Targeted NPR-C PET/MR imaging of carotid atherosclerosis in humans: correlation with ex vivo plaque immunohistochemistry (IHC) and patient outcomes

Objectives: It is unclear which asymptomatic carotid artery stenosis (ACAS) patients (≥ 70% diameter) benefit from carotid endarterectomy (CEA) intervention. We have identified a natriuretic peptide receptor (NPRC) that is present in deep intimal macrophages and vascular smooth muscle cells and is up-regulated in plaque with features of instability. We have developed a nanoparticle radiotracer, ⁶⁴Cu-CANF-Comb, directly targeting this receptor. Here we assess whether uptake of this radiotracer correlates with presence of NPRC on immunohistochemistry (IHC) of ex vivo carotid specimens in patients who went on to CEA surgery and to features of instability (large lipid pool, hemorrhage), and retrospectively assess for cerebrovascular events in patients who did not go on to CEA. Methods: Forty-four (44) patients (69.7±10.0 years; 19 women) with ACAS were recruited; 42 successfully completed carotid PET/MRI imaging approximately 18 hours after injection of 3.5-5.1 mCi ⁶⁴Cu-CANF-Comb. PET acquisition was list mode for ~ 30 min. MR imaging consisted of high-resolution (0.5-0.7 mm) bright blood 3D GRE, T2 3D SPACE, dark blood TSE T1, and T2/PD imaging using small neck surface coils. CEA specimens were collected post surgery for IHC. Fifteen (15) specimens were stained with an antibody (anti-NPRC, 1:100 in blocking serum) and a secondary antibody labeled with a blue chromogen and counterstained with nuclear fast red. Five 100X fields were randomly selected in the superficial intima, deep intima and media of the specimens. NPRC positive cells were counted in each field, and an average was derived for the 15 fields. A supervised classifier software program in Matlab was developed to segment the carotid MR images to determine the maximal morphological component (calcium, fibrous cap, lipid pool, hemorrhage). Highest PET SUV of plaque was compared to NPRC readout on IHC and to primary plaque component on MRI. Medical records of patients who did not go to surgery (n=27) were retrospectively assessed for downstream cerebrovascular event.

Results: Highest uptake in the region of carotid stenotic plaque removed for CEA across subjects (n=15) showed strong correlation with IHC presence of NPRC, r= 0.90, p < 0.0001 and correlation with features of plaque vulnerability (r=0.62, p=0.01). NPRC presence on IHC was highest in the deep intima. Logistic regression analysis showed that SUV (Odds Ratio (OR) 5.4, 95% CI [1.2, 24.3], p=0.02) had a stronger relationship to plaque vulnerability than MRI (OR 1.3, 95% CI [0.99, 1.6], p=.05). In patients who did not go on to surgery, five (5) later had cerebrovascular events. Patients with events showed higher average PET uptake than those who did not (SUVmean, 2.3±1.15 vs. 1.6±1.05). Conclusions: We have translated a receptor-targeted PET radiotracer, ⁶⁴Cu-CANF-Comb, into human subjects and show that PET uptake correlates with plaque expression of the receptor it targets and with features of plaque vulnerability. Preliminary retrospective data suggests the potential of ⁶⁴Cu-CANF-Comb PET to predict patient outcomes.

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