PO.ET02.04 · 实验与分子治疗
KY102：一种携带拓扑异构酶1抑制剂载荷的新型Glypican 3靶向纳米抗体药物偶联物，在肝细胞癌模型中展现出引人注目的临床前活性
KY102, a novel Glypican 3-targeting nanobody drug conjugate bearing a topoisomerase 1 inhibitor payload demonstrates compelling preclinical activity in hepatocellular carcinoma models
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摘要 Abstract
中文摘要
背景：Glypican-3（GPC3）是一种细胞表面癌胚糖蛋白，在肝细胞癌（HCC）中高表达，而在正常成人组织中存在极少。KY102是一种靶向人GPC3的纳米抗体药物偶联物（NDC），由一个单价纳米抗体经马来酰亚胺锚定和甘氨酰甘氨酰苯丙氨酰甘氨酸（GGFG）-氨甲基（AM）可裂解连接子共价偶联至拓扑异构酶1抑制剂DXD构成，药物与纳米抗体比（DAR）为3。
材料与方法：进行了广泛的功能表征，以评估KY102（NDC）在DAR为3时的作用机制和治疗潜力。
1.通过生物层干涉（BLI）和流式细胞术（FACs）评估纳米抗体与人、食蟹猴、大鼠和小鼠GPC3的结合。
2.使用177-Lu标记的纳米抗体在正常及细胞系来源动物模型中进行脱靶结合和靶点特异性的评估。
3.通过流式细胞术评估KY102纳米抗体在表达GPC3的肿瘤细胞系中的内化。
4.在一组HCC细胞系中评估体外KY102（NDC）对肿瘤单层的细胞毒性。
5.还进行了肿瘤细胞共培养测定，以评估KY102介导的旁观者杀伤作用。
6.在一组代表不同GPC3表达水平的细胞系来源异种移植（CDX）和患者来源异种移植（PDX）小鼠模型中研究了KY102的抗肿瘤活性。
7.在小鼠中的细胞系来源异种移植（CDX）中，通过单剂量和重复剂量（4次给药；每周一次）非GLP毒理学研究评估了KY102的耐受性。
结果：
1.KY102纳米抗体骨架对人、食蟹猴、大鼠和小鼠GPC3均展现出皮摩尔级结合亲和力，并与靶点表达的癌细胞系强结合。
2.在表达GPC3的HCC细胞系中观察到KY102纳米抗体的快速内化。
3.KY102在一组以单层培养的HCC细胞中表现出强效且靶点特异性的细胞毒性。
4.当与GPC3阳性癌细胞共培养时，KY102显示出对GPC3阴性癌细胞的有效旁观者介导杀伤。
5.单次给药KY102（2.5mg/kg和5.0 mg/kg）在一组代表不同GPC3表达水平的CDX和PDX模型中产生了稳健的肿瘤生长抑制。
6.KY102的单剂量和重复剂量小鼠毒理学研究在30和60 mg/kg剂量水平显示出令人鼓舞的耐受性。
7.在2.5 mg/kg和5 mg/kg剂量水平呈现剂量依赖性纳米抗体药代动力学，未确定最大耐受剂量（MTD）。
结论：总体而言，这些结果支持KY102作为针对包括HCC在内的GPC3阳性癌症的新型治疗药物的潜力。预计将于2026年提交IND申请。无利益冲突。
查看英文原文 English abstract
Background: Glypican-3(GPC3), a cell-surface oncofetal glycoprotein highly expressed in hepatocellular carcinoma (HCC) with minimal presence in normal adult tissues. KY102 is a nanobody-drug conjugate (NDC)targeting human GPC3, composed of a monovalent nanobody covalently conjugated to a topoisomerase 1 inhibitor, DXD. via a maleimide anchor and a glycyl glycyl phenylalanyl glycine (GGFG)- aminomethyl (AM) cleavable linker at a drug to nanobody ratio (DAR) of 3.
Materials and Methods: Extensive functional characterization was performed to assess the mechanism of action and therapeutic potential of the KY102 (NDC) at DARs of 3.
1.Nanobody binding to human,cynomolgus monkey rat and mouse GPC3 was assessed by biolayer interferometry (BLI) and flow cytometry(FACs).
2.Am assessment of off-target binding, and target specificity was conducted using 177-Lu labeled Nanobody in normal and cell line derived animalmodel.
3.KY102 nanobody internalization in GPC3-expressing tumor cell lines was assessed by flow cytometry.
4.In vitro KY102 (NDC) cytotoxicity against tumor monolayers was assessed in a panel of HCC cell lines.
5.Tumor cell co-culture assays were also performed to assess bystander-mediated cell killing by KY102.
6.Anti-tumor activity of KY102 was investigated in a panel of cell line-derived xenograft (CDX) and patient-derived xenograft (PDX) mouse models representing a range of GPC3 expression.
7.The tolerability of KY102 was assessed in a single dose and repeat-dose (4 doses; once every weeks) non-GLP toxicology study in a cell line-derived xenograft (CDX) in mice.
Results:
1.The KY102 nanobody backbone demonstrated pico-molar binding affinity to both human, cynomolgus monkey, rat and mouse GPC3, and strong binding to target-expressing cancer cell lines.
2.Rapid internalization of KY102 nanobody was observed in GPC3-expressing HCC cell lines.
3.KY102 exhibited potent and target-specific cytotoxicity in a panel of HCC cells cultured either in monolayer.
4.KY102 showed effective bystander-mediated killing of GPC3 negative cancer cells when in co-culture with GPC3 positive cancer cells.
5.A single administration of KY102 ( 2.5mg/kg and 5.0 mg/kg) resulted in robust tumor growth inhibition of a panel of CDX and PDX models representing a range of GPC3-expression.
6.A single dose and a repeat-dose mouse toxicology study of KY102 showed encouraging tolerability at 30, and 60 mg/kg dose levels.
7.A dose-dependent nanobody pharmacokinetics at 2.5 mg/kg, and 5 mg/kg dose levels, without the maximum tolerated dose (MTD) determined.
Conclusions: Overall, these results support the potential of KY102 as a novel therapeutic agent against GPC3-bearing cancers including HCC. An IND application is anticipated in 2026. No conflict of interest
利益披露 Disclosure
H. Shi, None.