Safety assessment after delivering drugs across the blood-brain barrier using Rapid Short-Pulse sequences in vivo

Safety assessment after delivering drugs across the blood-brain barrier using Rapid Short-Pulse sequences in vivo
Fateme Nasrollahi Boroujeni,^1,* Mohamad Taghi Ahmadian,² Dina Morshedi,³
1. Bioprocess Engineering Research Group, Department of Industrial and Environmental Biotechnology, National Institute of Genetic Engineering and Biotechnology, Tehran, Iran.
2. School of Mechanical Engineering, Sharif University of Technology, Azadi Ave, PO 11365-11155, Tehran, Iran
3. Bioprocess Engineering Research Group, Department of Industrial and Environmental Biotechnology, National Institute of Genetic Engineering and Biotechnology, Tehran, Iran.
Introduction: Focused ultrasound and microbubbles can alter the blood-brain barrier (BBB) permeability, allowing drugs to enter the brain. However, when using long ultrasound pulses (10 ms), the BBB remains open for several hours or days, which allows unwanted bloodborne proteins, such as albumin and immunoglobulins, to enter the brain and trigger a neuroimmune response (i.e., activation of microglial cells). Recently, we have developed a Rapid Short-Pulse (RaSP) ultrasound sequence that alters the BBB permeability more uniformly throughout the brain and for less than 10 minutes. Here, we explored whether the shorter duration of BBB disruption is representative of a safer delivery profile. We therefore evaluated whether the RaSP sequence reduces albumin and immunoglobulin extravasation and whether the neuroimmune system remains inactive.
Methods: We applied a RaSP sequence (pulse length: 5 cycles, repetition frequency: 1.25 kHz) or ms-long sequence (10,000 cycles; 0.5 Hz) onto the left murine hippocampus (1 MHz, 350 kPa, n = 5). Fluorescent 3 kDa dextran, our model drug, was systemically administered before the injection of SonoVue® microbubbles. Brains were extracted and sectioned either 0, 10 or 20 min after the ultrasound treatment to assess the extent of albumin and immunoglobulin extravasation; or 0, 2, 24 or 48 h after ultrasound exposure to investigate the involvement of microglia and astrocytes, both via immunofluorescence staining
Results: Although the RaSP sequence deposited 150 times less acoustic energy into the brain than long pulses, a similar dextran dose was delivered to the brain. Whereas long pulses resulted in albumin extravasating into the brain at all the time points tested, the RaSP sequence resulted in 3.7-fold less albumin extravasation at 0 min and an undetectable level of albumin at 10 and 20 min. With the RaSP sequence, no immunoglobulins were detected at all time points. Also, staining revealed that the RaSP sequence produced no uptake of the model drug within microglia and astrocytes, while with long pulses the microglia showed higher uptake and had a more rounded shape, a sign of activation. These results indicate that RaSP sequences can deliver drugs to the brain with a nearly negligible level of BBB disruption and neuroimmune response.
Conclusion: Safety features of RaSP and ms-long pulse sequences was confirmed.we demonstrated Less albumin and no immunoglobulin extravasates into the brain at 0, 10 and 20 min after ultrasound emitted in a RaSP sequence and Little or no uptake of dextran at 0, 2, 24 and 48 h in microglia and astrocytes, shows less of an immune response.
Keywords: Blood-Brain Barrier, Drug Dlivery, Focused Ultrasound, Rapid Sh