PO.ET01.02 · 实验与分子治疗
基于EV递送化学修饰siRNA在KRAS G12D突变胰腺癌中实现与MRTX1133相当的疗效
EV-based delivery of chemically modified siRNA achieves MRTX1133-comparable efficacy in KRAS G12D-mutant pancreatic cancer
作者与单位 Authors & Affiliations
摘要 Abstract
中文摘要
背景：KRAS G12D突变的胰腺导管腺癌（PDAC）在很大程度上仍无法治愈，尚无经临床验证的靶向疗法。RNA干扰可选择性抑制突变型KRAS，但当前的递送技术无法实现持续、安全的体内活性。我们开发了一种冲击波工程化的细胞外囊泡（EV）平台（SWEET™），它能够高效、稳定地包封靶向KRAS G12D的siRNA，并克服了脂质纳米颗粒（LNP）和小分子抑制剂的关键局限。
方法：我们设计并筛选了一个由105个成员组成的化学修饰siRNA panel，这些siRNA经工程改造以获得高稳定性和降低的先天免疫激活，以鉴定强效且等位基因特异的KRAS G12D沉默剂。优化的冲击波工程实现了>90%的包封率和>85%的核酸酶抗性。在PDAC细胞系、患者来源类器官（PDO）、异种移植、生物分布研究、毒性模型和符合GMP的放大工作流程中评估了先导候选物。
结果：五个siRNA表现出个位数纳摩尔级效力（IC50 < 10 nM）。先导物EV-siRNA #41产生了强效的KRAS G12D敲低和ERK/AKT信号的抑制。在体内，EV-siRNA #41相对于LNP实现了显著更优的肿瘤蓄积，并诱导了显著的肿瘤消退。值得注意的是，25 μg的EV-siRNA #41微剂量产生的抗肿瘤疗效与30-60 mg/kg的MRTX1133相当或更优。PDO测定证实对KRAS G12D突变肿瘤的选择性杀伤，而对野生型对照无影响。重复剂量GLP毒性研究确立了较高的NOAEL，无肝酶升高或体重下降。符合GMP的优化将EV产量从1%提高到7.5%，负载效率>80%。
结论：冲击波工程化的EV递送克服了PDAC中RNA疗法的主要障碍。EV-siRNA #41在微剂量水平展示出强效、选择性和持久的抗肿瘤活性，并优于LNP和一种领先的KRAS G12D抑制剂。这些数据将EV-siRNA #41定位为一种首创的、突变特异的RNA疗法，可进入支持IND的开发阶段。
致谢：本工作得到韩国药物开发基金（KDDF，资助号RS-2023-00282594）、科学和信息通信技术部技术开发计划（MSS，资助号RS-2023-00280797）以及2025年由韩国中小企业与创业部资助的旨在促进超级差距初创企业的超级差距初创企业1000+项目（资助号20266766）的支持。
查看英文原文 English abstract
Background: KRAS G12D-mutant pancreatic ductal adenocarcinoma (PDAC) remains largely incurable, with no clinically validated targeted therapies. RNA interference can selectively suppress mutant KRAS, yet current delivery technologies fail to achieve sustained, safe in vivo activity. We developed a shock wave-engineered extracellular vesicle (EV) platform (SWEET™) that enables efficient, stable encapsulation of KRAS G12D-targeting siRNA and overcomes key limitations of lipid nanoparticles (LNPs) and small-molecule inhibitors.
Methods: We designed and screened a 105-member panel of chemically modified siRNAs engineered for high stability and reduced innate immune activation to identify potent and allele-specific KRAS G12D silencers. Optimized shock-wave engineering achieved >90% encapsulation and >85% nuclease resistance. Lead candidates were evaluated across PDAC cell lines, patient-derived organoids (PDOs), xenografts, biodistribution studies, toxicity models, and GMP-aligned scale-up workflows.
Results: Five siRNAs exhibited single-digit nanomolar potency (IC50 < 10 nM). The lead, EV-siRNA #41, produced robust KRAS G12D knockdown and suppression of ERK/AKT signaling. In vivo, EV-siRNA #41 achieved markedly superior tumor accumulation relative to LNPs and induced significant tumor regression. Notably, a microdose of 25 μg EV-siRNA #41 produced antitumor efficacy comparable to or exceeding MRTX1133 at 30-60 mg/kg. PDO assays confirmed selective killing of KRAS G12D-mutant tumors with no effect on wild-type controls. Repeated-dose GLP toxicity studies established a high NOAEL without liver enzyme elevation or weight loss. GMP-aligned optimization increased EV yield from 1% to 7.5% with >80% loading efficiency.
Conclusions: Shock wave-engineered EV delivery overcomes the major barriers of RNA therapeutics in PDAC. EV-siRNA #41 demonstrates potent, selective, and durable antitumor activity at microdose levels and outperforms both LNPs and a leading KRAS G12D inhibitor. These data position EV-siRNA #41 as a first-in-class, mutation-specific RNA therapeutic ready for IND-enabling development.
Acknowledgments: This work was supported by the Korea Drug Development Fund (KDDF, Grant No. RS-2023-00282594), the Technology Development Program of the Ministry of Science and ICT (MSS, Grant No. RS-2023-00280797), and the Ultra-Gap Startup 1000+ Project for the promotion of ultra-gap startups, funded by the Ministry of SMEs and Startups of Korea in 2025 (Grant No. 20266766).
利益披露 Disclosure
H. kim, None..
Y. Choi, None..
K. Kim, None..
J. Kim, None..
S. An, None..
D. Bae, None..
M. Lee, None..
J. Chung, None..
S. Kim, None..
K. Kwon, None.