Focused Ultrasound Blood-Brain Barrier Opening Arrests the Growth and Formation of Cerebral Cavernous Malformations

Introduction: Cerebral cavernous malformations (CCM) are vascular lesions within the central nervous system, consisting of dilated and hemorrhage-prone capillaries. CCMs can cause debilitating neurological symptoms, and surgical excision or stereotactic radiosurgery are the only current treatment options. As an incisionless therapy with the ability to target eloquent brain locations, focused ultrasound (FUS) may represent an ideal alternative for CCM treatment. FUS delivers acoustic energy deep within the body. When FUS is combined with an intravenous (i.v.) injection of gas-filled microbubbles, the pressure waves induce microbubble oscillation, which pushes and pulls on blood vessel walls. If performed in the brain, this procedure can induce a temporary opening of the blood-brain barrier (BBB). FUS-mediated BBB opening (BBBO) has been deployed primarily to enable enhanced delivery of drugs and other therapeutic agents into the brain for various neurological conditions. However, FUS BBBO has also been shown to be beneficial in the absence of drug delivery for the treatment of Alzheimer’s disease, leading to several clinical trials that are testing this approach. In this study, we examined the effectiveness and safety profile of FUS BBBO applied to CCMs and its potential to, in the absence of drug delivery, therapeutically control the growth and de novo formation of CCMs.

Materials and

Methods: CCMs were induced in mice by postnatal, endothelial-specific Krit1 ablation. FUS BBBO was performed with a FUS Instruments RK-300 system with a 1.13 MHz single-element transducer using a 10 ms burst length over a 2000 ms period for 60 total sonications during a 2-min sonication duration. Fixed peak-negative pressure (PNP) application was performed using “Burst” mode, while passive cavitation modulated (PCD)-modulated PNP was performed using the “Blood-brain Barrier” mode. Parameters used for the feedback control system included a starting pressure of 0.2 MPa, pressure increment of 0.05 MPa, maximum pressure of 0.4 MPa, 20 sonication baselines without microbubbles, AUC bandwidth of 500 Hz, AUC threshold of 10 standard deviations, pressure drop of 0.95, and frequency selection of the subharmonic, first ultraharmonic, and second ultraharmonic. Gadolinium contrast agent (Multihance) was injected as a bolus prior to T1-RARE image acquisition. Albumin-shelled microbubbles were intravenously injected as a bolus dose of 10^5 microbubbles per gram body weight. High resolution T2-weighted images and T1-RARE images were used to guide FUS targeting to pre-selected CCMs. A single sonication target was used in all experiments, except in the case of PCD-modulated PNPs, in which two sonication targets were used. Mice receiving the repeat FUS BBBO regimens had all sonications staged 3 days apart with the same anatomical location targeted each time.

Results, Conclusions, and Discussions: FUS BBBO elicited gadolinium accumulation primarily at the perilesional boundaries of CCMs, rather than lesion cores (Figure 1A-C). Passive cavitation detection and gadolinium contrast enhancement were comparable in CCM and wild-type mice, indicating that Krit1 ablation does not confer differential sensitivity to FUS BBBO. Acutely, CCMs exposed to FUS BBBO remained structurally stable, with no signs of hemorrhage. Longitudinal MRI revealed that FUS BBBO halted the growth of 94% of CCMs treated in the study (Figure 1D-E). At 1 month, FUS BBBO-treated lesions lost, on average, 9% of their pre-sonication volume. In contrast, non-sonicated control lesions grew to 670% of their initial volume (Figure 1E). Strikingly, in mice receiving multiple BBBO treatments with fixed PNPs, de novo CCM formation was significantly reduced by 81% (Figure 1F, G). Lesion control with FUS BBBO was accompanied by a marked reduction in the area and mesenchymal appearance of Krit mutant endothelium (Figure 1H, I), suggesting that FUS BBBO controls CCM growth via the reversal of endothelial-to-mesenchymal transition in Krit1 mutant endothelial cells. Mock treatment plans on MRIs of patients with surgically inaccessible lesions revealed their lesions are amenable to FUS BBBO with current clinical technology. Our results establish FUS BBBO as a novel, non-invasive modality that can safely arrest murine CCM growth and prevent their de novo formation. As an incisionless, MR image-guided therapy with the ability to target eloquent brain locations, FUS BBBO offers an unparalleled potential to revolutionize the therapeutic experience and enhance the accessibility of treatments for CCM patients.

Acknowledgements (Optional): Supported by NIH R01CA279134, R01EB030409, and R01EB030744