血脑屏障（BBB）的存在限制了中枢神经系统疾病的药物治疗。近年来，在中枢神经系统药物传递促进剂领域取得了显著进展，如MR引导的聚焦超声技术已进入临床试验阶段。这激发了该领域的研究人员发明新的脑屏障开启（BBo）技术，这些技术需要简单、快速、安全和高效。我们最近开发的一种技术是基于低脉冲电场（L-PEFs）的屏障破坏电场（BDF），以控制、安全、可逆和非侵入的方式打开BBB。在这里，我们进行了活体研究，以展示BDF是一种将多柔比星（Doxo）输送到小鼠大脑中的可行技术。我们开发并应用了描述BBBo水平的方法，用于监测治疗并预测反应。总体而言，本研究的目标是证明将治疗性Doxo剂量输送到原始和带瘤小鼠脑中的可行性，并应用延迟对比度MRI（DCM）监测BBBo水平。L-PEFs是使用放置在原始小鼠完整颅骨上的板电极进行的。L-PEFs/Sham小鼠在程序结束后立即通过DCM进行扫描（“MRI实验”），或者注射Doxo和Trypan blue，然后延迟（4小时）灌注和提取脑组织（“Doxo实验”）。使用共聚焦显微镜在脑样本中测量Doxo浓度，并与体外胶质瘤细胞株中的Doxo IC50进行比较。为了绘制整个脑中的BBBo范围，使用DCM数据对像素进行MR图像分析。最后，在裸鼠携带的颅内人类胶质瘤肿瘤中测试了L-PEFs与Doxo的疗效。所有L-PEFs处理的小鼠脑皮层区域都有大量Doxo（0.50±0.06 μg Doxo/gr brain），而Sham小鼠脑中的Doxo浓度低于或接近检测限度（0.03±0.02 μg Doxo/gr brain）。这个浓度比在gl261小鼠胶质瘤细胞中计算的Doxo的IC50高97倍，比在U87人类胶质瘤细胞中计算的Doxo的IC50高8倍。DCM分析显示L-PEFs处理的小鼠的皮层区域有显著的BBBo水平；与Sham组相比，L-PEFs处理的小鼠中BBBo的平均体积增加了29倍。计算的BBBo水平随着BBBo阈值的指数下降，类似于大脑中电场的分布。最后，将非侵入性L-PEFs与Doxo联合使用显著降低了裸鼠脑肿瘤的生长速率。我们的结果显示，额外颅外L-PEFs诱导了显著的BBBo水平，使治疗性Doxo剂量能高效输送到大脑中，并减少肿瘤生长。由于标准对比增强MRI无法检测到BBBo，所以应用DCM生成了整个大脑中BBBo水平的图。这些发现表明，BDF是一种有前景的将药物高效输送到脑部的技术，并对未来治疗脑癌和其他中枢神经系统疾病具有重要意义。© 2023. 中央档案馆出版社有限公司， 斯普林格自然出版集团的一部分。

Pharmacological treatment of CNS diseases is limited due to the presence of the blood-brain barrier (BBB). Recent years showed significant advancement in the field of CNS drug delivery enablers, with technologies such as MR-guided focused ultrasound reaching clinical trials. This have inspired researchers in the field to invent novel brain barriers opening (BBo) technologies that are required to be simple, fast, safe and efficient. One such technology, recently developed by us, is BDF (Barrier Disrupting Fields), based on low pulsed electric fields (L-PEFs) for opening the BBB in a controlled, safe, reversible and non-invasive manner. Here, we conducted an in vivo study to show that BDF is a feasible technology for delivering Doxorubicin (Doxo) into mice brain. Means for depicting BBBo levels were developed and applied for monitoring the treatment and predicting response. Overall, the goals of the presented study were to demonstrate the feasibility for delivering therapeutic Doxo doses into naïve and tumor-bearing mice brains and applying delayed-contrast MRI (DCM) for monitoring the levels of BBBo.L-PEFs were applied using plate electrodes placed on the intact skull of naïve mice. L-PEFs/Sham mice were scanned immediately after the procedure by DCM ("MRI experiment"), or injected with Doxo and Trypan blue followed by delayed (4 h) perfusion and brain extraction ("Doxo experiment"). Doxo concentrations were measured in brain samples using confocal microscopy and compared to IC50 of Doxo in glioma cell lines in vitro. In order to map BBBo extent throughout the brain, pixel by pixel MR image analysis was performed using the DCM data. Finally, the efficacy of L-PEFs in combination with Doxo was tested in nude mice bearing intracranial human glioma tumors.Significant amount of Doxo was found in cortical regions of all L-PEFs-treated mice brains (0.50 ± 0.06 µg Doxo/gr brain) while in Sham brains, Doxo concentrations were below or on the verge of detection limit (0.03 ± 0.02 µg Doxo/gr brain). This concentration was x97 higher than IC50 of Doxo calculated in gl261 mouse glioma cells and x8 higher than IC50 of Doxo calculated in U87 human glioma cells. DCM analysis revealed significant BBBo levels in the cortical regions of L-PEFs-treated mice; the average volume of BBBo in the L-PEFs-treated mice was x29 higher than in the Sham group. The calculated BBBo levels dropped exponentially as a function of BBBo threshold, similarly to the electric fields distribution in the brain. Finally, combining non-invasive L-PEFs with Doxo significantly decreased brain tumors growth rates in nude mice.Our results demonstrate significant BBBo levels induced by extra-cranial L-PEFs, enabling efficient delivery of therapeutic Doxo doses into the brain and reducing tumor growth. As BBBo was undetectable by standard contrast-enhanced MRI, DCM was applied to generate maps depicting the BBBo levels throughout the brain. These findings suggest that BDF is a promising technology for efficient drug delivery into the brain with important implications for future treatment of brain cancer and additional CNS diseases.© 2023. BioMed Central Ltd., part of Springer Nature.