Characterization of ultrasound-mediated delivery of trastuzumab to normal and pathologic spinal cord tissue.

Extensive studies on focused ultrasound (FUS)-mediated drug delivery through the blood-brain barrier have been published, yet little work has been published on FUS-mediated drug delivery through the blood-spinal cord barrier (BSCB). This work aims to quantify the delivery of the monoclonal antibody trastuzumab to rat spinal cord tissue and characterize its distribution within a model of leptomeningeal metastases. 10 healthy Sprague-Dawley rats were treated with FUS + trastuzumab and sacrificed at 2-h or 24-h post-FUS. A human IgG ELISA (Abcam) was used to measure trastuzumab concentration and a 12 ± fivefold increase was seen in treated tissue over control tissue at 2 h versus no increase at 24 h. Three athymic nude rats were inoculated with MDA-MB-231-H2N HER2 + breast cancer cells between the meninges in the thoracic region of the spinal cord and treated with FUS + trastuzumab. Immunohistochemistry was performed to visualize trastuzumab delivery, and semi-quantitative analysis revealed similar or more intense staining in tumor tissue compared to healthy tissue suggesting a comparable or greater concentration of trastuzumab was achieved. FUS can increase the permeability of the BSCB, improving drug delivery to specifically targeted regions of healthy and pathologic tissue in the spinal cord. The achieved concentrations within the healthy tissue are comparable to those reported in the brain.

A Porcine Model of Transvertebral Ultrasound and Microbubble-Mediated Blood-Spinal Cord Barrier Opening.

Blood-spinal cord barrier opening, using focused ultrasound and microbubbles, has the potential to improve drug delivery for the treatment of spinal cord pathologies. Delivering and detecting ultrasound through the spine is a challenge for clinical translation. We have previously developed short burst, phase keying exposures, which can be used in a dual-aperture configuration to address clinical scale targeting challenges. Here we demonstrate the use of these pulses for blood-spinal cord barrier opening, in vivo in pigs. Methods: The spinal cords of Yorkshire pigs (n=8) were targeted through the vertebral laminae, in the lower thoracic to upper lumbar region using focused ultrasound (486 kHz) and microbubbles. Four animals were treated with a combination of pulsed sinusoidal exposures (1.0-4.0 MPa, non-derated) and pulsed short burst, phase keying exposures (1.0-2.0 MPa, non-derated). Four animals were treated using ramped short burst, phase keying exposures (1.8-2.1 MPa, non-derated). A 250 kHz narrowband receiver was used to detect acoustic emissions from microbubbles. Blood-spinal cord barrier opening was assessed by the extravasation of Evans blue dye. Histological analysis of the spinal cords was used to assess tissue damage and excised vertebral samples were used in benchtop experiments. Results: Ramped short burst, phase keying exposures successfully modified the blood-spinal cord barrier at 16/24 targeted locations, as assessed by the extravasation of Evans blue dye. At 4 of these locations, opening was confirmed with minimal adverse effects observed through histology. Transmission measurements through excised vertebrae indicated a mean transmission of (47.0 ± 7.0 %) to the target. Conclusions: This study presents the first evidence of focused ultrasound-induced blood-spinal cord barrier opening in a large animal model, through the intact spine. This represents an important step towards clinical translation.

Enhanced Detection of Bubble Emissions Through the Intact Spine for Monitoring Ultrasound-Mediated Blood-Spinal Cord Barrier Opening.

OBJECTIVE: We previously developed short burst, phase keying (SBPK) focused ultrasound (FUS) to mitigate standing waves in the human vertebral canal. Here, we show microbubble emissions from these pulses can be detected through the human vertebral arch and that these pulses are effective for blood-spinal cord barrier (BSCB) opening. METHODS: At f0 = 514 kHz, circulating microbubbles were sonicated through ex vivo human vertebrae (60 kPa-1 MPa) using a dual-aperture approach and SBPK exposures engineered to incorporate pulse inversion (PI). Signals from a 250 kHz receiver were analyzed using PI, short-time Fourier analysis and the maximum projection over the pulse train. In rats (n = 14), SBPK FUS+microbubbles was applied to 3 locations/spinal cord at fixed pressures (∼0.20-0.47 MPa). MRI and histology were used to assess opening and tissue damage. RESULTS: In human vertebrae between 0.2-0.4 MPa, PI amplified the microbubble/baseline ratio at f0/2 and 2f0 by 202 ± 40% (132-291%). This was maximal at 0.4 MPa, coinciding with the onset of broadband emissions. In vivo, opening was achieved at 40/42 locations, with mean MRI enhancement of 46 ± 32%(16%-178%). Using PI, f0/2 was detected at 14/40 opening locations. At the highest pressures (f0/2 present) histology showed widespread bleeding throughout the focal region. At the lowest pressures, opening was achieved without bleeding. CONCLUSION: This study confirmed that PI can increase sensitivity to transvertebral detection of microbubble signals. Preliminary in vivo investigations show that SBPK FUS can increase BSCB permeability without tissue damage. SIGNIFICANCE: SBPK is a clinically relevant pulse scheme and, in combination with PI, provides a means of mediating and monitoring BSCB opening noninvasively.