PO.TB04.05 · 肿瘤生物学
点亮你的靶点：内源性 HiBiT 敲入单克隆细胞系库存 + 快速定制
Illuminate your targets: Endogenous HiBiT knock-in monoclones-inventory + rapid custom
作者与单位 Authors & Affiliations
摘要 Abstract
中文摘要
背景与意义——应用 HiBiT 是一段 11 个氨基酸的标签，可与 LgBiT 互补以重构 NanoLuc，从而提供明亮、线性的生物发光读出信号，其强度随靶蛋白丰度按比例变化。与以 Western blot 为中心的工作流程相比，内源性 HiBiT 敲入可在天然调控下实现实时、定量的动力学检测，从而加速机制验证、效力排序、SAR 分析以及跨多种模态（PROTAC、分子胶、LYTAC 和 AbTAC）的联合设计。
Kyinno 案例系列——库中内容（代表性范围） Kyinno 已建立了一个广泛且可即用的 HiBiT-KI 单克隆细胞系组合，涵盖 30 多个内源性标记基因，横跨多个生物学类别——致癌信号通路、激酶、转录因子以及适合 E3 的靶点。这些包括代表性节点，如 KRAS、CDK 家族成员、PRMT5、EGFR、AR、BRD 家族蛋白、EZH2、STATs 等。因此，该组合全面覆盖了与靶向蛋白降解、信号转导和转录调控相关的主要靶点类别。快速定制敲入（N 端或 C 端）进一步将该平台扩展至项目特异性靶点和变体，使团队能够立即使用库存模型入手，或迅速获得定制构建体。
方法与部署——目前的工作原理 我们采用双稳定系统：(i) 基因组整合的 LgBiT 以最大限度减少信号漂移，以及 (ii) CRISPR-Cas9 HDR 在内源性基因座精确插入 HiBiT，同时保留基因剂量和蛋白功能。单细胞克隆经过序列验证、STR 鉴定，且无支原体污染。标准化的 NanoLuc 检测（10 点曲线、72-96 小时或实时动力学）输出定量的降解/周转轨迹，并具有板间可比性。该平台将检测建立时间从数周压缩至数天，统一了跨靶点和跨宿主的读出信号，并提供了一条从苗头化合物筛选到转化方法转移的可扩展路径——将 HiBiT 定量带到你需要的每一处。
查看英文原文 English abstract
Background & significance - applications HiBiT is an 11-aa tag that complements LgBiT to reconstitute NanoLuc, delivering a bright, linear bioluminescent readout that scales with target protein abundance. Compared with Western blot-centric workflows, endogenous HiBiT knock-ins enable real-time, quantitative kinetics under native regulation, accelerating mechanism confirmation, potency ranking, SAR, and combination design across modalities-PROTACs, molecular glues, LYTACs, and AbTACs.
Kyinno case series - what's in the panel (representative scope) Kyinno has established a broad, ready-to-run portfolio of HiBiT-KI monoclonal cell lines covering more than 30 endogenously tagged genes across diverse biological categories-oncogenic signaling, kinases, transcription factors, and E3-amenable targets. These include representative nodes such as KRAS, CDK family members, PRMT5, EGFR, AR, BRD family proteins, EZH2, STATs, and so on. The panel thus provides comprehensive coverage of major target classes relevant to targeted protein degradation, signal transduction, and transcriptional regulation. Rapid custom knock-in (N- or C-terminal) further extends the platform to program-specific targets and variants, allowing teams to start immediately with inventory models or quickly obtain bespoke constructs.
Methods & deployment - how it works now We implement a dual-stable system: (i) genome-integrated LgBiT to minimize signal drift, and (ii) CRISPR-Cas9 HDR to precisely insert HiBiT at endogenous loci while preserving gene dosage and protein function. Single-cell clones are sequence-verified, STR-authenticated, and mycoplasma-free. Standardized NanoLuc assays (10-point curves, 72-96 h or live kinetic) output quantitative degradation/turnover trajectories with plate-to-plate comparability. This platform compresses assay setup from weeks to days, unifies readouts across targets and hosts, and provides a scalable path from hit triage to translational method transfer-bringing HiBiT quantitation everywhere you need it.
利益披露 Disclosure
X. Gou,
Kyinno Biotechnology Co., LTD Employment.
Y. Huang,
Kyinno Biotechnology Co., LTD Employment.
Y. Wang,
Kyinno Biotechnology Co., LTD Employment.
J. Ning,
Kyinno Biotechnology Co., LTD Employment.
F. Hao,
Kyinno Biotechnology Co., LTD Employment.