crucial substrates for various epigenetic modulations, from post-translational modification of histones to DNA methylation. In turn, epigenetic changes can alter the expression of metabolic genes supporting on the one hand, the increased energetic demand of cancer cells and, on the other hand, influence the activity of tumor-associated immune cell populations. In this review, we will illustrate the most recent findings about metabolic reprogramming in liver cancer. We will focus on the metabolic changes characterizing the tumor microenvironment and on how these alterations impact on epigenetic mechanisms involved in the malignant progression. Furthermore, we will report our current knowledge about the influence of cancer-specific metabolites on epigenetic reprogramming of immune cells and we will highlight how this favors a tumor-permissive immune environment. Finally, we will review the current strategies to target metabolic and epigenetic pathways and their therapeutic potential in liver cancer, alone or in combinatorial approaches.

Background Hepatocellular carcinoma (HCC) is the most prevalent primary liver cancer, characterized by a poor prognosis. Many HCC patients are diagnosed at an advanced stage due to the lack of reliable prognostic biomarkers. G6PC1 (Glucose‐6‐Phosphatase Catalytic Subunit 1) is abnormally expressed in various cancers, including HCC. This study aimed to investigate the biomarker potential and biological functions of G6PC1 to elucidate its impact on HCC pathogenesis. Methods G6PC1 expression levels were assessed using TCGA and GEO datasets. Prognostic implications were explored through Kaplan-Meier survival analysis. Potential regulatory transcription factors (TFs) were identified using four prediction tools, and functional mechanisms were investigated via GO and KEGG enrichment analyses. Associations between G6PC1 and HCC metabolic reprogramming, as well as the tumor microenvironment were analyzed. Results G6PC1 exhibited low expression levels in HCC, which correlated with poor patient prognosis. HNF4A may act as a regulatory factor for G6PC1 in HCC. Functional analysis identified co-expressed genes associated with metabolism-related pathways. Furthermore, G6PC1 was implicated in metabolic reprogramming, immune infiltration, and immunotherapy response. Conclusion Low G6PC1 expression, associated with poor HCC prognosis, is a potential prognostic biomarker. Integrated multi-omics analyses underscore its clinical significance, involvement in metabolic reprogramming, and immunomodulatory functions, providing a foundation for further investigation into its prognostic potential and mechanistic contributions in HCC.

Background and aims Metabolic reprogramming has been found to be a typical feature of tumors. Hepatocellular carcinoma (HCC), a cancer with high morbidity and mortality, has been extensively studied for its metabolic reprogramming-related mechanisms. Our study aims to identify the hotspots and frontiers of metabolic reprogramming research in HCC and to provide guidance for future scientific research and decision-making in HCC metabolism. Methods Relevant studies on the metabolic reprogramming of HCC were derived from the Web of Science Core Collection (WoSCC) database up until November 2023. The bibliometrix tools in R were used for scientometric analysis and visualization. Results From 2011 to 2023, a total of 575 publications were obtained from WoSCC that met the established criteria. These publications involved 3,904 researchers and 948 organizations in 37 countries, with an average annual growth rate of 39.11% in research. These studies were published in 233 journals, with Cancers (n = 29) ranking first, followed by Frontiers in Oncology (n = 20) and International Journal of Molecular Sciences (n = 19). The top ten journals accounted for 26% of the 575 studies. The most prolific authors were Wang J (n = 14), Li Y (n = 12), and Liu J (n = 12). The country with the most publications is China, followed by the United States, Italy, and France. Fudan University had the largest percentage of research results with 15.48% (n = 89). Ally A’s paper in Cell has the most citations. A total of 1,204 keywords were analyzed, with the trend themes such as “glycolysis,” “tumor microenvironment,” “Warburg effect,” “mitochondria,” “hypoxia ,” etc. Co-occurrence network and cluster analysis revealed the relationships between keywords, authors, publications, and journals. Moreover, the close collaboration between countries in this field was elucidated. Conclusion This bibliometric and visual analysis delves into studies related to metabolic reprogramming in HCC between 2012 and 2023, elucidating the characteristics of research in this field, which has gradually moved away from single glycolipid metabolism studies to the integration of overall metabolism in the body, pointing out the trend of research topics, and the dynamics of the interaction between the tumor microenvironment and metabolic reprogramming will be the future direction of research, which provides blueprints and inspirations for HCC prevention and treatment programs to the researchers in this field. Systematic Review Registration: [https://www.bibliometrix.org].

Immune checkpoint inhibitors (ICIs), Ipilimumab, Nivolumab, Pembrolizumab and Atezolizumab, have been applied in anti-tumor therapy and demonstrated exciting performance compared to conventional treatments. However, the unsatisfactory response rates, high recurrence and adaptive resistance limit their benefits. Metabolic reprogramming appears to be one of the crucial barriers to immunotherapy. The deprivation of required nutrients and altered metabolites not only promote tumor progression but also confer dysfunction on immune cells in the tumor microenvironment (TME). Glycolysis plays a central role in metabolic reprogramming and immunoregulation in the TME, and many therapies targeting glycolysis have been developed, and their combinations with ICIs are in preclinical and clinical trials. Additional attention has been paid to the role of amino acids, lipids, nucleotides and mitochondrial biogenesis in metabolic reprogramming and clinical anti-tumor therapy. This review attempts to describe reprogramming metabolisms within tumor cells and immune cells, from the aspects of glycolysis, amino acid metabolism, lipid metabolism, nucleotide metabolism and mitochondrial biogenesis and their impact on immunity in the TME, as well as the significance of targeting metabolism in anti-tumor therapy, especially in combination with ICIs. In particular, we highlight the expression mechanism of programmed cell death (ligand) 1 [PD-(L)1] in tumor cells and immune cells under reprogramming metabolism, and discuss in detail the potential of targeting key metabolic pathways to break resistance and improve the efficacy of ICIs based on results from current preclinical and clinical trials. Besides, we draw out biomarkers of potential predictive value in ICIs treatment from a metabolic perspective.

Aim: Aberrant metabolism represents a hallmark feature of malignancies, which is crucial for facilitating adenosine triphosphate (ATP) production and biosynthesis of macromolecules that sustain cell proliferation, differentiation, and survival. In the context of tumorigenesis, fatty acids (FAs) have garnered substantial attention due to their dual role as secondary messengers and energy substrates. Notably, the pivotal role of FA metabolism in hepatocellular carcinoma (HCC) progression has been extensively explored. Therefore, this study aims to investigate the contributions of FA metabolism in the immunotherapy of HCC, which remain undefined. Methods: We analyzed messenger RNA expression and genetic alterations of regulators of FA metabolism from public HCC datasets. Based on their FA metabolism profiles, patients were classified into two distinct molecular subtypes: cluster A and cluster B. Using subtype-derived differentially expressed genes, we established an unsupervised FA_score algorithm. Immune infiltration analysis and prognostic screening of 2,484 immune genes were integrated to develop a risk model, ultimately classifying patients into four integrated subtypes: mixed index (MI)-1 to MI-4. Result: Cluster B exhibited significantly worse overall survival than cluster A. Higher FA_score correlated with shorter survival and increased infiltration of immunosuppressive cells. The MI-2 subgroup showed abundant CD4+ T cells, myeloid-derived suppressor cells, and regulatory T cells, indicating strong immunosuppression and poor prognosis, suggesting limited benefit from immunotherapy. Conclusion: We developed a novel classification system integrating FA metabolism and immune features. The MI-2 subtype is characterized by immunosuppression and poor outcomes, highlighting the clinical relevance of FA metabolic patterns in shaping the immune microenvironment and guiding personalized treatment in HCC.

Single-agent immune checkpoint inhibitors (ICIs) have been tested in patients with advanced hepatocellular carcinoma (HCC) showing an objective response rate of 15-20%, mostly without a significant overall survival (OS) benefit. Furthermore, approximately 30% of HCC shows intrinsic resistance to ICIs. In the absence of predictive biomarkers to identify patients likely to benefit most from immunotherapy, research has moved to exploring combinations with potential activity in broader patient populations. Basket trials, including cohorts of patients with HCC, and early phase studies tested the combination of ICIs with antiangiogenic agents as well as the combination of two different ICIs. The achieved promising results provided the rationale for the following phase 3 trials, which tested the combination of anti-PD-1/PD-L1 with bevacizumab, or tyrosine kinase inhibitors (TKIs), or anti-CTLA-4. Positive results from the IMbrave150 trial led to the practice-changing approval of atezolizumab-bevacizumab, the first regimen to demonstrate improved survival in the front-line setting, since the approval of sorafenib. More recently, the HIMALAYA trial demonstrated the superiority of durvalumab-tremelimumab (STRIDE regimen) over sorafenib, establishing a new first-line option. In contrast, inconsistent results have been achieved with combinations of ICIs and TKIs, with only one phase 3 trial showing an OS benefit. The rapidly evolving therapeutic landscape for patients with advanced HCC has left significant unmet needs to be addressed in future research. These include choice and sequencing of treatments, identification of biomarkers, combinations with locoregional therapies, and development of new immunotherapy agents. This review summarizes the scientific rationale and available clinical data for combination immunotherapy in advanced HCC.

Background Hepatocellular carcinoma (HCC) is the third leading cause of cancer-related death, worldwide. The predominant causative factor for HCC is hepatitis B virus (HBV) infection. We conducted a meta-analysis to estimate the efficacy and safety of PD-1/PD-L1 inhibitors combined with anti-angiogenic therapy for the first-line treatment of the unresectable HCC and to evaluate the benefits of different geographic regions and etiology stratifications. Methods Randomized clinical trials published up to 12th November 2022 were searched by online databases. Moreover, effects of hazard ratio (HR) for overall survival (OS) and progression-free survival (PFS) were extracted from included studies. Pooled odds ratio (OR) and 95% CI for objective response rate (ORR), disease control rate (DCR), and treatment-related adverse events (TRAEs) were calculated. Results A total of 3057 patients from five phase III randomized clinical trials were collected and reviewed for this meta-analysis. The pooled HR of OS (HR = 0.71; 95% CI: 0.60–0.85) and PFS (HR = 0.64; 95% CI: 0.53–0.77) demonstrated significantly better benefit in PD-1/PD-L1 inhibitors combination group than targeted monotherapy to treat unresectable HCC. In addition, combination therapy showed better ORR and DCR, with ORs of 3.29 (95% CI: 1.92–5.62) and 1.88 (95% CI: 1.35–2.61), respectively. The subgroup analysis indicated that PD-1/PD-L1 inhibitors combination therapy was significantly superior to anti-angiogenic monotherapy for HBV-related HCC in terms of OS (HR = 0.64; 95% CI: 0.55–0.74) and PFS (HR = 0.53; 95% CI:0.47–0.59), while there was no significant difference in patients with HCV (OS, HR = 0.81, p  = 0.1) or non-viral (OS, HR = 0.91, p  = 0.37; PFS, HR = 0.77, p  = 0.05). Conclusions Meta-analysis revealed for the first-time that PD-1/PD-L1 inhibitors combination therapy for unresectable HCC was associated with better clinical outcomes than anti-angiogenic monotherapy, especially for HBV infection and Asian population.

Hepatocellular carcinoma (HCC) is a malignancy with high morbidity and mortality but lacks effective treatments thus far. Although the emergence of immune checkpoint inhibitors in recent years has shed light on the treatment of HCC, a considerable number of patients are still unable to achieve durable and ideal clinical benefits. Therefore, refining the combination of immune checkpoint inhibitors (ICIs) to enhance the therapeutic effect has become a global research hotspot. Several histone deacetylase 2 inhibitors have shown advantages in ICIs in many solid cancers, except for HCC. Additionally, the latest evidence has shown that histone deacetylase 2 inhibition can regulate PD-L1 acetylation, thereby blocking the nuclear translocation of PD-L1 and consequently enhancing the efficacy of PD-1/PD-L1 inhibitors and improving anti-cancer immunity. Moreover, our team has recently discovered a novel HDAC2 inhibitor (HDAC2i), valetric acid (VA), that possesses great potential in HCC treatment as a monotherapy. Thus, a new combination strategy, combining HDAC2 inhibitors with ICIs, has emerged with significant development value. This perspective aims to ignite enthusiasm for exploring the application of ideal HDAC2 inhibitors with solid anti-tumor efficacy in combination with immunotherapy for HCC.

Antibodies targeting programmed cell death protein-1 (PD-1) or its ligand PD-L1 rescue T cells from exhausted status and revive immune response against cancer cells. Based on the immense success in clinical trials, ten α-PD-1 (nivolumab, pembrolizumab, cemiplimab, sintilimab, camrelizumab, toripalimab, tislelizumab, zimberelimab, prolgolimab, and dostarlimab) and three α-PD-L1 antibodies (atezolizumab, durvalumab, and avelumab) have been approved for various types of cancers. Nevertheless, the low response rate of α-PD-1/PD-L1 therapy remains to be resolved. For most cancer patients, PD-1/PD-L1 pathway is not the sole speed-limiting factor of antitumor immunity, and it is insufficient to motivate effective antitumor immune response by blocking PD-1/PD-L1 axis. It has been validated that some combination therapies, including α-PD-1/PD-L1 plus chemotherapy, radiotherapy, angiogenesis inhibitors, targeted therapy, other immune checkpoint inhibitors, agonists of the co-stimulatory molecule, stimulator of interferon genes agonists, fecal microbiota transplantation, epigenetic modulators, or metabolic modulators, have superior antitumor efficacies and higher response rates. Moreover, bifunctional or bispecific antibodies containing α-PD-1/PD-L1 moiety also elicited more potent antitumor activity. These combination strategies simultaneously boost multiple processes in cancer-immunity cycle, remove immunosuppressive brakes, and orchestrate an immunosupportive tumor microenvironment. In this review, we summarized the synergistic antitumor efficacies and mechanisms of α-PD-1/PD-L1 in combination with other therapies. Moreover, we focused on the advances of α-PD-1/PD-L1-based immunomodulatory strategies in clinical studies. Given the heterogeneity across patients and cancer types, individualized combination selection could improve the effects of α-PD-1/PD-L1-based immunomodulatory strategies and relieve treatment resistance.

Hepatocellular carcinoma (HCC) has a high prevalence and mortality rate worldwide. Sorafenib monotherapy has been the standard of first-line treatment for advanced HCC for a long time, but there are still many shortcomings. In recent years, with the deepening of research on tumor immune microenvironment, researchers have begun to explore new approaches in immunotherapy, and the introduction of immune checkpoint inhibitors has brought fundamental changes to the treatment of HCC. Programmed cell death protein 1 (PD-1) is an immune checkpoint molecule that plays an important role in down-regulating immune system function and promoting tolerance. Programmed cell death ligand 1 (PDL-1) is involved in tumor immune evasion by binding to PD-1, resulting in failure of treatment. Currently, immunotherapy targeting the PD-1/PD-L1 axis has achieved unprecedented success in HCC, but it also faces great challenges, with its low remission rate still to be solved. For most patients with HCC, the PD-1/PD-L1 pathway is not the only rate limiting factor of antitumor immunity, and blocking only the PD-1/PD-L1 axis is not enough to stimulate an effective antitumor immune response; thus, combination therapy may be a better option. In this study, changes in the immune microenvironment of HCC patients were reviewed to clarify the feasibility of anti-PD-1/PD-L1 therapy, and a series of monotherapy and combination therapy clinical trials were summarized to verify the safety and efficacy of this newly developed treatment in patients with advanced HCC. Furthermore, we focused on hyperprogressive disease and drug resistance to gain a better understanding of PD-1/PD-L1 blockade as a promising treatment.

ABSTRACT Introduction Underlying liver disease and the intrinsic chemoresistance have historically hampered the development of efficacious treatments in HCC. However, in the last few years, immunotherapy-based combinations have emerged as efficacious therapeutic strategy in this setting. This paper critically summarizes the recent therapeutic progress in the systemic treatment of HCC. Area covered This paper examines the preclinical rationale of the following combinations in HCC: dual checkpoint inhibitors, immune checkpoint inhibitors plus anti-angiogenic agents, and immune checkpoint inhibitors plus tyrosine kinase inhibitors. Results of recent clinical studies are presented, along with a brief overview of ongoing and future trials. Expert opinion The approval of atezolizumab plus bevacizumab and the positive results of the HIMALAYA trial have broadened the therapeutic scenario for advanced HCC, opening, at the same time, new challenges. First of all, predictive biomarkers to allocate patients to the best treatment are eagerly required; second, specific studies are urgently needed to define the use of new combinations in patients usually excluded from clinical trials, e.g. those with deranged liver function and HIV or transplant recipients. Finally, with new combinations being translated into earlier stages, profound changes are soon expected in the adjuvant and neoadjuvant setting.

BACKGROUND: Transarterial chemoembolization (TACE) is a main treatment for advanced hepatocellular carcinoma (HCC), but tumors often become resistant. Combining TACE programmed cell death (ligand) 1 [PD-(L)1] inhibitors and molecular targeted therapies (MTT) may improve outcomes, but its role in preventing TACE resistance requires further investigation. AIM: To compare if TACE plus PD-(L)1 inhibitors and MTT reduces TACE resistance and improves survival in advanced HCC compared to TACE alone. METHODS: We analyzed 721 patients: 532 received TACE only, and 72 received TACE with PD-(L)1 inhibitors and MTT. After matching patient characteristics, 144 patients (72 pairs) were compared. Tumor progression after 3 treatment cycles was measured. RESULTS: = 0.008). These findings underscore the efficacy of combination therapy in enhancing therapeutic outcomes in advanced HCC. CONCLUSION: Adding immunotherapy and targeted drugs to TACE significantly reduces treatment resistance and improves survival in advanced liver cancer, suggesting it may become a new standard treatment.

Objective To systematically evaluate the safety and efficacy of PD-1/PD-L1 inhibitor-based immunotherapy (hereafter referred to as “combination immunotherapy”) compared with that of sorafenib in the treatment of hepatocellular carcinoma (HCC). Methods Databases such as PubMed, Embase, and the Cochrane Library were searched from the date of their establishment to September 2023 to identify randomized controlled trials (RCTs) of combination immunotherapy versus sorafenib for the treatment of advanced HCC. Two reviewers independently evaluated the quality of the included studies, extracted the data, and cross-checked the information. The meta-analysis was performed using RevMan 5.3 software. Results A total of 5 RCTs were included. The results of the meta-analysis showed the following: (1) Effectiveness. Compared to sorafenib, combination immunotherapy significantly improved overall survival (OS, HR = 0.69, 95% CI: 0.58 ~ 0.82, p < 0.01) and progression-free survival (PFS, HR = 0.62, 95% CI: 0.50 ~ 0.78, p < 0.001) in patients with advanced HCC. (2) Safety. Both groups had comparatively high incidences of adverse events (AEs), but the difference in any treatment-related adverse events was not significant between the two arms (OR = 0.98, 95% CI: 0.95 ~ 1.02, p = 0.34). The difference in the incidence of grade 1–2 adverse reactions was statistically significant (OR = 0.66, 95% CI = 0.49–0.90, p = 0.001). There were no differences in grade 3/4 TRAEs or grade 5 TRAEs (OR = 1.46, 95% CI = 0.78 ~ 2.71, p = 0.24; OR = 1.08, 95% CI = 0.73 ~ 1.58, p = 0.71). Conclusion Combined immunotherapy can significantly prolong the OS and PFS of patients with advanced HCC without increasing the incidence of adverse effects in terms of safety, but the incidence of AEs in different systems is different.

Purpose To evaluate the efficacy and safety of TACE combined with regorafenib plus PD-1 inhibitor as a second-line therapy for hepatocellular carcinoma after sorafenib resistance. Materials and Methods The clinical data of 76 patients with hepatocellular carcinoma who were drug-resistant to sorafenib from September 2018 to May 2022 in the tumor intervention department were collected. Among them, 35 patients used TACE combined with regorafenib plus PD-1 inhibitor (TACE-R-P) as second-line treatment, and the remaining 41 patients used TACE combined with regorafenib (TACE-R) as second-line treatment. The mRECIST (modified Response Evaluation Criteria in Solid Tumors) standard was used to evaluate the therapeutic effect. The progression-free survival (PFS) and overall survival (OS) of the two groups were compared. Blood samples were collected before and after treatment to detect the changes in biochemical indicators, and the adverse events (AEs) related to treatment were recorded. Results A total of 76 patients were included in the study, including 35 patients receiving TACE-R-P treatment and 41 patients receiving TACE-R treatment. Patients in the TACE-R-P group had longer median OS (19.7months vs 15.2months, HR:0.7716, 95% CI:0.4767–1.2490, P=0.03), longer median PFS (6.3months vs 3.8months, HR:0.6032, 95% CI:0.3727–0.9763, P=0.0029), higher objective response rate (37.14% vs 19.51%, P=0.001) and higher disease control rate (71.43% vs 48.78%, P=0.001) than those in the TACE-R group. Multivariate analysis showed that Child–Pugh grade (B/A; HR=1.283, 95% CI: 0.623–1.707, P=0.014), PVTT (Yes/No, HR=1.455, 95% CI: 0.977–2.038, P=0.018), extrahepatic metastasis (Yes/No, HR=1.766, 95% CI: 1.135–2.302, P=0.022) and treatment option (TACE-R/TACE-R-P, HR=1.930, 95% CI: 1.461–2.850, P=0.017) were independent prognostic factors for OS. There was no significant difference in the incidence and severity of AEs between the two groups. Conclusion TACE-R-P treatment can be more effective than TACE-R treatment for HCC after sorafenib resistance and can be given priority as a second-line treatment for HCC.

A successful phase III trial for the combination of atezolizumab and bevacizumab (the IMbrave150 trial) in advanced hepatocellular carcinoma has recently been reported. This is groundbreaking because nivolumab and pembrolizumab, both programmed cell death-1 (PD-1) antibodies, have failed to show efficacy as first- and second-line therapeutics, respectively, in phase III clinical trials. Immunotherapy with a combination of atezolizumab and bevacizumab resulted in better survival than treatment with sorafenib for the first time since sorafenib was approved in 2007. The high efficacy of the combination of PD-1/programmed death ligand 1 (PD-L1) and vascular endothelial growth factor (VEGF) antibodies is not only due to their additive effects on tumor growth, but also to their reprogramming of the immunosuppressive microenvironment into an immunostimulatory microenvironment. These results were confirmed in a phase Ib trial that showed significantly longer progression-free survival in the atezolizumab plus bevacizumab group than in patients that received atezolizumab alone. These results demonstrate that immunotherapy with a combination of PD-1/PD-L1 and VEGF inhibitors is effective and may result in a reprogramming of the tumor microenvironment. The results of an ongoing phase III trial of a PD-1 antibody in combination with the VEGF receptor tyrosine kinase inhibitor (TKI) are highly anticipated.

… the therapeutic benefit and overcome drug resistance. … suggested that the combination therapy of PD-1 blockade and … a possible candidate for clinical trials of advanced HCC, although …

Simple Summary The cytotoxic T cell response against hepatocellular carcinoma antigens is exhausted and fails in its task of deleting tumoral cells. These cells are featured by the expression of negative immune checkpoints that can be modulated to restore T cell function. The blockade of the PD-1/PD-L1 pathway has shown promising results in rescuing hepatocellular carcinoma-specific CD8 T cells but only a reduced group of cases is sensitive to this treatment and the effect is usually temporary. Therefore, new anti-PD-1 based combinatory strategies are underway to increase the response by adding the effect of blocking neo-angiogenesis and other negative immune checkpoints, boosting positive immune checkpoints, blocking suppressive cytokines, or inducing the expression of tumoral neoantigens. The restoration of T cell responses with these anti-PD-1 based combinatory therapies will change the outcome of advanced hepatocellular carcinoma. Abstract Thirty to fifty percent of hepatocellular carcinomas (HCC) display an immune class genetic signature. In this type of tumor, HCC-specific CD8 T cells carry out a key role in HCC control. Those potential reactive HCC-specific CD8 T cells recognize either HCC immunogenic neoantigens or aberrantly expressed host’s antigens, but they become progressively exhausted or deleted. These cells express the negative immunoregulatory checkpoint programmed cell death protein 1 (PD-1) which impairs T cell receptor signaling by blocking the CD28 positive co-stimulatory signal. The pool of CD8 cells sensitive to anti-PD-1/PD-L1 treatment is the PD-1dim memory-like precursor pool that gives rise to the effector subset involved in HCC control. Due to the epigenetic imprints that are transmitted to the next generation, the effect of PD-1 blockade is transient, and repeated treatments lead to tumor resistance. During long-lasting disease, besides the TCR signaling impairment, T cells develop other failures that should be also set-up to increase T cell reactivity. Therefore, several PD-1 blockade-based combinatory therapies are currently under investigation such as adding antiangiogenics, anti-TGFβ1, blockade of other negative immune checkpoints, or increasing HCC antigen presentation. The effect of these combinations on CD8+ T cells is discussed in this review.

ABSTRACT Introduction Hepatocellular carcinoma (HCC) is one of the leading causes of cancer-related death worldwide with most of patients diagnosed at advanced stage. Thus, systemic therapy remains a cornerstone of treatment. In recent years, immunotherapy has changed therapeutic scenario, being investigated also in combination with anti-vascular endothelial growth factor (VEGF) agents. This approach has demonstrated safety and efficacy in several trials, paving the way for their investigation in earlier disease stages and in different settings. Areas covered A structured literature review was conducted using PubMed and ClinicalTrials.gov to identify published evidence supporting safety and efficacy of anti-programmed death-1/programmed death-ligand-1(PD-1/PD-L1) and anti-VEGF agents in different therapeutic settings and identifying ongoing clinical trials and key research directions. Expert opinion Combination of anti-PD-1/PD-L1 and anti-VEGF agents has demonstrated safety and efficacy as a first-line treatment for advanced HCC, supported by results from phase 3 trials. These results suggest that further investigation is warranted to optimize first-line efficacy, second-line choice, and potential application in earlier disease stages. The clinical benefit of anti-PD-1/PD-L1 and anti-VEGF agents have also opened the door to a new clinical paradigm, where transitioning from systemic therapy to locoregional therapies, resection or even liver transplantation could be a feasible treatment strategy.

Background The use of immune checkpoint inhibitors (ICIs) in treating hepatocellular carcinoma (HCC) has grown significantly. However, the therapeutic benefits of ICIs alone are notably modest. This meta-analysis assesses the efficacy and safety of using PD-1/PD-L1 inhibitors in conjunction with tyrosine kinase inhibitors (TKIs) for patients with advanced or unresectable HCC. Methods An extensive search of the literature was performed using databases such as PubMed, Web of Science, Embase, and the Cochrane Library, capturing randomized controlled trials (RCTs) until 16 October 2024. Efficacy was measured by progression-free survival (PFS), overall survival (OS), objective response rate (ORR), and disease control rate (DCR). Safety was gauged through the occurrence of treatment-related adverse events (TRAEs). Hazard ratios (HRs) for PFS and OS, along with risk ratios (RRs) for ORR, DCR, and TRAEs, were calculated, each with 95% confidence intervals (CIs). Heterogeneity among studies was quantified using Cochran’s Q test, I2 statistics, and 95% prediction intervals (PIs). Results This analysis incorporated 4 studies with a total of 2,174 patients. Treatment regimens combining PD-1/PD-L1 inhibitors with TKIs significantly improved PFS (HR = 0.694, 95% CI: 0.527–0.914; 95% PI: 0.228–2.114) and ORR (RR = 2.303, 95% CI: 1.360–3.902; 95% PI: 0.408–12.991) compared with first-line monotherapy or TKI monotherapy in the overall population. Subgroup analysis indicated that the improvements in PFS and OS were particularly significant among patients of Asian descent or those with hepatitis B virus (HBV) infection (all p < 0.05). While the occurrence of any grade TRAEs did not differ significantly between the two groups (RR = 1.016, 95% CI: 0.996–1.036; 95% PI: 0.941–1.097), the incidence of serious (RR = 2.068, 95% CI: 1.328–3.222; 95% PI: 0.487–8.776) and grade ≥3 TRAEs (RR = 1.287, 95% CI: 1.020–1.624; 95% PI: 0.574–2.883) increased in patients treated with the combination of PD-1/PD-L1 inhibitors and TKIs. Conclusion This study revealed that combining PD-1/PD-L1 inhibitors with TKIs in the treatment of advanced or unresectable HCC leads to superior clinical outcomes compared to first-line monotherapy or TKIs alone, particularly in patients with HBV infection and those of Asian descent. Clinicians are advised to be vigilant regarding the potential for TRAEs in clinical settings.

Simple Summary Immune checkpoint inhibitors (ICIs) had been explored extensively in patients affected by unresectable hepatocellular carcinoma. These agents were expected to be the keystones of the disease’s first-line treatment because they were theoretically able to revert the immune suppressive tumor microenvironment of the cancerous liver, and because of their manageable safety profile. However, when used as monotherapies, they showed important activity and efficacy limitations. In this mini-review, we summarize the characteristics of the different ICIs-based regimens which constitute the present gold standard of first-line treatment, then, moving from their shortcomings, we discuss the rationale supporting the strategies currently under investigation: systemic triplets and new paradigms of immune-therapeutic agents such as CAR-T and vaccines. Abstract Immune checkpoint inhibitors (ICIs) are a key component of different stages of hepatocellular carcinoma (HCC) treatment, particularly in the first line of treatment. A lesson on the primary resistance which hampers their efficacy and activity was learned from the failure of the trials which tested them as first-line mono-therapies. Despite the combination of anti-PD(L)1 agents with anti-VEGF, anti CTLA4, or TKIs demonstrating relevant improvements in efficacy, the “doublets strategy” still shows room for improvement, due to a limited overall survival benefit and a high rate of progressive disease as best response. In this review, we discuss the results from the currently tested doublet strategies (i.e., atezolizumab+bevacizumab, durvalumab+tremelimumab with a mention to the newly presented ICIs/TKIs combinations), which highlight the need for therapeutic improvement. Furthermore, we examine the rationale and provide an overview of the ongoing trials testing the treatment intensification strategy with triplet drugs: anti-PD1+anti-CTLA4+anti-VEGF/TKIs and anti-PD1+anti-VEGF+alternative immunity targets. Lastly, we report on the alternative strategy to integrate ICIs into the new paradigm of immune therapeutics constituted by CAR-T and anti-cancer vaccines. This review provides up-to-date knowledge of ongoing clinical trials of the aforementioned strategies and critical insight into their mechanistic premises.

Importance Combination therapies of anti-PD-1 and anti-angiogenesis regimens are emerging rapidly and exhibit more promising anti-tumor efficacy for advanced hepatocellular carcinoma (HCC), and consistently it is the hotspot in clinical studies. Objective To elaborate several issues which are warranted further consideration as more regimens are being investigated in combination therapies. Evidence Review We searched PubMed, MEDLINE, Cochrane Library and Google Scholar by 2021 February for publications on combination therapies for HCC. Findings Several clinical issues are worth reconsidering, such as the evaluation on appropriate primary endpoints in phase III clinical trials as for different practical problems, the translation of surrogate endpoint objective response rate (ORR) benefits into overall survival (OS) benefits, and whether conversion surgery contributes to initial expectations of long-term survival or not. New concepts in novel immunotherapy and targeted therapy in combination with loco-regional therapies may improve overall survival for HCC. Conclusions and Relevance for Reviews Comprehensive understanding of the mechanism of immunotherapy and targeted therapy contributes to better prognosis of advanced HCC and more explorative combination therapies are needed.

Background Immune cell infiltration in the tumor microenvironment (TME) affects tumor initiation, patients’ prognosis and immunotherapy strategies. However, their roles and interactions with genomics and molecular processes in hepatocellular carcinoma (HCC) still have not been systematically evaluated. Methods We performed unsupervised clustering of total 1000 HCC samples including discovery and validation group from available public datasets. Immune heterogeneity of each subtype was explored by multi-dimension analysis. And a support vector machine (SVM) model based on multi-omics signatures was trained and tested. Finally, we performed immunohistochemistry to verify the immune role of signatures. Results We defined three immune subtypes in HCC, with diverse clinical, molecular, and genomic characteristics. Cluster1 had worse prognosis, better anti-tumor characteristics and highest immune scores, but also accompanied by immunosuppression and T cell dysfunction. Meanwhile, a better anti-PD1/CTLA4 immunotherapeutic response was predicted in cluster1. Cluster2 was enriched in TAM-M2 and stromal cells, indicating immunosuppression. Cluster3, with better prognosis, had lowest CD8 T cell but highest immune resting cells. Further, based on genomic signatures, we developed an SVM classifier to identify the patient’s immunological status, which was divided into Type A and Type B, in which Type A had poorer prognosis, higher T cell dysfunction despite higher T cell infiltration, and had better immunotherapeutic response. At the same time, MMP9 may be a potential predictor of the immune characteristics and immunotherapeutic response in HCC. Conclusions Our work demonstrated 3 immune clusters with different features. More importantly, multi-omics signatures, such as MMP9 was identified based on three clusters to help us recognize patients with different prognosis and responses to immunotherapy in HCC. This study could further reveal the immune status of HCC and provide potential predictors for immune checkpoint treatment response.

Tumor treatment has undergone revolutionary changes with the development of immunotherapy, especially immune checkpoint inhibitors. Because not all patients respond positively to immune therapeutic agents, and severe immune-related adverse events (irAEs) are frequently observed, the development of the biomarkers evaluating the response of a patient is key for the application of immunotherapy in a wider range. Recently, various multi-omics features measured by high-throughput technologies, such as tumor mutation burden (TMB), gene expression profiles, and DNA methylation profiles, have been proved to be sensitive and accurate predictors of the response to immunotherapy. A large number of predictive models based on these features, utilizing traditional machine learning or deep learning frameworks, have also been proposed. In this review, we aim to cover recent advances in predicting tumor immunotherapy response using multi-omics features. These include new measurements, research cohorts, data sources, and predictive models. Key findings emphasize the importance of TMB, neoantigens, MSI, and mutational signatures in predicting ICI responses. The integration of bulk and single-cell RNA sequencing has enhanced our understanding of the tumor immune microenvironment and enabled the identification of predictive biomarkers like PD-L1 and IFN-γ signatures. Public datasets and machine learning models have also improved predictive tools. However, challenges remain, such as the need for large and diverse clinical datasets, standardization of multi-omics data, and model interpretability. Future research will require collaboration among researchers, clinicians, and data scientists to address these issues and enhance cancer immunotherapy precision.

Hepatocellular carcinoma (HCC) represents a significant global health challenge, with a complex etiology and limited treatment options. The integration of multi-omics technologies, including genomics, transcriptomics, proteomics, and metabolomics, has revolutionized our understanding of HCC, offering novel insights into its molecular underpinnings. This comprehensive review synthesizes the current knowledge on the application of multi-omics in HCC, highlighting its role in disease classification, early detection, and the development of targeted therapies. We discuss the identification of key driver mutations and single nucleotide polymorphisms (SNPs) that enhance risk prediction models, with implications for personalized medicine. The multi-omics approach has facilitated the discovery of distinct HCC subtypes, each with unique molecular signatures and tumor microenvironments (TME), which are critical for predicting prognosis and guiding treatment strategies. Furthermore, we explore the implications of these findings for precision medicine, emphasizing the potential of biomarker identification and targeted therapies, including immune checkpoint blockade (ICB). The review concludes by underscoring the transformative impact of multi-omics on HCC research and clinical practice, heralding a new era of personalized medicine with the promise of improved patient outcomes.

Background: Although current immunotherapies have achieved some successes for hepatocellular carcinoma (HCC) patients, their benefits are limited for most HCC patients. Therefore, the identification of biomarkers for promoting immunotherapeutic responses in HCC is urgently needed. Methods: Using the TCGA HCC cohort, we investigated correlations of various molecular features with antitumor immune signatures (CD8+ T cell infiltration and cytolytic activity) and an immunosuppressive signature (PD-L1 expression) in HCC. These molecular features included mRNAs, microRNAs (miRNAs), long non-coding RNAs (lncRNAs), proteins, and pathways. Results: We found that the mutations of several oncogenes and tumor suppressor genes significantly correlated with reduced antitumor immune signatures, including TTN, CTNNB1, RB1, ZFHX4, and TP53. It indicates that these genes’ mutations may inhibit antitumor immune responses in HCC. Four proteins (Syk, Lck, STAT5, and Caspase-7) had significant positive expression correlations with CD8+ T cell enrichment, cytolytic activity, and PD-L1 expression in HCC. It suggests that these proteins’ expression could be useful biomarkers for the response to immune checkpoint inhibitors Similiarly, we identified other types of biomarkers potentially useful for predicting the response to ICIs, including miRNAs (hsa-miR-511-5p, 150-3p, 342-3p, 181a-3p, 625-5p, 4772-3p, 155-3p, 142-5p, 142-3p, 155-5p, 625-3p, 1976, 7702), many lncRNAs, and pathways (apoptosis, cytokine-cytokine receptor interaction, Jak-STAT signaling, MAPK signaling, PI3K-AKT signaling, HIF-1 signaling, ECM receptor interaction, focal adhesion, and estrogen signaling). Further, tumor mutation burden showed no significant correlation with antitumor immunity, while tumor aneuploidy levels showed a significant negative correlation with antitumor immunity. Conclusion: The molecular features significantly associated with HCC immunity could be predictive biomarkers for immunotherapeutic responses in HCC patients. They could also be potential intervention targets for boosting antitumor immunity and immunotherapeutic responses in HCC.

Background/Aims Hepatocellular carcinoma (HCC) exhibits substantial morphological and biological heterogeneity. Clinical and molecular relevance of the infiltrative subtype remains poorly defined in the context of cancer immunotherapy. We aimed to evaluate the prognostic impact and molecular features of infiltrative HCC in patients treated with first-line atezolizumab plus bevacizumab (Ate/Bev). Methods We included 307 patients with advanced HCC treated with Ate/Bev and classified them into four gross morphological types based on imaging. Multi-omics profiling was conducted on tumor samples. Type IV infiltrative signature was derived and externally validated using five independent HCC cohorts, including IMbrave150. Results Infiltrative morphology, encompassing pure and mixed forms, was present in 42.7% of advanced HCC and associated with advanced disease features and compromised liver function. Patients with type IV infiltrative HCC showed lowest objective response rate (14.6%) and worst progression-free (median, 2.8 months) and overall survival (median, 7.1 months). Infiltrative morphology remained an independent predictor of poor outcomes after multivariable adjustment for confounders, including intrahepatic tumor extent. Genomic profiling revealed enriched TP53 and ATM loss-of-function mutations in type IV infiltrative HCC. Transcriptomic and proteomic analyses identified consistent activation of tumor proliferation, epithelial-mesenchymal transition, TGF-β signaling, and immunosuppressive pathways in type IV infiltrative HCC. Type IV infiltrative signature was significantly associated with poor survival across external datasets and retained independent prognostic value. Conclusions Infiltrative HCC is a clinically aggressive and molecularly distinct subtype of advanced HCC. Morphological classification and type IV infiltrative signatures may guide risk stratification and therapeutic decision-making in advanced HCC treated with immunotherapy.

Objective To explore the role of transketolase (TKT) in the immunotherapy and prognosis of hepatocellular carcinoma (HCC). Materials and methods TKT expression across various cancers and its associations with tumor immunity and prognosis were analyzed using nomogram models. A multi-omics approach was employed, including bulk RNA-seq analysis, methylation profiling, single-cell analysis, and spatial transcriptomics. Experimental methods included RT-qPCR, siRNA transfection, luciferase reporter assay, and chromatin immunoprecipitation. Results TKT was significantly upregulated in multiple cancers and correlated with immune cell infiltration, particularly in HCC. Elevated TKT expression was associated with poor overall survival (OS) in HCC and was an independent prognostic factor (p < 0.05). Drug sensitivity analysis suggested that higher TKT expression was associated with reduced sensitivity to several chemotherapeutic agents, including sorafenib (p < 0.01). Furthermore, hypermethylation of the TKT promoter and low TKT expression were linked to improved OS in HCC (log-rank test p = 0.005). Single-cell analysis revealed that TKT was predominantly expressed in the monocyte/macrophage cluster associated with HCC, and pseudo-time series analysis highlighted TKT’s role in cell differentiation within this cluster. Spatial transcriptomics confirmed the close association between TKT and macrophage distribution in HCC. Moreover, STAT3 was found to directly regulate TKT expression by binding to its promoter region. Conclusion Our findings suggest that TKT may play a role in tumor immunity and prognosis in HCC. Although these results provide insights into the potential involvement of TKT in immune cell infiltration and survival outcomes, further studies are required to fully elucidate its role in immunotherapy.

The neural precursor cell expressed developmentally down-regulated protein 1 (NEDD1) is implicated in tumorigenesis, but its role in hepatocellular carcinoma (HCC) remains unclear. This study aims to explore the oncogenic role, regulatory mechanisms, and tumor microenvironment interactions of NEDD1 in HCC. Multi-omics analyses were performed using public datasets (TCGA, GEO) and in-house clinical samples. These included expression and survival analysis, epigenetic (DNA methylation) and post-translational (phosphorylation) profiling, functional pathway enrichment, and drug sensitivity prediction. Functional validation was conducted via NEDD1 knockdown in HCC cells and a subcutaneous xenograft model. The co-expression and spatial distribution of NEDD1 and its predicted partner MZT2B were investigated using single-cell (GSE140228) and spatial transcriptomic (HRA000437) datasets. NEDD1 was significantly overexpressed in HCC tissues and correlated with poor prognosis. Its overexpression was potentially linked to promoter hypomethylation and aberrant phosphorylation. NEDD1 knockdown suppressed HCC cell proliferation, migration, and tumor growth in vivo. Notably, NEDD1 expression positively correlated with immune checkpoint molecules (PD-1, CTLA-4), and low NEDD1 expression was associated with better predicted response to immunotherapy. Single-cell and spatial transcriptomics revealed that NEDD1 and MZT2B co-expression was highly enriched in specific macrophage subsets (e.g., APOE+) and exhibited cell context-dependent heterogeneity, suggesting they may constitute a dynamic functional module within the HCC microenvironment. This multi-omics study suggests NEDD1 as a potential prognostic biomarker and therapeutic target in HCC. We propose a novel model wherein the NEDD1-MZT2B module may operate in both tumor cells and immunosuppressive macrophages, potentially influencing disease progression and immunotherapy response. Supplementary Information The online version contains supplementary material available at 10.1038/s41598-026-42505-z.

Background Hepatocellular carcinoma (HCC) is the most prevalent form of liver cancer, characterized by elevated mortality rates and heterogeneity. Despite advancements in treatment, the development of personalized therapeutic strategies for HCC remains a substantial challenge due to the intricate molecular characteristics of the disease. A multi-omics approach has the potential to offer more profound insights into HCC subtypes and enhance patient stratification for personalized treatments. Methods A comprehensive data set comprising clinical, transcriptomic, genomic and epigenomic information from HCC patients was retrieved from the TCGA, ICGC, GEO and CPTAC databases. To identify distinct molecular subtypes, a multi-omics data integration approach was employed, utilizing 10 distinct clustering algorithms. Survival analysis, immune infiltration profiling and drug sensitivity predictions were then used to evaluate the prognostic significance and therapeutic responses of these subtypes. Furthermore, machine learning models were employed to develop the artificial intelligence-derived risk score (AIDRS) with the aim of predicting patient outcomes and guiding personalized therapy. In vitro and vivo experiments were conducted to assess the role of CEP55 in tumor progression. Results The present study identified two distinct HCC subtypes (CS1 and CS2, respectively), each exhibiting different clinical outcomes and molecular characteristics. CS1 was associated with better overall survival, while CS2 exhibited higher mutation burden and immune suppression. The AIDRS, constructed using a multi-step machine learning approach, effectively predicted patient prognosis across multiple cohorts. High AIDRS score correlated with poor prognosis and a limited response to immunotherapy. Furthermore, the study identified CEP55 as a potential therapeutic target, as it was found to be overexpressed in CS2 and associated with poorer outcomes. In vitro experiments confirmed that CEP55 knockdown reduced HCC cell proliferation, migration, and invasion. Moreover, in xenograft models, CEP55 knockdown significantly reduced tumor growth and proliferation. Conclusions The integration of multi-omics data has been demonstrated to provide a comprehensive understanding of HCC subtypes, thus enhancing the prediction of prognosis and guiding personalized treatment strategies. The development of the AIDRS offers a robust tool for risk stratification, while CEP55 has emerged as a promising target for therapeutic intervention in HCC.

Background Hepatocellular carcinoma (HCC) is a highly heterogeneous tumor, and the development of accurate predictive models for prognosis and drug sensitivity remains challenging. Methods We integrated laboratory data and public cohorts to conduct a multi-omics analysis of HCC, which included bulk RNA sequencing, proteomic analysis, single-cell RNA sequencing (scRNA-seq), spatial transcriptomics sequencing (ST-seq), and genome sequencing. We constructed a tumor purity (TP) and tumor microenvironment (TME) prognostic risk model. Proteomic analysis validated the TP-TME-related signatures. Joint analysis of scRNA-seq and ST-seq revealed characteristic clusters associated with TP high-risk subtypes, and immunohistochemistry confirmed the expression of key genes. We conducted functional enrichment analysis, transcription factor activity inference, cell-cell interaction, drug efficacy analysis, and mutation information analysis to identify a novel subtype of HCC. Results Our analyses constructed a robust HCC prognostic risk prediction model. The patients with TP-TME high-risk subtypes predominantly exhibit hypoxia and activation of the Wnt/beta-catenin, Notch, and TGF-beta signaling pathways. Furthermore, we identified a novel subtype, XPO1+Epithelial. This subtype expresses signatures of the TP risk subtype and aligns with the biological behavior of high-risk patients. Additional analyses revealed that XPO1+Epithelial is influenced primarily by fibroblasts via ligand-receptor interactions, such as FN1-(ITGAV+ITGB1), and constitute a significant component of the TP-TME subtype. Moreover, XPO1+Epithelial interact with monocytes/macrophages, T/NK cells, and endothelial cells through ligand-receptor pairs, including MIF-(CD74+CXCR4), MIF-(CD74+CD44), and VEGFA-VEGFR1R2, respectively, thereby promoting the recruitment of immune-suppressive cells and angiogenesis. The ST-seq cohort treated with Tyrosine Kinase Inhibitors (TKIs) and Programmed Cell Death Protein 1 (PD-1) presented elevated levels of TP and TME risk subtype signature genes, as well as XPO1+Epithelial, T-cell, and endothelial cell infiltration in the treatment response group. Drug sensitivity analyses indicated that TP-TME high-risk subtypes, including sorafenib and pembrolizumab, were associated with sensitivity to multiple drugs. Further exploratory analyses revealed that CTLA4, PDCD1, and the cancer antigens MSLN, MUC1, EPCAM, and PROM1 presented significantly increase expression levels in the high-risk subtype group. Conclusions This study constructed a robust prognostic model for HCC and identified novel subgroups at the single-cell level, potentially assisting in the assessment of prognostic risk for HCC patients and facilitating personalized drug therapy.

Hepatocellular carcinoma (HCC) was the sixth common malignancies characteristic with highly aggressive in the world. It was well established that tumor mutation burden (TMB) act as indicator of immunotherapeutic responsiveness in various tumors. However, the role of TMB in tumor immune microenvironment (TIME) is still obscure. The mutation data was analyzed by employing “maftools” package. Weighted gene co-expression network analysis (WGCNA) was implemented to determine candidate module and significant genes correlated with TMB value. Differential analysis was performed between different level of TMB subgroups employing R package “limma”. Gene ontology (GO) enrichment analysis was implemented with “clusterProfiler”, “enrichplot” and “ggplot2” packages. Then risk score signature was developed by systematical bioinformatics analyses. K-M survival curves and receiver operating characteristic (ROC) plot were further analyzed for prognostic validity. To depict comprehensive context of TIME, XCELL, TIMER, QUANTISEQ, MCPcounter, EPIC, CIBERSORT, and CIBERSORT-ABS algorithm were employed. Additionally, the potential role of risk score on immune checkpoint blockade (ICB) immunotherapy was further explored. The quantitative real-time polymerase chain reaction was performed to detect expression of HTRA3. TMB value was positively correlated with older age, male gender and early T status. A total of 75 intersection genes between TMB-related genes and differentially expressed genes (DEGs) were screened and enriched in extracellular matrix-relevant pathways. Risk score based on three hub genes significantly affected overall survival (OS) time, infiltration of immune cells, and ICB-related hub targets. The prognostic performance of risks score was validated in the external testing group. Risk-clinical nomogram was constructed for clinical application. HTRA3 was demonstrated to be a prognostic factor in HCC in further exploration. Finally, mutation of TP53 was correlated with risk score and do not interfere with risk score-based prognostic prediction. Collectively, a comprehensive analysis of TMB might provide novel insights into mutation-driven mechanism of tumorigenesis further contribute to tailored immunotherapy and prognosis prediction of HCC.

NAD(P)H dehydrogenase quinone 1 (NQO1) is overexpressed in various cancers and is strongly associated with an immunosuppressive microenvironment and poor prognosis. In this study, we explored the role of NQO1 in the microenvironment, prognosis and immunotherapy of Hepatocellular carcinoma (HCC) using multi-omics analysis and machine learning. The results revealed that NQO1 was significantly overexpressed in HCC cells. NQO1+HCC cells were correlated with poor prognosis and facilitated tumor-associated macrophages (TAMs) polarization to M2 macrophages. We identified core NQO1-related genes (NRGs) and developed the NRGs-related risk-scores in hepatocellular carcinoma (NRSHC). The comprehensive nomogram integrating NRSHC, age, and pathological tumor-node-metastasis (pTNM) Stage achieved an area under the curve (AUC) above 0.7, demonstrating its accuracy in predicting survival outcomes and immunotherapy responses of HCC patients. High-risk patients exhibited worse prognoses but greater sensitivity to immunotherapy. Additionally, a web-based prediction tool was designed to enhance clinical utility. In conclusion, NQO1 may play a critical role in M2 polarization and accelerates HCC progression. The NRSHC model and accompanying tools offer valuable insights for personalized HCC treatment.

Background: Nonalcoholic steatohepatitis (NASH)-driven hepatocellular carcinoma (HCC) is becoming a major health-related problem. The exploration of NASH-related prognostic biomarkers and therapeutic targets is necessary. Methods: Data were downloaded from the GEO database. The “glmnet” package was used to identify differentially expressed genes (DEGs). The prognostic model was constructed by the univariate Cox and LASSO regression analyses. Validation of the expression and prognosis by immunohistochemistry (IHC) in vitro. Drug sensitivity and immune cell infiltration were analyzed by CTR-DB and ImmuCellAI. Results: We constructed a prognostic model that identified the NASH-related gene set (DLAT, IDH3B, and MAP3K4), which was validated in a real-world cohort. Next, seven prognostic transcription factors (TFs) were identified. The prognostic ceRNA network included three mRNAs, four miRNAs, and seven lncRNAs. Finally, we found that the gene set was associated with drug response which was validated in six clinical trial cohorts. Moreover, the expression level of the gene set was inversely correlated with CD8 T cell infiltration in HCC. Conclusions: We established a NASH-related prognostic model. Upstream transcriptome analysis and the ceRNA network provided clues for mechanism exploration. The mutant profile, drug sensitivity, and immune infiltration analysis further guided precise diagnosis and treatment strategies.

Background: Hepatocellular carcinoma (HCC) is a highly malignant tumor with high incidence and mortality rates. Aging-related genes are closely related to the occurrence and development of cancer. Therefore, it is of great significance to evaluate the prognosis of HCC patients by constructing a model based on aging-related genes. Method: Non-negative matrix factorization (NMF) clustering analysis was used to cluster the samples. The correlation between the risk score and immune cells, immune checkpoints, and Mismatch Repair (MMR) was evaluated through Spearman correlation test. Real Time Quantitative PCR (RT-qPCR) and immunohistochemistry were used to validate the expression levels of key genes in tissue and cells for the constructed model. Result: By performing NMF clustering, we were able to effectively group the liver cancer samples into two distinct clusters. Considering the potential correlation between aging-related genes and the prognosis of liver cancer patients, we used aging-related genes to construct a prognostic model. Spearman correlation analysis showed that the model risk score was closely related to MMR and immune checkpoint expression. Drug sensitivity analysis also provided guidance for the clinical use of chemotherapy drugs. RT-qPCR showed that TFDP1, NDRG1, and FXR1 were expressed at higher levels in different liver cancer cell lines compared to normal liver cells. Conclusion: In summary, we have developed an aging-related model to predict the prognosis of hepatocellular carcinoma and guide clinical drug treatment for different patients.

Despite advances in immunotherapy treatment, nonresponse rates remain high, and mechanisms of resistance to checkpoint inhibition remain unclear. To address this gap, we performed spatial transcriptomic and proteomic profiling on human hepatocellular carcinoma tissues collected before and after immunotherapy. We developed an interpretable, multimodal deep learning framework to extract key cellular and molecular signatures from these data. Our graph neural network approach based on spatial proteomic inputs achieved outstanding performance (ROC-AUC > 0.9) in predicting patient treatment response. Key predictive features and associated spatial transcriptomic profiles revealed the multi-omic landscape of immunotherapy response and resistance. One such feature was an interface niche expressing restrictive extracellular matrix factors that physically separates tumor tissue and lymphoid aggregates in nonresponders. We integrate this and other spatially-resolved signatures into SPARC, a multi-omic “fingerprint” comprising scores for immunotherapy response and resistance mechanisms. This study lays groundwork for future patient stratification and treatment strategies in cancer immunotherapy.