溶瘤新城疫病毒静脉给药的生物分布及安全性评价

Biodistribution and safety of intravenously administered oncolytic Newcastle disease virus

摘要

目的 评估溶瘤病毒静脉给药的安全性与体内分布。方法 以新城疫病毒 (NDV) Italien 速发型毒株为对象, 在免疫健全 BALB/c 小鼠中系统评价其经静脉注射后的安全性及生物分布。将小鼠随机分为 4 组: 对照组 (PBS)、低剂量组 (5 × 10⁸ PFU)、中剂量组 (1 × 10⁹ PFU) 和高剂量组 (5 × 10⁹ PFU), 每组单次尾静脉注射给药。通过观察一般状况、体质量变化、死亡率及组织病理学 (HE 染色) 评估急性毒性。采用 qRT-PCR 检测单次注射 (5 × 10⁸ PFU) 后不同时间点 (1 h、6 h、1 d、3 d、7 d) 及多次注射 (5 × 10⁷ PFU/次, 每 3 d 一次, 共 3 次) 后心、肝、脾、肺、肾及血浆中的病毒基因组拷贝数, 并计算药代动力学参数。另设 rNDV-Luci (表达荧光素酶的重组病毒) 组, 通过小动物活体成像观察病毒在体内 (24、72 h) 的复制动态。结果 单次静脉注射 NDV Italien 后, 未观察到不良效应的最高剂量为 1 × 10⁹ PFU, 为有效治疗剂量的 100 倍。低剂量组 (5 × 10⁸ PFU) 和中剂量组 (1 × 10⁹ PFU) 小鼠全部存活, 仅表现为短暂体质量下降和轻微组织病理改变; 高剂量组 (5 × 10⁹ PFU) 小鼠在 48 h 内全部死亡, 并出现弥漫性肺出血和局灶性肝坏死等严重损伤。病毒基因组定量分析表明, 肺是病毒分布的首选靶器官, 注射后 6 h 达到峰值, 随后迅速清除; 肝脏、脾脏等器官病毒载量较低, 呈单相清除特征。活体成像进一步证实病毒主要在肺部呈现一过性复制, 72 h 内基本清除。多次给药未导致病毒在血浆或组织中蓄积, 脾脏表现出最显著的清除能力。结论 NDV Italien 株在免疫健全小鼠中经静脉注射给药在治疗剂量范围内具有良好安全性, 但 5 × 10⁹ PFU 剂量可引起肺和肝脏损伤。肺作为主要靶器官, 提示其对肺部肿瘤具有潜在治疗优势, 为重组速发型 NDV 的临床转化提供了重要的安全性依据。

Abstract

Objective To evaluate the safety and in vivo distribution of oncolytic viruses administered intravenously. Methods The velogenic strain Newcastle disease virus (NDV) Italien was used to systematically assess its safety and biodistribution after intravenous injection in immunocompetent BALB/c mice. Mice were randomly divided into four groups: control group (PBS), low-dose group (5 × 10⁸ PFU), medium-dose group (1 × 10⁹ PFU), and high-dose group (5 × 10⁹ PFU), each receiving a single tail vein injection. Acute toxicity was evaluated by observing general condition, body mass changes, mortality, and histopathology (HE staining). qRT-PCR was used to detect viral genome copy numbers in the heart, liver, spleen, lung, kidney, and plasma at different time points (1 h, 6 h, 1 d, 3 d, 7 d) after a single injection (5 × 10⁸ PFU) and after multiple injections (5 × 10⁷ PFU per injection, once every 3 d for a total of 3 injections). Pharmacokinetic parameters were calculated. Additionally, an rNDV-Luci (recombinant virus expressing luciferase) group was established to observe viral replication dynamics in vivo (at 24 h and 72 h) using small animal in vivo imaging. Results After a single intravenous injection of NDV Italien, the no-observed-adverse-effect level was 1 × 10⁹ PFU, which was 100 times the effective therapeutic dose. All mice in the low-dose (5 × 10⁸ PFU) and medium-dose (1 × 10⁹ PFU) groups survived, showing only transient body mass loss and mild histopathological changes. Mice in the high-dose group (5 × 10⁹ PFU) all died within 48 h, presenting severe injuries including diffuse pulmonary hemorrhage and focal hepatic necrosis. Quantitative viral genome analysis showed that the lung was the primary target organ for viral distribution, reaching peak levels at 6 h post-injection followed by rapid clearance. Viral loads in the liver, spleen, and other organs were lower and exhibited a monophasic clearance pattern. In vivo imaging further confirmed that viral replication was transient and mainly localized in the lung, being largely cleared within 72 h. Multiple injections did not lead to viral accumulation in plasma or tissues, with the spleen demonstrating the most prominent clearance capacity. Conclusion Intravenous administration of NDV Italien strain is well tolerated within the therapeutic dose range in immunocompetent mice, although a dose of 5 × 10⁹ PFU can cause lung and liver injuries. The lung is the primary target organ, suggesting a potential therapeutic advantage for pulmonary tumors. This study provides important safety evidence for the clinical translation of recombinant velogenic NDV.

关键词

溶瘤病毒 / 新城疫病毒 / 静脉注射 / 生物分布 / 安全性 / 肿瘤生物治疗 / 肿瘤免疫治疗 / 溶瘤病毒治疗

Key words

oncolytic virus / Newcastle disease virus / intravenous injection / biodistribution / safety / tumor biotherapy / tumor immunotherapy / oncolytic virotherapy

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基金资助

陕西省重点研发计划项目(2024SF-YBXM-400)