First-in-Human Studies of [¹⁸F]Fluorohydroxyphenethylguanidines: PET Radiotracers for Quantifying Regional Cardiac Sympathetic Nerve Density

Abstract

Background:

Disease-induced damage to cardiac autonomic nerve populations is associated with increased risk of sudden cardiac death (SCD). The extent of cardiac sympathetic denervation, assessed using planar scintigraphy or PET, has been shown to predict the risk of arrhythmic events in heart failure patients staged for implantable cardioverter defibrillator (ICD) therapy. The goal of this study was to perform first-in-human evaluations of 4-[¹⁸F]fluoro-meta-hydroxyphenethylguanidine ([¹⁸F]4F-MHPG) and 3-[¹⁸F]fluoro-para-hydroxyphenethylguanidine ([¹⁸F]3F-PHPG), two new PET radiotracers developed for quantifying regional cardiac sympathetic nerve density.

Methods and Results:

Cardiac PET studies with [¹⁸F]4F-MHPG and [¹⁸F]3F-PHPG were performed in normal subjects (n = 4 each) to assess their imaging properties and organ kinetics. Patlak graphical analysis of their myocardial kinetics was evaluated as a technique for generating nerve density metrics. Whole-body biodistribution studies (n = 4 each) were acquired and used to calculate human radiation dosimetry estimates. Patlak analysis proved to be and effective approach for quantifying regional nerve density. Using 960 left ventricular volumes of interest, across-subject Patlak slopes averaged 0.107 ± 0.010 mL/min/g for [¹⁸F]4F-MHPG and 0.116 ± 0.010 mL/min/g for [¹⁸F]3F-PHPG. Tracer uptake was highest in heart, liver, kidneys and salivary glands. Urinary excretion was the main elimination pathway.

Conclusions:

[¹⁸F]4F-MHPG and [¹⁸F]3F-PHPG each provide high quality PET images of the distribution of sympathetic nerves in human heart. Patlak analysis provides reproducible measurements of regional cardiac sympathetic nerve density at high spatial resolution. Further studies of these tracers in heart failure patients will be performed to identify the best agent for clinical development.

Clinical Trial Registration:

https://clinicaltrials.gov; Unique identifier: NCT02385877.

Introduction

Scintigraphic imaging of cardiac sympathetic innervation is an established noninvasive approach to assessing risk of sudden cardiac death (SCD) in patients with heart failure and may be used to balance risks and benefits of implantable cardioverter defibrilator (ICD) therapy. Evidence supporting this assertion has been building for almost 20 years, starting with small scale trials using the sympathetic nerve imaging agent [¹²³I]meta-iodobenzylguanidine ([¹²³I]mIBG).^1–3 Early studies with [¹²³I]mIBG demonstrated that a single index of global cardiac sympathetic denervation, the heart-to-mediastinum ratio (H/M), was able to separate heart failure patients with high risk of SCD from those with low risk. This finding has been remarkably consistent in subsequent trials with [¹²³I]mIBG in heart failure patients, using either planar scintigraphy or more advanced assessments with SPECT imaging, including the multicenter ADMIRE-HF trial.^4,5 ADMIRE-HF confirmed that the H/M ratio of [¹²³I]mIBG is an independent predictor of SCD in heart failure patients, and can be used to guide appropriate ICD therapy.⁶

Further confirmation that regional cardiac sympathetic denervation promotes malignant arrhythmias comes from the recent PAREPET trial.^7,8 This study used [¹¹C]-(–)-meta-hydroxyephredrine ([¹¹C]HED) and PET to generate measures of the extent of regional cardiac sympathetic denervation in more than 200 heart failure patients staged for ICD placement. In addition, resting perfusion was measured using [¹³N]ammonia and myocardial viability was assessed using [¹⁸F]fludeoxyglucose ([¹⁸F]FDG. Among all imaging parameters, the extent of denervated myocardium (as defined by [¹¹C]HED retention) was found to be the strongest predictor of potentially fatal arrhythmic events. Subjects with denervation extent measures exceeding 37.4% of the left ventricle defined the upper tertile of patients at highest risk of sudden cardiac arrest. These results strongly suggest that PET-derived metrics of the regional extent of cardiac denervation provide an even stronger predictor of SCD than the global H/M ratio of [¹²³I]mIBG. However, the short half-life of carbon-11 (20.4 min) limits use of [¹¹C]HED to academic hospitals with an on-site cyclotron and radiochemistry facility.

Two new fluorine-18 labeled sympathetic innervation radiotracers, 4-[¹⁸F]fluoro-meta-hydroxyphenethylguanidine ([¹⁸F]4F-MHPG) and its structural isomer 3-[¹⁸F]fluoro-para-hydroxyphenethylguanidine ([¹⁸F]3F-PHPG), have been developed in our laboratory (Figure 1). In addition to the benefits of the longer half-life of fluorine-18 (109.8 min), these tracers were designed to have slower neuronal uptake rates than [¹²³I]mIBG and [¹¹C]HED, and irreversible neuronal retention through efficient trapping inside norepinephrine storage vesicles. We hypothesized that this combination of kinetic properties would make measures of their cardiac retention more sensitive in detecting the mild to moderate nerve losses that occur early in the progression of disease. In addition, their kinetics could be analyzed using tracer kinetic analysis methods previously applied to other irreversible cardiac radiotracers, such as [¹³N]ammonia and [¹⁸F]FDG.^9,10 Our goal was to develop a fluorine-18 labeled sympathetic nerve tracer that could accurately quantify the extent and severity of cardiac sympathetic denervation. Preclinical tests with [¹⁸F]4F-MHPG and [¹⁸F]3F-PHPG in non-human primates were highly promising, including successful kinetic analysis results using Patlak graphical analysis, supporting the clinical translation of these radiotracers.^11,12

Figure 1.

Chemical structures of [¹⁸F]4F-MHPG and [¹⁸F]3F-PHPG.

The purpose of this study was to perform first-in-human assessments of [¹⁸F]4F-MHPG and [¹⁸F]3F-PHPG as cardiac PET radiotracers, including their imaging properties, metabolism, and kinetics in heart, liver and blood. Patlak graphical analysis of the myocardial kinetics of [¹⁸F]4F-MHPG and [¹⁸F]3F-PHPG was evaluated as an approach to quantifying regional nerve density. Also, whole-body PET scans were performed to collect the tissue biodistribution data required to calculate human radiation dosimetry estimates. The results show that [¹⁸F]4F-MHPG and [¹⁸F]3F-PHPG each provide high quality cardiac PET images, and Patlak analysis of their myocardial kinetics provides reproducible quantitative metrics of regional sympathetic nerve density.

Methods

The data, analytic methods, and study materials for this study are available from the corresponding author upon reasonable request, contingent on compliance with applicable HIPAA requirements and appropriate IRB approval.

Study Design

Two series of PET studies were performed to compare [¹⁸F]4F-MHPG and [¹⁸F]3F-PHPG as cardiac imaging agents. Dynamic cardiac PET studies assessed the cardiac imaging properties of the two tracers and their ability to provide quantitative metrics of regional nerve density using kinetic analysis methods. Whole-body PET studies determined the tissue distribution and organ clearance kinetics of the tracers, data needed to generate human radiation dosimetry estimates. For the cardiac PET studies, 8 healthy subjects (4 females, 4 males, 29 ± 9 y old, range 21–49 y) were recruited. Four subjects (2 female, 2 male) were scanned with [¹⁸F]4F-MHPG, the other four (2 female, 2 male) with [¹⁸F]3F-PHPG. Eight healthy subjects (4 females, 4 males, 22 ± 3 y old, range 19–28 y) completed whole-body PET studies, with each tracer studied in 2 female and 2 male subjects. Subjects were screened in an initial clinic visit to exclude subjects with occult cardiovascular disease. Screening tests included: medical history, vital signs, body weight and height, BMI, inventory of concomitant medications and evaluation of a resting supine 12-lead ECG. Blood tests included: comprehensive metabolic panel, lipid profile and hemoglobin A1c levels. Inclusion criteria included: age 18–55y, non-obese (BMI < 30), normal blood pressure, no history of cardiovascular disease, and normal ECG. Exclusion criteria included: risk factors for coronary artery disease (age > 55y, hypertension, smoking, hypercholesterolemia, diabetes, obesity), history of coronary ischemic events (infarction, atrial fibrillation, ventricular tachycardia, or exertional angina), pregnancy or lactation, claustrophobia, and medications affecting sympathetic nerves (tricyclic antidepressants, phenylephrine, etc.). The study protocol was approved by the University of Michigan Institutional Review Board. Signed informed consent statements were obtained from all participants prior to enrollment. This study was performed under an exploratory Investigational New Drug (IND) clearance from the U.S. FDA, and was registered at the ClinicalTrials.gov website (NCT02385877).

Radiochemistry

[¹⁸F]4F-MHPG and [¹⁸F]3F-PHPG were prepared as previously described.¹² Specific activities were 46 ± 20 GBq/μmol and 84 ± 27 GBq/μmol for [¹⁸F]4F-MHPG and [¹⁸F]3F-PHPG, respectively. Injected doses of [¹⁸F]4F-MHPG averaged 254 ± 9 MBq, containing an average mass of 1.8 ± 0.9 μg (range 1.2 – 3.9 μg). [¹⁸F]3F-PHPG injected doses averaged 248 ± 23 MBq, with an average mass of 0.8 ± 0.1 μg (range 0.5 – 0.9 μg).

Dynamic Cardiac PET Studies

A Siemens ECAT Exact HR+ PET camera was used for all studies. For the cardiac PET studies, a 90-min dynamic sequence (12×10 s, 2×30 s, 2×60 s, 2×150 s, 2×300 s, and 7×600 s) was acquired after i.v. injection of [¹⁸F]4F-MHPG or [¹⁸F]3F-PHPG. Images were reconstructed using iterative OSEM (4 iterations, 8 subsets), zoom = 2.2 and 7-mm Gaussian filter. Six venous blood samples (2–3 mL) were drawn during each scan to assess radiotracer metabolism in plasma and activity partitioning between plasma and red blood cells.

Image Processing

Transaxial PET images were reoriented into orthogonal views, the short-axis images interpolated into 4 mm slices and saved using the PMOD image analysis package (PMOD Technologies). These data were analyzed using a custom tracer kinetic analysis software package written in IDL (Harris Geospatial Solutions). On each of 16 short-axis slices, encompassing the left ventricle, an algorithm defined 60 angular sectors, centered on the left ventricular wall, generating 16 × 60 = 960 volumes of interest (VOIs). Time-activity curves for tissue, C_t(t), were generated from the dynamic PET data for each VOI. Another VOI was placed in the left ventricular blood pool at the base of the heart to extract a whole-blood time-activity curve, C_wb(t).

Input Functions

Blood samples were processed into purified plasma and analyzed on an HPLC system with an in-line radiation detector to determine the fraction of plasma activity associated with intact parent radiotracer, f_intact(t).^11,12 Whole-blood and plasma aliquots were counted in a gamma counter to determine the ratio of activity concentrations in plasma and whole-blood, C_p/C_wb. These data were used to convert the whole-blood curve C_wb(t) to an intact tracer in plasma curve, C_p(t), as: C_p(t) = C_wb(t)·f_intact(t)·[C_p/C_wb]. Plasma curves were used as input functions for Patlak analysis.

Tracer Kinetic Analysis

Patlak graphical analysis was used to analyze the myocardial kinetics of [¹⁸F]4F-MHPG and [¹⁸F]3F-PHPG. For several subjects, patient motion during the last 30 min caused artifacts in the time-activity data. In light of this, and considering that clinical studies with these tracers are unlikely to exceed 40–60 min, kinetic analyses used only the first 60 min of data. For Patlak analysis, the regional tissue time-activity curve C_t(t) and the plasma time-activity curve C_p(t) undergo mathematical transformations to generate a ‘Patlak plot’ of the transformed data which has a distinct linear phase.¹³ Linear regression of the linear portion of the plot provides a Patlak slope, K_p (mL/min/g), which is a measure of the rate of irreversible tracer uptake into sympathetic nerve terminals. Patlak analysis was performed using the myocardial kinetics of each VOI to determine regional values of the Patlak slope K_p, the y-intercept, and the linear regression coefficient R.

Biodistribution PET Studies and Radiation Dosimetry Estimates

Whole-body PET scans were acquired at four times, starting at: 5, 60, 150 and 360 min after tracer injection. In some cases, the last two scans started at different times due to competing clinical responsibilities of the PET technologists. Human radiation dosimetry estimates were calculated using OLINDA/EXM Version 2.0 (Vanderbilt University, Hermes Medical Solutions) following published guidelines^14–16. Briefly, whole-body PET scans were analyzed using the PMOD image analysis software suite to determine the total activity (kBq) in each organ of interest. Decay-corrected activity data were normalized to the total radiotracer dose injected (kBq) to determine the % injected dose (%ID) as a function of time for each organ. Biokinetic data for each organ were fit to a single exponential or bi-exponential clearance model using nonlinear regression analysis (GraphPad Prism, GraphPad Software). Uptake fractions and clearance half-times for each organ analyzed were entered into OLINDA/EXM 2.0 to determine the number of distintegrations in the organ N_i (MBq-h/MBq). In a few cases where tracer accumulation was evident in the early organ kinetics (e.g. salivary glands), it was necessary to take an interpolated curve fit to the data and apply fluorine-18 decay factors to estimate the effective organ kinetics. This curve was then numerically integrated to directly determine N_i (GraphPad Prism). The fraction of the injected dose eliminated by urinary excretion and the half-time for elimination were estimated from kinetic data for the bladder. The maximum fraction of the injected dose measured in the gastrointestinal tract was entered into the ICRP Human Alimentary Tract (HAT) model incorporated into OLINDA/EXM 2.0, assuming the activity entered the small intestine. Each subject’s biokinetic data were used to calculate radiation dosimetry estimates for the ICRP 89 Adult Male and Adult Female models. Dose estimates were averaged for each gender (n = 4 each). The mean data for each gender were then averaged following ICRP 128 guidelines to obtain the final table of sex-averaged dose estimates. These values were multiplied by the appropriate tissue weighting factors and summed to determine the effective dose.

Safety and Tolerability Tests

Tests were performed to evaluate the safety of [¹⁸F]4F-MHPG and [¹⁸F]3F-PHPG. Before and after each PET session, resting supine 12-lead ECG, body temperature, heart rate, and blood pressure were recorded, and blood and urine samples were collected. Laboratory tests included a comprehensive metabolic panel, complete blood count, plasma catecholamine levels, and urinalysis. For 90 min after radiotracer administration, heart rate and peripheral capillary oxygen saturation (SpO₂) were continuously monitored, and blood pressure was measured every 10 min. Female subjects took a urine pregnancy test before the PET session. Subjects were contacted 24 h and 30 d after the PET session to enquire if they had experienced any adverse events related to the study.

Statistical Analyses

All presented data are mean ± SD. Reported coefficients of variation (CV) were calculated as CV = (SD/mean)×100%.

Results

Safety

Intravenous administrations of [¹⁸F]4F-MHPG and [¹⁸F]3F-PHPG were well-tolerated by all subjects, causing no significant changes in hemodynamic measures, ECG data, vital signs, or laboratory measures. No adverse events or detectable pharmacological effects were observed in any of the 16 subjects.

Dynamic Cardiac PET Studies

[¹⁸F]4F-MHPG and [¹⁸F]3F-PHPG provide high quality images of the regional distribution of sympathetic nerve terminals in the heart (Figure 2). The myocardial kinetics of the two tracers are consistent with irreversible uptake and retention in sympathetic neurons (Figure 3). Neuronal accumulation of [¹⁸F]4F-MHPG is typically complete within 10 min, while neuronal levels of [¹⁸F]3F-PHPG climb for 30 min or more. The more prolonged neuronal uptake of [¹⁸F]3F-PHPG contributes to its higher heart-to-blood contrast late in the study (5.3 ± 0.8 vs. 3.7 ± 0.6 for [¹⁸F]4F-MHPG). Liver uptake of the tracers is similar early, but [¹⁸F]4F-MHPG clears more quickly than [¹⁸F]3F-PHPG, affording better heart-to-liver contrast (2.4 ± 0.4 vs. 1.2 ± 0.4, respectively). For both tracers, lung uptake was extremely low.

Figure 2.

Representative short axis (SA), horizontal long axis (HLA) and vertical long axis (VLA) images of [¹⁸F]4F-MHPG (top row) and [¹⁸F]3F-PHPG (bottom row).

Figure 3.

Kinetics of [¹⁸F]4F-MHPG (top) and [¹⁸F]3F-PHPG (bottom) in blood, liver and heart.

Blood Sample Analyses

Metabolism of [¹⁸F]4F-MHPG and [¹⁸F]3F-PHPG in plasma is biphasic (Figure 4). [¹⁸F]4F-MHPG is metabolized more quickly than [¹⁸F]3F-PHPG. The slower metabolism of [¹⁸F]3F-PHPG explains why it accumulates into sympathetic nerves for a longer time than [¹⁸F]4F-MHPG. Each tracer is metabolized into one major metabolite that is more polar than the parent compound (data not shown). Based on HPLC retention times, the metabolites formed in humans appear to be the same ones detected in preclinical PET studies in non-human primates.^11,12 [¹⁸F]4F-MHPG is metabolized by sulfate conjugation at the meta-hydroxyl position.¹¹ The metabolite of [¹⁸F]3F-PHPG remains unknown, but prior tests suggest that sulfate conjugation and glucuronidation are unlikely pathways.¹² Since highly linear Patlak plots were obtained in all subjects using plasma curves that excluded the [¹⁸F]3F-PHPG radiometabolite, this strongly suggests that the radiometabolite is not transported into cardiac sympathetic nerves. Activity concentration ratios in plasma over whole-blood, C_p(t)/C_wb(t), were consistent in each study. Mean values of C_p(t)/C_wb(t) ranged from 1.29 ± 0.14 to 1.38 ± 0.12 for [¹⁸F]4F-MHPG, and from 1.28 ± 0.06 to 1.45 ± 0.14 for [¹⁸F]3F-PHPG. Across subjects, C_p/C_wb averaged 1.38 ± 0.12 for [¹⁸F]4F-MHPG and 1.35 ± 0.12 for [¹⁸F]3F-PHPG. Since C_p/C_wb was nearly constant during a study, the mean ratio for each subject was used to estimate their input function.

Figure 4.

Metabolic breakdown of [¹⁸F]4F-MHPG (top) and [¹⁸F]3F-PHPG in plasma (n = 4 each). Mean ± SD of the times at which 50% of parent tracer was still intact (T_50%) are shown.

Tracer Kinetic Analysis

Patlak graphical analysis of the kinetics of [¹⁸F]4F-MHPG and [¹⁸F]3F-PHPG provided very consistent results (Table 1). Patlak plots and corresponding linear regression lines for a few VOI from an [¹⁸F]4F-MHPG subject are shown in Figure 5. Regional Patlak plot data were highly linear (R > 0.97 in all VOI). Measured Patlak slopes K_p for individual subjects had coefficients of variation (CV) values ranging from 8.8% to 16.1%. The highest CV of 16.1% for the first subject studied with [¹⁸F]4F-MHPG (4F-S1) was likely related to patient motion starting around 20 min into the PET scan causing a partial mismatch between the attenuation correction map derived from the transmission data and the scan emission data. Averaging the mean Patlak slopes of subjects within groups, the inter-individual CV values were about 9% for each tracer. Representative examples of regional Patlak slopes of [¹⁸F]4F-MHPG and [¹⁸F]3F-PHPG, presented in polar map format, are shown in Figure 6.

Table 1.

Patlak Graphical Analysis Results

[18F]4F-MHPG				[18F]3F-PHPG
Subject	Kp (mL/min/g)	CV (%)	R	Subject	Kp (mL/min/g)	CV (%)	R
4F-S1	0.120 ± 0.019	16.1%	0.987 ± 0.009	3F-S1	0.108 ± 0.011	10.4%	0.998 ± 0.002
4F-S2	0.114 ± 0.011	9.8%	0.999 ± 0.001	3F-S2	0.107 ± 0.009	8.8%	0.998 ± 0.001
4F-S3	0.096 ± 0.010	9.9%	0.995 ± 0.003	3F-S3	0.123 ± 0.014	11.0%	0.997 ± 0.003
4F-S4	0.111 ± 0.015	13.4%	0.999 ± 0.001	3F-S4	0.126 ± 0.015	11.8%	0.999 ± 0.001
Mean ± SD:	0.107 ± 0.010	9.0%	‒	Mean ± SD:	0.116 ± 0.010	8.7%	‒

Values are mean ± SD. K_p indicates Patlak slope, CV, coefficient of variation, R, linear regression coefficient.

Figure 5.

Examples of Patlak plots and linear regression fits for [¹⁸F]4F-MHPG from six volumes-of-interest in a single normal subject (#4F-S4).

Figure 6.

Polar maps of regional Patlak slopes for [¹⁸F]4F-MHPG (top) and [¹⁸F]3F-PHPG (bottom). Averages of the Patlak slopes for all 960 volumes of interest (mean ± SD) are shown. Interpolation between adjacent VOIs and adjacent short axis slices was performed to smooth the map.

Radiation Dosimetry Estimates

Maximum intensity projection (MIP) images of whole-body PET scans are presented in Figure 7. Each tracer exhibited high uptake and long retention times in the heart. The fraction of the injected dose (%ID) in the heart early after injection averaged 1.74% ± 0.09% for [¹⁸F]4F-MHPG and 2.02% ± 0.09% for [¹⁸F]3F-PHPG. Clearance half-times from the heart were very slow, averaging 40.5 ± 7.5 h (range 36.0 – 51.8 h) and 50.2 ± 7.0 h (range 44.1 – 57.7 h) for [¹⁸F]4F-MHPG and [¹⁸F]3F-PHPG, respectively. To determine the number of fluorine-18 disintegrations N_i (MBq-h/MBq) in organs of interest, VOI analysis was performed for brain, heart, liver, kidneys, pancreas, urinary bladder, intestines, salivary glands, thyroid gland and prostate gland (male subjects). Gallbladder kinetics were measured for [¹⁸F]4F-MHPG, but no gallbladder uptake was seen for [¹⁸F]3F-PHPG. Urinary excretion was the main elimination pathway. The %ID excreted in the urine averaged 31.9% ± 4.3% for [¹⁸F]4F-MHPG and 28.9% ± 3.6% for [¹⁸F]3F-PHPG. The maximum %ID excreted in the gastrointestinal tract was 4.5% ± 1.0% for [¹⁸F]4F-MHPG. Excretion of [¹⁸F]3F-PHPG in the GI tract was much lower (maximum %ID = 0.4% ± 0.1%). Adult organ equivalent doses and effective dose estimates generated with the biokinetic data and OLINDA/EXM 2.0 are shown in Table 2.

Figure 7.

Representative whole-body images of [¹⁸F]4F-MHPG (top) and [¹⁸F]3F-PHPG (bottom). Mid-times of scan acquisition times are shown. In the first [¹⁸F]4F-MHPG image, the circular lesion in the liver was later established by MRI to be a focal nodular hyperplasia in this 22-year old female subject.

Table 2.

Adult Radiation Dose Estimates

	Absorbed dose per unit activity administered (mSv/MBq)
Site	[18F]4F-MHPG	[18F]3F-PHPG
Adrenals	1.45E-02	1.61E-02
Brain	3.53E-03	4.06E-03
Breasts	9.82E-03	1.02E-02
Esophagus	1.16E-02	1.29E-02
Eyes	8.51E-03	8.73E-03
Gallbladder Wall	1.78E-02	1.64E-02
Left colon	2.14E-02	1.45E-02
Small Intestine	3.45E-02	1.57E-02
Stomach Wall	1.31E-02	1.37E-02
Right colon	3.63E-02	1.53E-02
Rectum	2.49E-02	2.26E-02
Heart Wall	3.32E-02	3.83E-02
Kidneys	2.47E-02	2.11E-02
Liver	1.57E-02	3.12E-02
Lungs	1.12E-02	1.21E-02
Ovaries	2.14E-02	2.00E-02
Pancreas	2.32E-02	2.87E-02
Prostate	4.03E-02	3.47E-02
Salivary Glands	1.92E-02	3.40E-02
Red Marrow	1.14E-02	1.15E-02
Osteogenic Cells	9.37E-03	9.52E-03
Spleen	1.26E-02	1.24E-02
Testes	1.13E-02	1.12E-02
Thymus	1.20E-02	1.26E-02
Thyroid	1.57E-02	1.96E-02
Urinary Bladder Wall	1.75E-01	1.65E-01
Uterus	3.08E-02	2.87E-02
Total Body	1.26E-02	1.26E-02
Effective Dose	2.11E-02	2.03E-02

Discussion

This report describes first-in-human PET studies of [¹⁸F]4F-MHPG and [¹⁸F]3F-PHPG, two new cardiac sympathetic nerve radiotracers. The results demonstrate that administration of these tracers is safe, their biodistribution is consistent with localization in peripheral tissues with dense sympathetic innervation, including the heart, and analysis of their myocardial kinetics can provide robust regional metrics of cardiac sympathetic nerve density.

An advantage of the irreversible kinetics of [¹⁸F]4F-MHPG and [¹⁸F]3F-PHPG is the ability to use Patlak graphical analysis, a method developed for radiotracers with irreversible tissue kinetics. Since this approach uses linear regression of the Patlak plot data to estimate only two parameters (slope and intercept), it is possible to use smaller VOIs than would typically be used for other tracer kinetic analysis approaches, such as compartmental modeling. Although smaller VOIs have greater levels of statistical noise in the myocardial kinetic data, the Patlak approach can obtain robust regional nerve density measures at high spatial resolution. The Patlak slope is a measure of the net uptake or influx of tracer into irreversible tissue compartments, in this case, into the storage vesicles in sympathetic nerve terminals. Thus the regional value of the Patlak slope provides a useful quantitative metric of regional sympathetic nerve density. Our results showed that Patlak slopes K_p had low coefficients of variations within subjects, as well as across subjects. This finding suggests that normal control databases of regional Patlak slopes can be used to quantitatively assess regional nerve losses in patients with heart diseases.

The radiation dosimetry estimates of [¹⁸F]4F-MHPG and [¹⁸F]3F-PHPG in Table 2 show that the urinary bladder wall is the ‘critical organ’, the organ receiving the highest radiation dose for these radiotracers. Comparing the dosimetry of [¹⁸F]4F-MHPG and [¹⁸F]3F-PHPG with recent [¹⁸F]FDG dose estimates by Stabin and Siegel¹⁵ calculated using OLINDA/EXM 2.0, most of the organ doses are similar, with a few exceptions. Doses to the heart wall are about 50% lower than the heart wall dose for [¹⁸F]FDG, brain doses are only 10% of the [¹⁸F]FDG brain dose, and urinary bladder wall doses are about 15% higher than the bladder dose for [¹⁸F]FDG.¹⁵ Overall, the radiation dosimetry profiles for [¹⁸F]4F-MHPG and [¹⁸F]3F-PHPG compare favorably to that of the widely used PET radiopharmaceutical [¹⁸F]FDG.

[¹⁸F]4F-MHPG and [¹⁸F]3F-PHPG are not the first fluorine-18 labeled radiotracers to be used for imaging cardiac sympathetic innervation in human subjects. Goldstein and coworkers at the National Institutes of Health have performed many pioneering studies of the impact of cardiovascular and neurological diseases on cardiac sympathetic nerve populations using 6-[¹⁸F]fluorodopamine^17–21. Recently, [¹⁸F]-LMI1195, a fluorine-18 labeled analog of [¹²³I]MIBG, was evaluated in normal human subjects, including biodistribution studies and human radiation dosimetry estimates.²² Comparing radiation absorbed dose estimates, like [¹⁸F]4F-MHPG and [¹⁸F]3F-PHPG, the critical organ for these tracers is the urinary bladder wall. One noteworthy difference is in the absorbed dose to the kidneys, which for [¹⁸F]4F-MHPG and [¹⁸F]3F-PHPG is about 11% – 12% of the value reported for 6-[¹⁸F]fluorodopamine and 25% - 29% of the dose estimated for [¹⁸F]-LMI1195.^21,22 In terms of myocardial kinetics, our institution has not worked with either of these agents, so we do not have sufficient kinetic data to make a detailed comparison between them and our new tracers. Based on the existing literature, it appears that 6-[¹⁸F]fluorodopamine and [¹⁸F]-LMI1195 each are rapidly transported into sympathetic nerve terminals and exhibit reversible tissue kinetics.²³ Thus, one advantage of [¹⁸F]4F-MHPG and [¹⁸F]3F-PHPG is their irreversible neuronal retention kinetics, which allows application of the Patlak graphical analysis technique to obtain quantitative metrics of regional nerve density.

Summary and Conclusion

Phase I studies with [¹⁸F]4F-MHPG and [¹⁸F]3F-PHPG have demonstrated that both tracers provide high quality cardiac PET images, with extremely low uptake in lungs and acceptable uptake levels in the liver. Patlak analysis of the myocardial kinetics of [¹⁸F]4F-MHPG and [¹⁸F]3F-PHPG provides robust quantitative nerve density metrics, generating polar maps of regional sympathetic nerve density at high spatial resolution. Comparing the two tracers, one advantage of [¹⁸F]4F-MHPG is its more rapid clearance from the liver. This not only improves image interpretation, but reduces spillover of counts from the liver into areas of the left ventricle, which can confound kinetic analyses. On the other hand, the more prolonged accumulation of [¹⁸F]3F-PHPG into sympathetic neurons may be an advantage in accurately measuring low nerve densities in areas of severe denervation. Since each tracer possesses advantages over the other, they will be compared head-to-head in heart failure patients staged for ICD placement to determine which is better for clinical assessments of regional denervation in diseased hearts. In conclusion, the findings of these pilot studies in normal subjects suggest that [¹⁸F]4F-MHPG and [¹⁸F]3F-PHPG may represent a significant advance in our ability to precisely quantify the extent and severity of cardiac sympathetic denervation in heart diseases using PET. Future clinical studies in patients with a wide range of cardiac denervation levels, including heart failure patients, are required to confirm this conclusion. If they do, [¹⁸F]4F-MHPG or [¹⁸F]3F-PHPG could one day be distributed commercially to cardiac PET centers for routine clinical assessments of cardiac sympathetic denervation in patients with heart disease.

Clinical Perspective.

Cardiac autonomic dysfunction can promote malignant arrhythmias, leading to sudden cardiac arrest. Recent clinical trials with radiolabeled sympathetic nerve imaging agents have established that higher levels of regional cardiac sympathetic denervation in heart failure are associated with a greatly elevated risk of ventricular arrhythmias and sudden death. These results point to a potential role of cardiac neuroimaging in improved sudden death risk stratification in heart failure patients being staged for placement of implantable cardioverter defibrillators (ICDs). This study presents first-in-human studies of two new fluorine-18 labeled radiotracers developed for more accurate quantification of the extent of regional cardiac denervation using PET. The findings show that these new radiotracers provide high quality cardiac PET images, are safe and well-tolerated, and have human radiation dosimetry profiles comparable to existing PET radiopharmaceuticals. In addition, their myocardial kinetics can be analyzed using Patlak graphical analysis to provide quantitative metrics of regional nerve density at high spatial resolution in the heart. Achieving similar results in patients with heart failure could ultimately allow one of these two radiotracers to be commercialized and distributed to PET centers, with the goal of providing improved risk stratification for ICD therapy.