Benefit of cardiac N-13 PET CFR for combined anatomical and functional diagnosis of ischemic coronary artery disease: a pilot study

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1 Ann Nucl Med (2014) 28: DOI /s y ORIGINAL ARTICLE Benefit of cardiac N-13 PET CFR for combined anatomical and functional diagnosis of ischemic coronary artery disease: a pilot study Srikara V. Peelukhana Hanan Kerr Kranthi K. Kolli Mariano Fernandez-Ulloa Myron Gerson Mohamed Effat Imran Arif Tarek Helmy Rupak Banerjee Received: 25 March 2014 / Accepted: 5 June 2014 / Published online: 21 June 2014 Ó The Japanese Society of Nuclear Medicine 2014 Abstract Background Cardiac positron emission tomography (PET) can lead to flow impairment quantification using PET coronary flow reserve (CFR p : ratio of stress flow to rest flow) and is superior to the current standard, singlephoton emission computed tomography. In this study, our first aim was to assess the benefit of CFR p in place of invasive CFR (CFR i ) by comparing the correlations of each of the indices with combined pressure and flow index CDP, and combined functional (pressure-flow) and anatomical (%area stenosis, %AS) index, LFC. The second aim was to test the correlation between CFR p and CFR i. Methods N-13 ammonia PET scans were performed and CFR p was obtained using a 1-compartment 2K-dynamic volume (DV)-constant kinetic model in Flowquant. During S. V. Peelukhana K. K. Kolli R. Banerjee (&) Department of Mechanical and Materials Engineering, University of Cincinnati, 598 Rhodes Hall, PO Box 0072, Cincinnati, OH , USA rupak.banerjee@uc.edu S. V. Peelukhana H. Kerr K. K. Kolli M. Fernandez-Ulloa M. Effat I. Arif T. Helmy R. Banerjee Veteran Affairs Medical Center, Cincinnati, OH, USA M. Fernandez-Ulloa Department of Nuclear Medicine, University of Cincinnati, Cincinnati, OH 45221, USA M. Gerson M. Effat I. Arif T. Helmy Division of Internal Medicine, Cardiology, University of Cincinnati, Cincinnati, OH 45221, USA R. Banerjee Department of Biomedical Engineering, University of Cincinnati, Cincinnati, OH 45221, USA catheterization, simultaneous pressure and flow readings were obtained in 10 vessels (three vessels in one patient, one vessel each in 7 patients) using a dual sensor tipped Combowire, and CFR i, CDP, LFC, and FFR were computed. %AS was obtained using quantitative coronary angiography. CDP was correlated with invasive pressure index (FFR) and CFR p and with FFR and CFR i. LFC was correlated with the %AS, FFR, and CFR p /CFR i, individually and in combination. Correlation analysis was done in SAS; p \ 0.05 was used for statistical significance. Results The correlations between CDP vs FFR and CFR p (r = 0.62, p = 0.19) in combination, as well as CDP vs FFR and CFR i in combination (r = 0.58, p = 0.24) remained similar. The correlation between LFC vs FFR, CFR p and %AS in combination improved (r = 0.82) with a near-significant p = 0.06, in comparison to the correlation between LFC vs FFR, CFR i and %AS in combination (r = 0.75, p = 0.15). CFR p correlated strongly and significantly (r = 0.82, p = 0.003) with CFR i, and the values were within 11 %. Conclusion The novelty of the PET procedure in this study is that the noninvasive CFR p can be used instead of invasive CFR i for the functional diagnosis of CAD. Therefore, a PET scan can reduce procedure time and cost while simplifying the diagnostic protocol for assessing coronary artery disease, thus benefitting both the patients and clinicians. Keywords Myocardial perfusion imaging Non-invasive stress test Coronary artery disease Anatomical and functional diagnosis Lesion flow coefficient FFR CFR Catheterization Abbreviations LAD Left anterior descending artery LCX Left circumflex artery

2 Ann Nucl Med (2014) 28: RCA LV EF CAD PET MPI MBF FFR CFR CDP LFC APV %AS CFR p CFR i Introduction Right coronary artery Left ventricle Ejection fraction Coronary artery disease Positron emission tomography Myocardial perfusion imaging Myocardial blood flow (ml/min/g) Fractional flow reserve Coronary flow reserve Pressure drop coefficient Lesion flow coefficient Average peak velocity (cm/s) %Area stenosis Noninvasive CFR obtained from PET scan Invasive CFR obtained using guidewire Standard assessment of myocardial perfusion in current clinical practice often involves either single-photon emission computed tomography (SPECT) or positron emission tomography (PET). Although SPECT myocardial perfusion imaging (MPI) is a useful noninvasive modality for detection of coronary artery disease (CAD), it is limited by issues including low attenuation artifacts, particularly in obese subjects and women, low-resolution, and the lack of quantification of the myocardial perfusion [1]. PET is currently the most robust technique to quantify perfusion noninvasively in the human heart and its use in cardiac imaging is rapidly increasing. Cardiac PET imaging has superior resolution, higher diagnostic accuracy than SPECT imaging, and can result in quantitative myocardial perfusion measures [2 5]. Moreover, in addition to being an accurate imaging modality, PET allows for the detection of CAD using noninvasive blood flow quantification [2, 3, 6 17]. Therefore, the PET MPI scan is superior to SPECT scan and can result in noninvasive quantification of flow impairment through PET-CFR (CFR p, subscript p: PET) [2, 3, 6 17]. Based on the results of MPI, invasive angiography maybe used to diagnose significant coronary epicardial stenosis. Therapeutic decision-making during coronary angiography is based on the visual assessment of the lesion and is often followed by evaluation of the functional severity of the stenosis. AHA/ACC guidelines suggest that a visually ambiguous stenosis may need to be further evaluated using the pressure-based fractional flow reserve (FFR: ratio of pressure distal to the stenosis to the pressure proximal to the stenosis [18 20]). It is a dimensionless parameter with a range from 0 (completely occluded artery) to 1 (normal artery), with a cut-off value of FFR \0.75 for a single-vessel stenosis and FFR \0.8 [18, 19] for multi-vessel stenoses indicating the likely benefit of percutaneous coronary intervention (PCI). Another parameter is the flow-based coronary flow reserve measured invasively using guidewire, CFR i, (subscript i: invasive) [21, 22], which is defined as the ratio of flow at hyperemia to the flow at rest. A CFR i value of is indicative of mild abnormality while a CFR i \2 is considered consistent with significant CAD [12]. The presence of concomitant microvascular disease, causing reduced hyperemia due to impaired microcirculation, leads to an increase in FFR values; this, in turn, results in an underestimation of epicardial disease. While CFR i can quantify the flow capacity of the entire coronary vasculature, it does not distinguish if a flow impediment is caused by the epicardial conduit or the microvascular bed. In view of the above shortcomings, two non-dimensional parameters based on fundamental fluid dynamics principles that can simultaneously distinguish between epicardial and microvascular disease were introduced. First one is the pressure drop coefficient (CDP) [23, 24], ratio of trans-stenotic pressure drop to distal dynamic pressure, which combines both pressure and flow measurements, and has a wider range of The second one is the lesion flow coefficient (LFC) [23, 24], ratio of %area stenosis (%AS) to the square root of CDP at the throat region (CDP m ), that combines both the anatomical and functional measurements. LFC is a normalized parameter with a range from 0 to 1, similar to FFR. Both these parameters are obtained from the pressure and flow measurements obtained invasively using a dual-sensor tipped guidewire. In a recent clinical study, CDP has been evaluated for clinical application [25]. In a meta-analysis study, clinical cut-off values of CDP for the delineation of epicardial and microvascular disease have been proposed [26]. In addition, both the parameters have been extensively validated in vitro [27 32] and in vivo [33 37] and shown to simultaneously distinguish the presence of epicardial stenosis with concomitant microvascular disease [23] in animal model. Both the CDP and LFC have also been shown to be independent of hemodynamic factors like HR and contractility in the presence of epicardial stenosis alone [33, 37] and in the presence of epicardial stenosis with concomitant microvascular disease [34, 36]. Further information about these parameters has been included in the discussion section. The prognostic value of CFR p has been assessed [2, 6, 9, 14, 17, 38, 39] and compared against FFR, and %diameter and area stenosis [6, 9]. CFR p obtained using N-13 ammonia [3, 7] has been validated in animal models [7, 16]. A few previous studies have correlated CFR p with intracoronary Doppler CFR [40 44]. However, a study

3 748 Ann Nucl Med (2014) 28: comparing the noninvasive CFR p and invasive CFR i with the functional (pressure and flow) parameter CDP and combined functional and anatomic parameter LFC in ischemic coronary arteries is currently lacking. Therefore, in this pilot clinical study, using N-13 ammonia as the radionuclide, PET MPI cardiac scans were used for noninvasive quantification of flow impairment by CFR p. Further, in the same patient group, the invasive diagnostic parameters, FFR, CFR i, CDP, and LFC were obtained. The objective of this study was to assess the benefits of PET MPI scan and the quantifiable CFR p in augmenting the anatomical and functional diagnosis of ischemic coronary artery disease. We hypothesize that the noninvasive CFR p can be used in place of the invasive CFR i. To test this hypothesis, our first aim was to compare the correlations of CFR p and CFR i with existing diagnostic parameters. In particular, we correlated the combined pressure and flow index, CDP vs FFR and CFR p ; and CDP vs FFR and CFR i. Similarly, we correlated the combined functional (pressureflow) and anatomical (%AS) index LFC vs FFR, CFR p, and %AS; and LFC vs FFR, CFR i, and %AS. The second aim was to test the correlation between CFR p and CFR i. Methods The Institutional Review Board (IRB) at the University of Cincinnati and the research and development (R&D) committee at the Cincinnati Veteran Affairs Medical Center (CVAMC) approved the protocol for this study. Patient population Inclusion criteria consisted of patients who were 18 years of age or above, symptomatic with acute chest pain and/or strong family history, and in need of a rest-stress perfusion scan. Excluded group of patients were those with LVEF \25 %, chronic kidney disease with baseline serum creatinine [2.5 gm/dl, history of type II heparin-induced thrombocytopenia, significant co-morbid conditions, pregnant women, and patients unable to comprehend the consent process. Further, patients with multiple stents, severe aortic stenosis, and coronary artery bypass grafts were also excluded. Justification of sample size The sample size in our study (n = 10) is similar to the previous studies, which had a sample size of 9 [43], 10 [40], and 11 [42]. Based on the study protocol, an explanation of the sample size used in our study is given below in sequential order along with the enrolment numbers (Fig. 1). PET MPI scan is not a standard-of-care procedure at the VA medical center. Due to this, we needed additional consenting steps to recruit patients for the PET scanning procedure. Based on the study inclusion/exclusion criteria, 34 patients were found eligible. As per steps detailed in Fig. 1, 19 out of the 34 patients were included while only 16 patients could successfully finish the rest/stress PET MPI scans. Subsequently, out of these 16 patients, 2 patients were not further evaluated using coronary angiography due to negative or low-risk test. Our protocol for coronary angiography consisted of standard-of-care invasive measurements. Hence, 6 patients with mild and severe stenoses out of the 14 patients were not further assessed using invasive measurements. The remaining 8 patients had visually intermediate lesions, which were assessed using invasive measurements. We had 10 data points for these 8 patients, where 3 data points were from one patient and 7 data points were from 7 patients. Patient background A summary of the characteristics of enrolled patients is provided in Table 1. All the patients were monitored for any adverse cardiac events during the stress part of the scan. Of the 8 patients enrolled, 4 patients were obese (BMI [30). No significant difference in heart rate or blood pressure was observed between the obese and the patients with normal weight (Table 2). One patient, P3, was on statin (40 mg/day, for 48 days), beta-blocker (25 mg/day, 80 days), calcium channel blocker (10 mg/day, 42 days), and nitrate (30 mg/day, 18 days). Another patient, P7, was on a low dose statin (40 mg/day, 70 days) prior to the study. These patients did not take the medication on the day of the scan. Compared to the patients without medications, no noticeable difference in the HR or blood pressure was observed in these two patients (Table 2). In addition, patient P1 had a heart attack (MI) 13 years prior to the study. At that time, he was treated with a balloon angioplasty and no infarcted myocardial tissue was observed during the angiography. Therefore, we believe that the flow data in this patient is reliable. PET MPI scan protocol All patients refrained from drinking coffee or any caffeinerelated products and medications 24 h prior to the scan. In addition, they were on fasting 8 h prior to the study. All patients underwent PET MPI scan according to the protocol using a whole-body CT-PET hybrid scanner (Discovery ST; GE healthcare) with a FOV of 41.9 cm with a left

4 Ann Nucl Med (2014) 28: Fig. 1 Flow-chart summarizing the diagnostic steps for patients offset of 4 cm. PET images were blurred to 6.4-mm FWHM in the transaxial direction using a Hanning filter for the reconstruction of data. For the rest scan, a low-voltage computerized tomography scan for attenuation correction (CTAC, 140 kv, 40 ma) was performed. A resting dose of ± 5.15 mci of N-13 ammonia was injected followed by an 18-min rest emission scan. After the completion of the rest scan, we waited for min for the radioactivity of the rest dose to decay, following which the stress agent, lexiscan/regadenoson, was administered as bolus dose of 0.4 mg/5 ml over 10 s followed by radionuclide injection. A stress dose of ± 7.65 mci of N-13 ammonia was injected and the stress emission scan was performed for 18 min, followed by a low-dose CTAC scan. Therefore, the minimum amount of time between the first injection of N-13 ammonia dosage and the second injection is 50 min (18 min of scan time? 30 min wait period? 2 min buffer time). The data were acquired in 2D, list mode and reconstructed using OSEM algorithm with 30 subsets and 2 iterations. The first 5 min of the data was reconstructed into 21 dynamic frames of S, S, S. Quantification of the myocardial blood flow using 1-compartment 2K-constant-dynamic volume (DV) kinetic model in the Flowquant Ò [14, 45] (Ottawa heart institute, Ottawa, CA, USA), is illustrated in Fig. 2. Cardiac catheterization and functional measurements Using both the visual analysis and 4-grade scoring system, final diagnosis of the perfusion abnormality was performed by an experienced nuclear cardiologist or a nuclear medicine physician as well as confirmed by a second expert reader. A reversible defect was defined as any reduction in the flow during stress images when compared to the flow in the same region during rest images. Based on the summed stress- and rest-scores, and the presence of reversible perfusion defects as indicated by PET MPI scan, patients were referred to coronary angiography. Using standard-of-care

5 750 Ann Nucl Med (2014) 28: catheterization procedures, the coronary arteries were visually assessed and ambiguous lesions were further evaluated using functional measurements at rest and at adenosine-induced maximal arterial dilatation (hyperemia). The aortic pressure (p a ) and ECG tracings were continuously recorded through Combomap Ò system (Volcano Therapeutics Inc., CA, USA). A Combowire with a flow sensor at its tip and pressure sensor at 1.5 cm offset was advanced into the artery of interest and equalized with p a before insertion into the artery. The Combowire was then advanced distal to the stenosis and the baseline pressure and average peak velocity (APV) readings were Table 1 Summary of the characteristics of the recruited patients Variable Sex (M/F) 7/1 Age (years) 63 ± 1.95 Body weight (Kgs) 89 ± 8.9 BMI 28 ± 2.45 PET ejection fraction (%) 55 ± 5 Clinical history Diabetes 5/8 Hypertension 8/8 Dyslipidemia 6/8 Smoking history 5/8 Family history of CAD 2/8 Myocardial infarction 1/8 Medications Statins (P3, P8) 2/8 Beta blockers (P3) 1/8 Calcium channel blockers (P3) 1/8 Nitrates (P3) 1/8 Arteries with functional readings a LAD 4 LCX 2 RCA 4 a One subject had readings from 3 arteries Study/group (mean ± SE) obtained on the Combomap machine. Using IV adenosine (140 lg/kg/min) as the stress-inducing agent, functional readings were obtained at hyperemia. The vessel diameter (D v ), diameter at the stenosis (D m ), and length of the lesion were obtained in the arteries through quantitative coronary angiography (QCA) of the angiograms, using automatic edge detection techniques available in the GE centricity software. The %AS was calculated based on these diameter values. Invasive parameters The pressure-based FFR [18, 20, 46] and flow-based CFR i [21, 22] were obtained from Combomap based on the following formula: FFR ¼ p d p a hyperemia and CFR i ¼ APV hyperemia APV baseline ; where p d is the pressure distal to the stenosis, p a is the pressure proximal to the stenosis, and APV is the average peak velocity obtained using the Combowire. CDP [23, 24, 47] is a functional parameter that combines flow and pressure, and is defined as Dp CDP ¼ 0:5 q APV 2 ; where Dp is the pressure drop across the stenosis, p a (aortic pressure) - p d (downstream pressure measured from Combowire), and q is the density of the blood (1.05 g/ cm 3 ). LFC [23, 24, 36] combines the lesion geometry (%AS, anatomical endpoint) and pressure and flow measurements (functional endpoints). It is defined as ð1 kþ or %AS LFC ¼ p ffiffiffiffiffiffiffiffiffiffiffiffi ; k ¼ A m A g CDP m A v A g where the numerator 1 - k is the %AS; in other words, k is the area ratio A m /A v, where A m and A v are area of the throat and vessel obtained from QCA (cm 2 ). On a similar note, ; Table 2 Summary of body mass index (BMI), heart rate (HR), blood pressure (BP), and rate-pressure product (RPP) for the enrolled patients ID BMI Rest readings Stress readings HR (bpm) BP (mmhg) RPP (mmhg bpm) HR (bpm) BP (mmhg) P / / P / / P / / P / / P / / P / / P / / P / / RPP (mmhg bpm)

6 Ann Nucl Med (2014) 28: Fig. 2 Figure showing the steps involved in the kinetic analysis of data. A representative case is shown for the stress images. a Reorientation of acquired images to register left ventricle (LV) in the three orthogonal slices and generation of an uptake image by summing the uptake frames. b LV sampling to determine the exact location of LV the CDP m in the denominator is. The APV 0:5 q APV 2 m m is calculated from the measured distal APV (cm/s). A g is the area of the guidewire (¼ p 4 0:35562 = mm 2 ). Data analysis Using the standard segmentation model for interpretation of nuclear images [48], the defect interpretation was assigned to one of the three arteries: LAD, LCX, and RCA. The defects obtained from PET imaging modality were then compared with the stenoses observed during coronary angiography. Representative polar maps for three patients showing regional defect quantification obtained using PET data and analyzed using Flowquant are shown in Fig. 3 (discussed more in later section). Figure 4 shows (a) the polar maps summarizing the CFR p values for mild stenosis (Fig. 4a) and intermediate Dp myocardium. c Polar map and a 3D model of the LV myocardium with superimposed tracer uptake. d Time activity curves obtained by applying the orientation and sampling to all the timeframes of the image sequence stenosis (Fig. 4d) in the left ventricle (LV), LAD, LCX, and RCA; (b) angiograms showing the QCA analyses for mild (56.14 %AS, Fig. 4b) and intermediate (77.67 %AS, Fig. 4e) stenoses; and (c) the invasive pressure, flow, FFR, and CFR readings obtained using the Combowire for the mild stenosis (Fig. 4c) and intermediate stenosis (Fig. 4f). Ten readings (n = 10; one vessel each in seven patients and three vessels in one patient) in abnormal coronary arteries were obtained during the coronary catheterization. CDP was correlated with CFR p and invasive pressure (FFR) in combination. Further, CDP was also correlated with FFR and CFR i in combination. Similarly, LFC was correlated with CFR p, FFR, and %AS in combination, further with CFR i, FFR, and %AS in combination. The CFR p values were correlated with CFR i values. The correlation analysis was performed using SAS (SAS institute, Cary, NC, USA). A value of p \ 0.05 was

7 752 Ann Nucl Med (2014) 28: Fig. 3 Regional defect quantification using kinetic analysis as obtained in the Flowquant for three representative cases. a Key to the interpretation of regional distribution defects, adopted from Johnson et al. [12]. b Patient 4. c Patient 5. d Patient 6 used for statistical significance. The strength of the correlation was given by the correlation coefficient (r value). All the values are reported as mean ± standard error (SE). Results A summary of the heart rate (HR), blood pressure (BP), and rate-pressure product (RPP: product of the HR and systolic BP) of the enrolled patients is given in Table 2. During the PET MPI scans, there was a significant (p \ 0.001) increase in the HR values from 66 beats per minute (bpm) at rest to 93 bpm at stress. The BP slightly decreased from 144 ± 6/ 70 ± 3 mmhg at rest to 137 ± 6/67 ± 3 mmhg at stress with an insignificant p value (=0.57). Further, the RPP increased significantly (p = 0.01) from 9595 bpm mmhg at rest to bpm mmhg at stress. During coronary angiography, out of the 24 arteries (8 patients 9 3 arteries), 14 arteries were stenosed and 10 arteries were normal. A comparison of PET defects vs coronary stenoses observed during angiography showed that out of the 14 stenosed arteries, PET agreed with angiographic stenoses in 12 arteries, an agreement of 87 %. Further, we have compared the rest myocardial blood flow (MBF), stress-mbf, and CFRp values between the 14 stenosed arteries and the 10 normal arteries (Fig. 5). The restmbf values showed an insignificant (p = 0.71) decrease from 0.81 ± 0.1 to 0.76 ± 0.09 ml/min/g between normal and stenosed arteries. For the normal arteries, the stress-mbf was 2.21 ± 0.24 ml/min/g, which decreased insignificantly (p = 0.22) to 1.82 ± 0.2 ml/min/g for the stenosed arteries.

8 Ann Nucl Med (2014) 28: Fig. 4 Representative images of mild stenosis. a CFR p values; b QCA analysis showing %AS; c pressure, flow, FFR, and CFR readings from Combomap. Similar images of intermediate stenosis d CFR p values; e QCA analysis showing %AS; f pressure, flow, FFR, and CFR readings from Combomap. All values are in mm On the other hand, there was a significant (p = 0.04) decrease in the CFR p values between the normal arteries (2.82 ± 0.13) and stenosed arteries (2.44 ± 0.11). Of the 14 vessels with defects observed during angiography, 10 vessels had invasive readings. A total of 4 vessels were not further assessed (1 vessel with chronic total occlusion, 1 vessel with serial lesions and 2 vessels with diffuse lesions). Table 3 provides a summary of the invasive parameters, FFR, CDP, LFC, the %AS obtained through QCA, invasive rest-, stress-apv values and CFR i, and the corresponding PET rest-, stress-mbf and CFR p (Fig. 4)values. A summary of correlations between CDP, LFC, and FFR with CFR p is given in Table 4. Correlations between CDP, LFC, and FFR with CFR i are provided in Table 5. Similar to a previous study by Werner et al. [49], FFR correlations with both the CFR p (Table 4) and CFR i (Table 5) were insignificant (high p values). Correlations of CDP, LFC with CFR p The functional parameter CDP had an insignificant correlation (r = 0.17, p = 0.63) with flow-based CFR p alone. When CDP was correlated with FFR and CFR p in combination, the r value increased to 0.62 and the p value improved to 0.19 (Table 4). The combined anatomical and functional parameter LFC, when correlated with CFR p alone, resulted in an insignificant (r = 0.43, p = 0.22) correlation. Further, when LFC was correlated in

9 754 Ann Nucl Med (2014) 28: Correlation of CFR p with CFR i Fig. 5 Bar graphs comparing the rest-myocardial blood flow (MBF), stress-mbf and CFR p values between the stenosed and normal arteries combination with %AS, FFR, and CFR p the r value increased to 0.82 with a near significance p value of 0.06 (Table 4). Correlations of CDP, LFC with CFR i CDP when correlated with CFR i alone had an insignificant correlation (r = 0.10, p = 0.80). When CDP was correlated with FFR and CFR i in combination, the r value remained similar at 0.58, with a p value of 0.24 (Table 5). The correlation of LFC with CFR i alone was insignificant (r = 0.02, p = 0.96). When LFC was correlated with %AS, FFR, and CFR i in combination, the r value increased to 0.75 and the p value improved to 0.15 (Table 5). The correlations of CDP with FFR and CFR i, LFC with %AS, FFR, and CFR i remained similar to the correlations of CDP with FFR and CFR p, LFC with %AS, FFR, and CFR p reported above. Though the strength of correlations (r values) were higher between CDP vs FFR and CFR p in combination and CDP vs FFR and CFR i in combination as well as for LFC vs FFR, CFR p and %AS in combination and LFC vs FFR, CFR i and %AS in combination, the p values were somewhat above 0.05 for these correlations. This is possibly due to the limited number of data points in this study. It is expected that the p values will be significant with an increase in the sample size as was the case in three previous invasive clinical studies [25, 26, 50]. Statistically significant linear correlations were reported between CDP vs FFR and CFR i, in a clincal study [25] with 27 patients (n = 27, r = 0.77, p = 0.01) as well as in a meta-analysis study [26] with 313 patients (n = 329, r = 0.82, p \ 0.001). Similarly, in a clinical study consisting of 20 patients [50], the LFC correlated significantly when FFR, CFR i, and %AS were combined (n = 23, r = 0.82, p \ 0.05). A comparison of the CFR p values and their corresponding CFR i values along with %difference is also presented in the Table 3. For P1 (P: patient), the CFR p values were 2.26, 2.05, and 2.39, in the LAD, LCX, and RCA, respectively. The corresponding CFR i values were 2.41, 2.19, and Values in RCA had the highest %difference of 11 % (%difference = PET CFR Invasive CFR PET CFR 100; Table 3). For P2, the CFR p and CFR i values were 2.17 and 2.24 in RCA, respectively, a %difference of 3 %. For P3, the CFR p (2.95) and CFR i values (2.93) in RCA were within 1 %. In case of P4, the CFR p (2.40) and CFR i (2.63) values were within 10 % in the RCA. The CFR p and CFR i values for the stenoses in the LAD matched well for P5 (2.00 vs 2.00), while they were within 1 % (2.85 vs 2.88) for P6. For P7 and P8, the CFR p and CFR i values in the LAD matched within 11 % (2.66 vs 2.38) and LCX (2.38 vs 2.41), respectively. Further, correlations between the PET rest- and stress- MBF values vs invasively obtained rest- and stress-apv values were performed. There was an insignificant correlation (r = 0.19, p = 0.61) between PET rest-mbf and invasive rest-apv values (Fig. 6a). There was a better correlation (r = 0.37) with an increased p value of 0.29 between the PET stress-mbf and invasive stress-apv values (Fig. 6a). More importantly, when the CFR p was correlated with the CFR i, there was a highly linear and significant correlation (r = 0.82, p = 0.003, Fig. 6b). Therefore, flow impairment quantification through CFR p had a significant linear correlation with the CFR i and the values matched within 11 % difference for this patient group. Discussion In this clinical study, we sought to test the benefits of PET MPI scan procedure and the quantifiable CFR p in augmenting the anatomical and functional diagnosis of ischemic coronary artery disease. Our main goal was to assess if the noninvasive CFR p can be used in place of the invasive CFR i. In particular, we correlated the combined pressure and flow index, CDP vs FFR and CFR i ; and CDP vs FFR and CFR p. Similarly, we correlated the combined functional (pressure-flow) and anatomical (%AS) index LFC vs FFR, CFR p, and %AS; and LFC vs FFR, CFR i, and %AS. Further, we also tested the correlation between CFR p and CFR i. The novelty of PET procedure in this study is that the noninvasive CFR p can be used instead of CFR i for the

10 Ann Nucl Med (2014) 28: Table 3 Summary of the PET rest- and stress-myocardial blood flow (MBF) values and CFR p, and the invasive rest- and stress-average peak velocity (APV) values, FFR, CFR i, CDP, and LFC PET readings Invasive readings %Variation in CFR Lesion flow coefficient (LFC) Pressure drop coefficient (CDP) %AS values Fractional flow reserve (FFR) S.no ID Defect territory values CFR i Stress APV (cm/s) CFR p Rest APV (cm/s) Stress MBF (ml/min/g) Rest MBF (ml/min/g) 1 P1a LAD P1b LCX P1c RCA P2 RCA P3 RCA P4 RCA P5 LAD P6 LAD P7 LAD P8 LCX Average values ± ± ± ± ± ± ± ± ± ± 0.1 Also shown are the %variations between CFR p and CFR i functional diagnosis of CAD. Consequently, a PET-scan can reduce the procedure time and cost while simplifying the diagnostic protocol for assessing CAD; thus benefitting both patients and the clinicians. In this way, PET MPI scan resulting in quantifiable CFR p can augment the functional and anatomical diagnosis of CAD. A discussion of our results along with the advantages of the functional parameter CDP and the combined functional and anatomical parameter LFC is presented below. Advantages of CDP and LFC The pressure drop (Dp)-flow characteristics in a stenosed vessel are non-linear. The Dp includes (a) viscous loss, a linear relationship of Dp and flow (or velocity), resulting from the friction between the blood flow and the lumen of the stenosis wall; and (b) loss due to the momentum change, a quadratic relationship of Dp and velocity, caused by the area change in the stenosis. In case of multiple or diffuse lesions or in the presence of concomitant microvascular disease, the complex interplay between the pressure and flow might not be sufficiently explained by FFR and CFR i or CFR p alone. This is because FFR is a pressure-based parameter, and CFR i is a flow-based parameter. On the other hand, CDP combines both pressure and flow, and LFC combines anatomical (%AS) and functional (pressure and flow) information. This explains the reason for the insignificant correlation of CDP and LFC with either CFR i or CFR p. In contrast, based on fluid dynamics principles, CDP showed an improved correlation when pressure-based FFR and flow based-cfr i /CFR p were combined in comparison to individual correlations. Similarly, LFC showed an improved correlation when %AS (anatomical index), FFR, and CFR i /CFR p (functional indices) were combined in comparison to the individual correlations. Therefore, these two parameters can account for the complex hemodynamic variations within the coronary artery. As mentioned previously, CDP has a wider range of In addition to accounting for the pressure drop in numerator, CDP has a squared velocity term in its denominator (APV 2 ). Due to this, CDP can account for the reduction in flow and has an increased resolving power for delineation of the concomitant epicardial and microcirculation disease. In the presence of epicardial stenosis alone, the value of CDP increases as the stenosis severity increases. Similarly, in the presence of microvascular disease alone, the value of CDP increases with an increase in the severity of microvascular impairment. In the presence of concomitant epicardial stenosis with microvascular disease, the value of CDP increases with increase in either the severity of the epicardial stenosis or microvascular impairment. These trends have been observed in a

11 756 Ann Nucl Med (2014) 28: Table 4 Correlations of CFR p with FFR, CDP, and LFC Parameter Equation r p FFR 0:02 CFR p þ 0: CDP 404:83 FFR þ 41:47 CFR p þ 257: * 0.19* LFC 0:68 FFR þ 0:18 CFR p þ 0:01 %AS 1: * 0.06* * The correlation when LFC and CDP, based on their definition, were correlated in combination with other parameters Table 5 Correlations of invasive parameters with CFR i Parameter Equation r p FFR 0:07 CFR i þ 0: CDP 405:55 FFR þ 11:1 CFR i þ 332: * 0.24* LFC 0:72 FFR þ 0:1 CFR i þ 0:01 %AS 1: * 0.15* * The correlation when LFC and CDP, based on their definition, were correlated in combination with other parameters recent clinical and meta-analyses studies by Kolli et al. [25, 26]. The futuristic parameter, LFC has a range from 0 to 1. A LFC value of zero represents no stenosis and values near one represent significant stenosis in the artery. In general, LFC value increases with increasing stenosis severity [50]. In the presence of microvascular disease alone, lower values of LFC are indicative of abnormal microcirculation, whereas elevated LFC values represent normal microcirculation. When there is a combination of epicardial and microvascular disease, for a fixed stenosis, the LFC value decreases with an increase in the severity of downstream microvascular impairment. For a fixed microvascular status, the LFC value increases as the epicardial disease severity increases. Such a trend was discussed in a previous animal study [23]. CDP and LFC in discordant FFR and CFR cases Fig. 6 Regression analysis between PET imaging modality and invasive readings. a PET rest-myocardial blood flow (MBF) vs invasive rest-average peak velocity (APV) and PET stress-mbf vs invasive stress-apv. b Noninvasive PET-CFR (CFR p ) vs invasive CFR (CFR i ) In a study comprising a combination of diffuse and focal lesions, Johnson et al. [12] have shown a moderate (r = 0.34) but significant correlation (p \ 0.001) between FFR and CFR i (n = 438). On the other hand, several studies, e.g., Werner et al. [49], showed a non-significant correlation between FFR and CFR i (r = 0.03, p = 0.87), similar to the values reported in our study. Therefore, with an increase in the sample size, it is expected that the statistical significance between the FFR and CFR p, CFR i will increase, as shown by Johnson et al. [12]. Johnson et al. [12] reported the concordance and discordance between FFR and CFR for different epicardial and microvascular conditions. Discordant values of FFR [0.8 and CFR \2.0 were observed in the presence of a

12 Ann Nucl Med (2014) 28: predominant diffuse stenosis compared to focal stenosis or in the presence of concomitant microvascular disease (Fig. 3 of Johnson et al. [12]). Similarly, values of FFR \0.8 and CFR [2 were attributed to the presence of primarily focal stenosis compared to diffuse stenosis. FFR and CFR, being pressure- and flow-based parameters, will need to have separate cut-offs for different vascular disease conditions. On the other hand, the functional (pressure and flow) parameter CDP, and the combined functional and anatomical parameter LFC, will be able to account for the complex pressure-flow variations using different cut-off values corresponding to each disease condition. Based on the established cut-off values of FFR and CFR for detecting significant epicardial stenosis and microvascular disease, four different cut-off values for CDP have been reported by Kolli et al. [26]. Using CDP, one can distinguish the matched FFR-CFR reduction and normal FFR- CFR values as well as mismatch FFR-CFR (normal-ffr and low-cfr; low-ffr and normal-cfr). Similar trend is expected for LFC. Therefore, based on these distinct cutoff values for various combinations of FFR and CFR, the fluid-dynamic based parameters, CDP and LFC, would better diagnose the discordant FFR and CFR cases in clinical practice. Using a single index, such as CDP or LFC, a cardiologist may get a comprehensive knowledge about the relative contribution of each of the epicardial and microvascular resistances. This will allow better diagnostics and therapeutic decision-making. CFR p vs CFR i The sample size in our study (n = 10) is similar to the sample size in previous studies by Kaufman et al. [40] (n = 10), Miller et al. [42](n = 11), and Shelton et al. [43] (n = 9). In our study, CFR p correlated significantly with CFR i. Except Kaufman et al. [40], all the previous studies [41 44] reported similar results showing a significant correlation between CFR p and CFR i. Further, there are a few notable differences between these previous studies and our study. Merlet et al. [41] and Shelton et al. [43] used a Doppler wire while Miller et al. [42] used a Doppler wire, as compared to the standard Doppler wire ( ) used in this study, for the measurement of CFR i. The presence of a bigger diameter guidewire leads to significant flow obstruction [51, 52]. While all the other studies used O-15 labeled water as a radiotracer, Stewart et al. [44] and our study used N-13 as a radiotracer. However, the subject group of Stewart et al. [44] consisted of patients with myocardial infarction whereas our subject group consisted of patients with ischemic coronary artery disease. Therefore, to the best of our knowledge, the present study is the first to report the concordance between CFR p and CFR i using N-13 in patients with ischemic coronary artery disease. MBF values In this study, we have compared the MBF values between arteries without stenosis and stenosed arteries in a patient group with significant risk factors indicating CAD (Fig. 5). The CFR p in the stenosed group significantly reduced to 2.4 ± 0.23 compared to arteries without stenosis (2.8 ± 0.23). However, arteries without stenosis were in the same patient group with significant risk factors. Therefore, the rest- and stress-mbf, and the CFR values in arteries without stenosis were compared with the previously published data in Table 1 (patients with risk-factors only, n = 3592) of Gould et al. [2]. The rest-mbf value in the present study was 0.81 ± 0.1 compared to the 0.85 ± 0.08, reported by Gould et al. [2], showing only 5 % difference. The stress-mbf values were within 2 % (2.21 ± 0.24 for present study vs 2.25 ± 1.07 by Gould et al. [2]). Similarly, the CFR values matched well within 1 % (2.82 ± 0.13 vs 2.80 ± 1.39). Limitations QCA In this study, QCA was used to obtain %AS. It is limited by the fact that a 3D vessel is being analyzed in a 2D plane. We performed blinded QCA measurements to reduce observer bias and used multiple frames to get average diameter and area. In future studies, the plan is to utilize the either IVUS or OCT technique to obtain better anatomical details. Radiotracer N-13 has been proven to have higher prognostic value in comparison to Rb-82 [8, 53]. N-13 ammonia is a cyclotron product and is not accessible at places without an onsite cyclotron, which might limit the usage of N-13 ammonia as a radionuclide. While N-13 ammonia is an excellent tracer, there might be a roll-off with ammonia uptake possibly not increasing linearly at high coronary blood flows that are achievable with pharmacologic coronary artery vasodilation. Hence, the myocardial perfusion quantitation using N-13 ammonia is expected to underestimate CFR p at high flow range. In our study, we have selectively assessed the patients in the intermediate stenoses range and correlated the CFR i vs CFR p. The regression line was obtained to explain the trend for this range of data only. Since a complete range of CFR values was not studied, we believe that an extrapolation of the present data is not appropriate.

13 758 Ann Nucl Med (2014) 28: We expect this trend to improve with an increased sample size and range. One may also note that the functional assessment of patients with visually moderate (\75 % areas obstruction) or severe ([95 % area obstruction) stenoses is not conducted since such a procedure is not typically a standard-of-care practice. Thus, it is often difficult to obtain the full range of CFR i values during catheterization. Sample size The sample size in our study (n = 10) is similar to the sample size in previous studies [40, 42, 43]. This study was primarily aimed as a proof of concept to show the feasibility of the use of a PET MPI scan and the quantifiable CFR p in place of CFR i for augmenting the anatomical and functional diagnosis of CAD. The plan is to improve upon this pilot study using a larger sample size for better statistical significance. Variations of CAD In this study, we have focused on discrete focal lesions, mainly in the intermediate stenosis range. However, the concordance of CFR p and CFR i needs to be tested in wider range of stenosis, complex and diffuse lesions, and particularly, in patients with known microvascular disease. Kinetic modeling All the analyses were performed using a 1-compartment 2K constant DV kinetic model. The results might vary somewhat based on the type of compartmental model used. However, the model we have chosen has been proven to have high reproducibility and reliability. Future directions We plan to test the concordance of CFR p and CFR i in a larger sample of patients. Alongside, we also plan to use CFR p and CFR i to obtain separate cut-off values for assessing the CDP and LFC to detect the epicardial stenosis and microvascular diseases. This can be achieved using a CFR p and CFR i cut-off value of 2 and an invasive FFR cutoff value of 0.8. Interestingly, there is a methodology to obtain the CDP and LFC using invasive pressure measurements and noninvasive PET MBF measurements (in ml/min/gm). In order to obtain the velocity values from the noninvasive MBF values, the myocardial mass (gm) and the vessel diameters of the individual arteries need to be known. The LV myocardial mass can be obtained through gated reconstruction of the PET images. Therefore, the myocardial mass in each of the coronary territories can be obtained by sub-dividing the LV myocardial mass in the three regions perfused by LAD, LCX, and RCA. The PET MBF measurements (ml/min/gm) when multiplied by the myocardial mass will provide the volumetric flow rate (ml/min). Once the volumetric flow rate (ml/min) in each of the territories is obtained using the above methodology, the vessel diameters of the coronary arteries can be obtained through QCA measurements. The velocity values (in cm/s) in each of the arteries can thus be obtained by dividing the volumetric flow rate values by area. Using these noninvasive velocity values and the invasive pressure measurements, CDP and LFC can be calculated. The values of CDP and LFC obtained in this alternate way are less invasive, cost-effective, and technically simplistic considering only one measurement (pressure) being invasive in nature. Such alternate way of determining the CDP and LFC needs to be tested in the future. However, this methodology has some assumptions and is applicable only to the large epicardial arteries, LAD, LCX, and RCA. Conclusion In this pilot clinical study, we performed N-13 ammonia PET MPI scans to non-invasively quantify the perfusion impairment using CFR p values. Our main goal was to assess if the noninvasive CFR p can be used in place of the invasive CFR i. In particular, we correlated the combined pressure and flow index, CDP vs FFR and CFR p ; and CDP vs FFR and CFR p. Similarly, we correlated the combined functional (pressure-flow) and anatomical (%AS) index LFC vs FFR, CFR p, and %AS; and LFC vs FFR, CFR i, and %AS. In addition, we correlated the CFR p and CFR i. The correlations between CDP vs FFR and CFR p (r = 0.62, p = 0.19) in combination, as well as CDP vs FFR and CFR i in combination (r = 0.73, p = 0.15) remained similar. The correlation between LFC vs FFR, CFR p and %AS in combination improved (r = 0.85) with a near-significant p = 0.06, in comparison to the correlation between LFC vs FFR and CFR i, %AS in combination (r = 0.75, p = 0.15). The flow impairment quantification through CFR p had a significant linear correlation (r = 0.85, p = 0.01) with the CFR i and the values matched within a %difference of 11 % in this patient group. The novelty of the PET procedure in this study is that the noninvasive CFR p can be used instead of invasive CFR i for the functional diagnosis of CAD. Consequently, a PET scan can reduce procedure time and cost while simplifying the diagnostic protocol for assessing CAD, thus benefitting both the patients and clinicians. In this way, PET MPI scan resulting in quantifiable CFR p can augment the functional and anatomical diagnosis of CAD.

14 Ann Nucl Med (2014) 28: Acknowledgments The authors would like to thank Kevin Fischio in the department of nuclear medicine, University medical center, Cincinnati, and Mike Grannen at the VA medical center for their help with the PET data acquisition and reconstruction. The authors are grateful towards Judy Hughes, all the technicians and nurses in the department of nuclear medicine at the UCMC, Gary Henry and Sherri Rosser at the VA, Ginger Conway, Rachel Mardis, Cindy Werner, and Cindy Mulcahy at the UCMC for help with the patient consent process. The authors would also like to thank all the nurses and technicians at the VA and UCMC catheterization laboratories for their support during the procedures. This work is funded by the VA Merit Review Grant (I01CX ), Department of Veteran Affairs. Conflict of interest The authors report no financial relationships or conflicts of interest regarding the content herein. References 1. 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