Study design and participants

This open label, prospective, proof-of-concept, phase I trial was designed to study the safety and feasibility of repeated BBB opening in patients with AD with demonstrated amyloid deposition in the targeted area. To improve safety, the study was divided into two stages, graded by the volume of brain tissue for BBB opening. Moreover, presumed non-eloquent cortex in the right frontal lobe, namely the superior frontal gyrus white matter of the dorsolateral prefrontal cortex (DLPFC), was selected to minimize potential complications in the event of bleeding or mass effect from vasogenic edema. The study was approved by the Research Ethics Board at Sunnybrook Health Sciences Centre (SHSC) and Health Canada. This study was registered with ClinicalTrial.gov number NCT02986932, and Health Canada number 195168. Prior to enrollment, all patients and their primary caregivers provided informed consent to the study, and publication of radiologic images.

Figure 1 outlines the study design. Patients between age 50 and 85 with mild-to-moderate AD35 with an MMSE score equal to or greater than 18 were eligible for the study. Detailed inclusion and exclusion criteria are listed in Table 3. In general, patients were referred to the study by neurologists and geriatricians. They were excluded if they had any contraindications to MRI, gadolinium or ultrasound contrast (Definity®), increased risk of bleeding, active intracranial diseases such as brain tumors or vascular malformations, or significant cardiovascular, pulmonary, and renal disease. During screening, patients underwent confirmation of their diagnosis by an expert in cognitive neurology, a pre-surgical anesthetic evaluation, baseline psychometric tests, and radiographic investigations with CT, MRI, and [18F]-florbetaben PET CT scans. Twenty-one patients were screened, of which five entered and completed the study, between March and September 2017.

Table 3 Inclusion and exclusion criteria Full size table

MRgFUS procedure

We used a focused ultrasound device consisting of 1024 individual transducers with a frequency of 220 kHz (ExAblate Neuro; InSightec Haifa). The device integrates intraoperative imaging, which was used for interim evaluations of the patient, and real-time acoustic monitoring to support decision-making on sonication parameters. On the day of the procedure, a Cosman-Roberts-Wells (CRW) stereotactic frame was fixed to the patient’s head under local anesthetic. The frame was then coupled to the helmet transducer array, with the patient entering the MRI supine and awake. A safety switch was given to the patient to abort the procedure in case of discomfort or pain. The patient was examined and questioned for adverse events after each sonication.

A 3-Tesla MRI (Signa MR750; GE Healthcare, Milwaukee, Wis.) was used to obtain T1, T2 (fast spin echo), and T2* (gradient echo) weighted images for surgical planning. A region in the right frontal lobe was then selected for BBB opening. To minimize the risk, we avoided areas containing sulci and vessels within two contiguous MRI slices in each of the axial, sagittal, and coronal planes. Once the target region was identified, patients received a weight-based intravenous injection of microbubble contrast (Definity®) (4 μl/kg), followed shortly by the application of low-frequency focused ultrasound to the target. MR thermometry was used to monitor tissue temperature at the sonicated region in real time. The sonication parameters were limited by the clinical device hardware and software, and corresponded to those previously tested in large animal models36.

At each new target, a power ramp test was performed with the first microbubble injection. This test involves applying short sonications with increasing power in 5% increments until the device hydrophones detect a sub-harmonic acoustic feedback from the target, indicating a cavitation. Subsequent sonications are then performed at 50% of this ‘cavitation threshold’ power. The ramp test was developed from preclinical studies to determine the optimal power required for safe opening of the BBB36,37. Sonication volumes covered a rectangular spot approximately 9 mm by 9 mm, comprised of 3-by-3 grid of spots, each 3 mm in diameter. For the last three patients, given the extent of atrophy on their MRI, a 2-by-2 grid was utilized, yielding a spot approximately 5 mm by 5 mm. The device electronically steered the ultrasound through each grid for 50 s total, sonicating each spot with 2 ms on and 28 ms off bursts for 300 ms, with a repetition interval of 2.7 s (duty cycle 0.74%). For stage 2, performed approximately 1 month following stage 1, the procedure was repeated, opening the BBB at the original location as well as at an adjacent area, following the same protocol, but doubling the volume of tissue opened.

After completion of the sonication protocol, a gadolinium-enhanced T1 sequence was performed to verify definitive evidence of BBB opening. Contrast enhancement at the targeted region signified the end of the procedure. The patient was then taken out of the scanner, the stereotactic frame removed, and additional high-resolution MRI sequences obtained. Patients were admitted to the surgical short stay unit for overnight observation.

Outcomes

The primary outcomes were clinical and radiographic safety as well as technical feasibility of reversible and repeated BBB opening. Successful barrier opening and restoration was determined, respectively, by gadolinium leakage immediately after sonication and by the absence of enhancement 1 day after sonication at the target region on T1-weighted contrast images. Safety was measured by clinical exam during the procedure and at each follow-up, as well as radiographic examination for any adverse events, including hemorrhage, swelling, or mass effect. Follow-up visits were scheduled for 1 day, 1 week, and 1 month after each procedure, as well as 2 months following the second procedure (Fig. 1). Adverse events were recorded and monitored in a prospective fashion.

Secondary outcomes were Alzheimer’s-specific psychometrics, and exploratory outcomes included regional changes in [18F]-florbetaben binding on PET. Psychometric tests administered at 1-day, 1-week, 1-month, and 2-months after sonication included the MMSE, Geriatric Depression Scale (GDS), Alzheimer’s Disease Assessment Scale-cognitive (ADAS-Cog), NPI-Q, and Alzheimer’s Disease Cooperative Study Activity of Daily Living Scale (ADCS-ADL).

[18F]-Florbetaben PET CT image acquisition and analysis

PET CT scans to measure beta-amyloid deposition were performed at baseline and 1 week following each procedure. A transmission scan followed by a 20-min emission scan (four frame/5 min each) were acquired on the Phillips Gemini PET CT (3D mode) starting at 90 min after an 8 mCi ± 20% (n = 14) radiotracer injection of [18F]-florbetaben. The SUVr was calculated on a voxel-wise basis by dividing the summed PET images by the cerebellar gray matter ROI, consistent with other [18F]-florbetaben studies29. To derive the cerebellar gray matter ROI, the T1-weighted MR images were processed with the Freesurfer pipeline (version 5.1; http://surfer.nmr.mgh.harvard.edu/). The radioactivity in the cerebellar reference region was extracted after mapping the cerebellar gray matter ROI to the co-registered [18F]-florbetaben scans. Image preprocessing was performed with statistical parametric mapping, version eight (SPM8, Institute of Neurology, London). PET-to-PET and MR-to-PET registrations were performed using the normalized mutual information algorithm, and images were spatially normalized into standard 3D space relative to the anterior commissure using the Alzheimer’s Disease Neuroimaging Initiative (ADNI) template38.

To measure the effects of MRgFUS-mediated BBB opening on beta-amyloid deposition, the sonication ROIs were manually delineated as the contrast enhanced areas plus adjacent gray and white matter on gadolinium MR images over 6–8 contiguous slices. These ROIs were mapped onto the co-registered T1-weighted MR and PET scans. Of note, the ROIs for stage 2 were larger than stage 1, consistent with the larger sonication volumes.

Role of funding source

InSightec, the manufacturer of the ExAblate device used in this study, was the regulatory sponsor and had no role in study design, data collection, analysis, or interpretation. This study was funded by a grant from the Focused Ultrasound Foundation, a non-profit organization that funds research into clinical applications of ultrasound. The corresponding author had full access to all the data in the study and had final responsibility for the decision to submit for publication.

Data availability

The data generated during the current study are available from the corresponding author on request.