Detection of Atherosclerotic Inflammation by 68Ga-DOTATATE PET Compared to [18F]FDG PET Imaging

Abstract

Background: Inflammation drives atherosclerotic plaque rupture. Although inflammation can be measured using fluorine-18-labeled fluorodeoxyglucose positron emission tomography ([¹⁸F]FDG PET), [¹⁸F]FDG lacks cell specificity, and coronary imaging is unreliable because of myocardial spillover.

Objectives: This study tested the efficacy of gallium-68-labeled DOTATATE (⁶⁸Ga-DOTATATE), a somatostatin receptor subtype-2 (SST₂)-binding PET tracer, for imaging atherosclerotic inflammation.

Methods: We confirmed ⁶⁸Ga-DOTATATE binding in macrophages and excised carotid plaques. ⁶⁸Ga-DOTATATE PET imaging was compared to [¹⁸F]FDG PET imaging in 42 patients with atherosclerosis.

Results: Target SSTR2 gene expression occurred exclusively in "proinflammatory" M1 macrophages, specific ⁶⁸Ga-DOTATATE ligand binding to SST₂ receptors occurred in CD68-positive macrophage-rich carotid plaque regions, and carotid SSTR2 mRNA was highly correlated with in vivo ⁶⁸Ga-DOTATATE PET signals (r = 0.89; 95% confidence interval [CI]: 0.28 to 0.99; p = 0.02). ⁶⁸Ga-DOTATATE mean of maximum tissue-to-blood ratios (mTBR_max) correctly identified culprit versus nonculprit arteries in patients with acute coronary syndrome (median difference: 0.69; interquartile range [IQR]: 0.22 to 1.15; p = 0.008) and transient ischemic attack/stroke (median difference: 0.13; IQR: 0.07 to 0.32; p = 0.003). ⁶⁸Ga-DOTATATE mTBR_max predicted high-risk coronary computed tomography features (receiver operating characteristics area under the curve [ROC AUC]: 0.86; 95% CI: 0.80 to 0.92; p < 0.0001), and correlated with Framingham risk score (r = 0.53; 95% CI: 0.32 to 0.69; p <0.0001) and [¹⁸F]FDG uptake (r = 0.73; 95% CI: 0.64 to 0.81; p < 0.0001). [¹⁸F]FDG mTBR_max differentiated culprit from nonculprit carotid lesions (median difference: 0.12; IQR: 0.0 to 0.23; p = 0.008) and high-risk from lower-risk coronary arteries (ROC AUC: 0.76; 95% CI: 0.62 to 0.91; p = 0.002); however, myocardial [¹⁸F]FDG spillover rendered coronary [¹⁸F]FDG scans uninterpretable in 27 patients (64%). Coronary ⁶⁸Ga-DOTATATE PET scans were readable in all patients.

Conclusions: We validated ⁶⁸Ga-DOTATATE PET as a novel marker of atherosclerotic inflammation and confirmed that ⁶⁸Ga-DOTATATE offers superior coronary imaging, excellent macrophage specificity, and better power to discriminate high-risk versus low-risk coronary lesions than [¹⁸F]FDG. (Vascular Inflammation Imaging Using Somatostatin Receptor Positron Emission Tomography [VISION]; NCT02021188).

Keywords: atherosclerosis; inflammation; macrophages; molecular imaging; positron emission tomography; somatostatin receptor.

Graphical abstract

Figure 1

The VISION Study Patient (A) and procedure (B) flowcharts. ∗Did not meet study criteria, n = 8; other clinical factors, n = 3; declined/cancelled, n = 49. †Coronary artery PET data excluded in ACS patients with ambiguous culprit arteries (n = 2). ‡Carotid artery PET data excluded in patients with prior carotid surgery (n = 2). §[¹⁸F]-FDG PET imaging not completed because of timing of surgery (n = 1). ‖Tissue samples excluded owing to insufficient mRNA extracted for quantitative PCR (n = 2). ¶CT scans not completed (calcium scan, n = 1; coronary angiogram, n = 5; carotid angiogram, n = 2). ACS = acute coronary syndrome; CT = computed tomography; CVD = cardiovascular disease; FDG = fluorodeoxyglucose; PCR = polymerase chain reaction; PET = positron emission tomography; TIA = transient ischemic attack; VISION = Vascular Inflammation imaging using Somatostatin receptor positron emissION tomography.

Figure 2

Target SSRT2 Expression in Proinflammatory Macrophages Heatmap of population-based RNA sequencing data (A) showing high SSTR2 expression in proinflammatory M1 macrophages (n = 4), very low levels of SSTR2 expression in unstimulated M0 macrophages (n = 4), and alternatively activated M2 macrophages (n = 5). For comparison, a heatmap of GLUT1 and GLUT3 shows significant gene expression in all cell types (note, different scales for SSRT and GLUT genes; mean values are log2 fragments per kilobase of transcript for million mapped reads [FPKM+1]). SSTR2 expression in LPS-stimulated macrophages from CVD patients versus age- and sex-matched healthy volunteers (n = 3 for both) using quantitative PCR (B). Photomicrograph shows green fluorescent immunoreactive SST₂ staining in macrophages (C), with blue nuclear DAPI-stained ([inset] concentration and isotype-matched IgG negative control). Brightfield photomicrograph shows brown immunoreactive SST₂-stained cultured macrophages, with nuclear counterstain (D). Phosphor autoradiographic image shows total binding of ⁶⁸Ga-DOTATATE (E) in clusters of cultured macrophages ([inset] parallel incubation with ⁶⁸Ga-DOTATATE and cold competing ligand showing very low levels of nonspecific binding). IgG = immunoglobulin G; LPS = lipopolysaccharide; other abbreviations as in Figure 1.

Figure 3

Coronary PET Inflammation Imaging: ACS Culprit Versus Bystander Lesions X-ray angiography images from a 59-year old man with ACS, showing a culprit first obtuse marginal lesion ([A] hatched oval) and nonculprit (bystander) right coronary artery disease ([E] circle). Identification of a culprit artery was aided by electrocardiographic findings of lateral T-wave inversion. Corresponding CT angiography images (B, F) show stented culprit lesion (*) and native bystander lesion with high-risk plaque morphology ([inset] low attenuation, cross-section of artery with outer wall boundary marked by dotted outline). In both lesions, intense inflammation (arrows) detected by ⁶⁸Ga-DOTATATE PET (C, G) is reproduced by [¹⁸F]FDG PET (D, H). Graphs of culprit versus highest nonculprit coronary ⁶⁸Ga-DOTATATE TBR_max values in patients (n = 8) with ACS and stented culprit ACS lesions (n = 6) versus stable stented (n = 18) lesions (I). ROC analysis demonstrates good diagnostic accuracy of ⁶⁸Ga-DOTATATE for culprit coronary lesions (J). Note stable stented lesions are coronary stents that were inserted >3 months prior to PET imaging in all but 1 patient. AUC = area under curve; mTBR_max = mean of maximum tissue-to-blood ratios; ROC = receiver operating characteristic; other abbreviations as in Figures 1 and 2.

Figure 4

Coronary PET Inflammation Imaging: High-Risk CT Features (A) X-ray and (D) CT coronary angiograms of a 67-year-old man with stable angina, showing minor LCx atheroma (hatched oval) with spotty calcification ([inset] *calcium scan) and calcified plaque in the LAD artery. Although ⁶⁸Ga-DOTATATE PET (B, E) allows unimpeded interpretation of inflammation in the LCx lesion (B, arrow), and lack of signal in the LAD, coronary [¹⁸F]FDG imaging is obscured by patchy myocardial tracer uptake (C). Graphs compare ⁶⁸Ga-DOTATATE (F) with [¹⁸F]FDG (G) coronary TBR_max values by CT plaque morphology in coronary segments (⁶⁸Ga-DOTATATE: NCP or MP, n = 86; normal, n = 45; spotty calcium, n = 30; large calcium, n = 72; LA or PR, n = 11; no high-risk CT, n = 186; [¹⁸F]FDG: NCP or MP, n = 43; normal, n = 13; spotty calcium, n = 15; large calcium n = 14; LA or PR, n = 4; no high-risk CT, n = 66), and ROC analysis demonstrating good diagnostic accuracy for high-risk coronary lesions. LA = low attenuation; LAD = left anterior descending; LCx = left circumflex; NCP = noncalcified plaque; MP = mixed plaque; PR = positive remodeling; other abbreviations in Figures 1, 2, and 3.

Figure 5

Carotid PET Inflammation Imaging: TIA/Stroke Views from a 66-year-old man ([top] axial plane) and a 70-year-old man ([bottom] sagittal plane), both of whom had TIAs resulting from right internal carotid artery lesions, shown on CT (A [hatched circle], D [∗]), with intense culprit plaque inflammation (hatched circles/arrows) detected by ⁶⁸Ga-DOTATATE (B, E) and reproduced by [¹⁸F]FDG (C, F). Graphs compare culprit versus nonculprit ⁶⁸Ga-DOTATATE (G) and [¹⁸F]FDG (H) TBR_max values (⁶⁸Ga-DOTATATE: culprit n = 14; contralateral n = 14; nonculprit [unstable CVD] n = 31; nonculprit [stable CVD] n = 24; contralateral [TIA/stroke] n = 14; diseased stable CVD n = 19; [¹⁸F]FDG: culprit n = 13; contralateral n = 13; nonculprit [unstable CVD] n = 31; nonculprit [stable CVD] n = 24; contralateral [TIA/stroke] n = 13; diseased [stable CVD] n = 19). TIA = transient ischemic attack; other abbreviations as in Figures 1, 2, and 3.

Figure 6

Vascular Inflammation Versus Clinical Risk Factors Graphs show correlations of vascular inflammation detected by ⁶⁸Ga-DOTATATE (A to C) and [¹⁸F]FDG (D) versus clinical cardiovascular disease risk factors. (Carotid arteries n = 62; aortas, n = 38; note data from patients not taking statins [n = 4] were excluded to control for this variable). Abbreviations as in Figure 1.

Figure 7

⁶⁸Ga-DOTATATE Versus [¹⁸F]FDG-Defined Inflammation and Other Clinical Factors Graphs show (A) the strong correlations among coronary, carotid, and aortic ⁶⁸Ga-DOTATATE mTBR_max versus [¹⁸F]FDG mTBR_max (n = 123 mean arterial values per tracer); (B) carotid ⁶⁸Ga-DOTATATE mTBR_max grouped by FRS (<8%, n = 16; 8% to 16%, n = 14; >16%, n = 32); (C) negative correlation of coronary aortic mTBR_max versus calcium score in patients with CAC <400 (n = 19); and (D) carotid ⁶⁸Ga-DOTATATE TBR_max ROI values in non TIA/stroke patients grouped by statin dosages (n = 20 patients [14 ROIs per artery]; low-dose n = 4; moderate dose, n = 9; high-dose, n = 7). Log transformed CAC values used to account for the non-normal distribution in the general population. Median difference between low-dose vs. high-dose: −0.13; (95% CI: −0.17 to −0.016); p = 0.02; %Δ −8.65. Low dose: simvastatin, 20 mg or lower; high-dose: atorvastatin, 80 mg or equivalent; moderate dose: all other dosages. CAC = coronary artery calcium score; FRS = %10-year Framingham risk score; ROI = region of interest; TIA = transient ischemic attack.

Figure 8

⁶⁸Ga-DOTATATE Ligand Binding to Macrophage SST₂ in Carotid Plaque In vivo CT angiography views of culprit carotid artery (hatched oval = internal carotid artery) in axial (A) and sagittal (E) views, with corresponding fused ⁶⁸Ga-DOTATATE PET-CT (B). Ex vivo views of macrographic images of the explanted carotid specimen (I, hatched line signifies location of carotid section); phosphor autoradiographic image shows the total binding of ⁶⁸Ga-DOTATATE to SST₂ receptors in macrophages within a transverse carotid section (C) corresponding to the level shown in clinical images. Adjacent section was incubated with ⁶⁸Ga-DOTATATE and cold competing ligand (D) showing very low levels of nonspecific binding. Brightfield photomicrographs show brown immunoreactive SST₂ staining (G, J, M) of macrophages identified with the panmacrophage marker CD68 (H, K, N), colocalized SST₂(brown), and CD68 (blue) staining in the same section (L); Movat’s pentachrome stain (F).

Figure 9

Carotid SSTR2/CD68 mRNA Versus In Vivo ⁶⁸Ga-DOTATATE Activity Graphs show correlations of SSTR2 versus CD68 mRNA within ex vivo carotid plaques measured by quantitative PCR (A); SSTR2(B) and CD68(C) mRNA versus corresponding in vivo ⁶⁸Ga-DOTATATE TBR_max values measured from clinical images (n = 6). Representative photomicrograph shows red SST₂ and green CD68 fluorescent immunoreactive staining of macrophages within carotid plaque (D), with blue nuclear DAPI staining. Note presence of both double positive (+) and double negative (−) staining indicating high cell specificity (E).

Central Illustration

Comparison Between ⁶⁸Ga-DOTATATE and [¹⁸F]FDG Coronary PET Inflammation Imaging Images from a 57-year old man with acute coronary syndrome who presented with deep anterolateral T-wave inversion (arrow) on electrocardiogram (A) and serum troponin-I concentration elevated at 4,650 ng/l (NR: <17 ng/l). Culprit left anterior descending artery stenosis (dashed oval) was identified by X-ray angiography (B). After the patient underwent percutaneous coronary stenting (C), residual coronary plaque (*inset) with high-risk morphology (low attenuation and spotty calcification) is evident on CT angiography (D, E). Use of ⁶⁸Ga-DOTATATE PET (F, H, I) clearly detected intense inflammation in this high-risk atherosclerotic plaque/distal portion of the stented culprit lesion (arrow) and recently infarcted myocardium (*). In contrast, using [¹⁸F]FDG PET (G, J), myocardial spillover completely obscures the coronary arteries. CT = computed tomography; [¹⁸F]FDG = fluorine-18-labeled fluorodeoxyglucose; ⁶⁸Ga-DOTATATE = gallium-68-labeled DOTATATE; PET = positron emission tomography.