Cannabimimetic activities of cumyl carboxamide-type synthetic cannabinoids

1-Pentyl-N-(2-phenylpropan-2-yl)-1H-indazole-3-carboxamide (CUMYL-PINACA) is a carboxamide-type synthetic cannabinoid comprising a cumylamine moiety. Recently, the detection of CUMYL-PINACA and some analogs in illicit drug products has been reported by the European Monitoring Centre for Drugs and Drug Addiction. In this study, we synthesized seven cumyl carboxamide-type synthetic cannabinoids (CUMYL-PINACA, CUMYL-5F-PINACA, CUMYL-PICA, CUMYL-5F-PICA, CUMYL-THPINACA, CUMYL-BICA, and CUMYL-5F-P7AICA) and evaluated their activities as CB₁ and CB₂ receptor agonists. We also showed the analytical characterization of these compounds using gas chromatography–electron ionization-mass spectrometry. All of the evaluated compounds exhibited moderate to strong activities as agonists acting on both CB₁ and CB₂ receptors with EC₅₀ values in the range of 8.1 × 10^?10–7.8 × 10^?7 mol/L for CB₁ and from 2.5 × 10^?10 to 9.1 × 10^?6 mol/L for CB₂. The EC₅₀ data presented will be helpful to understand the effects of these compounds in the forensic cases. Furthermore, other new cumyl carboxamide-type synthetic cannabinoids, which will be potentially distributed in the future, will probably have the activities as agonists acting on both CB₁ and CB₂ receptors.     

Sensitive identification and quantitation of parent forms of six synthetic cannabinoids in urine samples of human cadavers by liquid chromatography–tandem mass spectrometry

Human urine samples are easier to obtain than human blood samples due to noninvasiveness. The urine levels of synthetic cannabinoids (SCs) in unchanged forms, however, are usually much lower than their blood and tissue levels and cannot be detected in most cases. Therefore, in the present work a sensitive analytical method was devised for the determination of urine levels of six SCs in unchanged forms such as N-(1-amino-3-methy-1-oxobutan-2-yl)-1-pentyl-1H-indazole-3-carboxamide (AB-PINACA), N-(1-amino-3-methy-1-oxobutan-2-yl)-1-(4-fluorobenzyl)-1H-indazole-3-carboxamide (AB-FUBINACA), N-[(1S)-1-(1-aminocarbonyl)-2-methyl-propyl]-1-(cyclohexylmethyl)-1H-indazole-3-carboxamide (AB-CHMINACA), N-(1-amino-3,3-dimethyl-1-oxobutan-2-yl)-1-(cyclohexylmethyl)-1H-indazole-3-carboxamide (MAB-CHMINACA), methyl-2-[1-(5-fluoropentyl)-1H-indazole-3-carboxamido]-3-methylbutanoate (5F-AMB) and methyl-2-[1-(5-fluoropentyl)-1H-indazole-3-carboxamido]-3,3-dimethylbutanoate (5F-ADB). These SCs were extracted from urine via liquid–liquid extraction. The identification and quantitation were performed by a relatively new type of an instrument for liquid chromatography–tandem mass spectrometry. The limits of detection were as low as 3–8 pg/mL, and the quantitation range was 10–1000 pg/mL using 400 μL of urine. The urine levels of AB-PINACA and AB-FUBINACA of victim 1 were 23 and 10 pg/mL, those of AB-CHMINACA and 5F-AMB of victim 2 were 239 and 19 pg/mL, and those of MAB-CHMINACA and 5F-ADB of victim 3 were 229 and 19 pg/mL, respectively. To our knowledge, this is the first report dealing with successful analysis of low levels of parent synthetic cannabinoids in authentic human urine specimens.     

MDMB-CHMICA induces thrashing behavior,bradycardia, and slow pressor response in a CB<Subscript>1</Subscript>- and CB<Subscript>2</Subscript>-receptor-dependent manner in conscious rats

Purpose

MDMB-CHMICA, a new synthetic cannabinoid (SC), has become prevalent since 2014 as an ingredient of recreational drugs. Reports on intoxication due to the drug have been increasing, which show diverse cardiovascular, psychiatric, and neuronal symptoms. Reports on sudden death and accidental death related to psychiatric disorders in MDMB-CHMICA intoxication have also increased, but the underlying mechanisms are largely unknown.

Methods

As there has been no experimental study on the drug, we investigated the effects of peripheral injection of MDMB-CHMICA in conscious rats.

Results

MDMB-CHMICA induced rapid bradycardia and a slow pressor response. Cardiovascular responses to other SCs have been shown to be inhibited only by cannabinoid receptor-1 (CB₁)-antagonists. However, the MDMB-CHMICA-induced bradycardia was inhibited not only by a CB₁-antagonist, AM281, but also by a CB₂-antagonist, AM630. Unlike other SCs, MDMB-CHMICA induced a gradual increase in mean blood pressure, which was marginally enhanced by the CB₁- and CB₂- antagonists. For the first time, we demonstrated that MDMB-CHMICA induces a thrashing hypermobile behavior in a CB₁- and CB₂-receptor-dependent manner, following catalepsy-like hypomobile behavior.

Conclusions

This unexpected response to MDMB-CHMICA may help explain the mechanisms underlying the sudden deaths and accidents associated with its use.

     

The 2-alkyl-2<Emphasis Type="Italic">H</Emphasis>-indazole regioisomers of synthetic cannabinoids AB-CHMINACA,AB-FUBINACA,AB-PINACA,and 5F-AB-PINACA are possible manufacturing impurities with cannabimimetic activities

Indazole-derived synthetic cannabinoids (SCs) featuring an alkyl substituent at the 1-position and l-valinamide at the 3-carboxamide position (e.g., AB-CHMINACA) have been identified by forensic chemists around the world, and are associated with serious adverse health effects. Regioisomerism is possible for indazole SCs, with the 2-alkyl-2H-indazole regioisomer of AB-CHMINACA recently identified in SC products in Japan. It is unknown whether this regiosiomer represents a manufacturing impurity arising as a synthetic byproduct, or was intentionally synthesized as a cannabimimetic agent. This study reports the synthesis, analytical characterization, and pharmacological evaluation of commonly encountered indazole SCs AB-CHMINACA, AB-FUBINACA, AB-PINACA, 5F-AB-PINACA and their corresponding 2-alkyl-2H-indazole regioisomers. Both regioisomers of each SC were prepared from a common precursor, and the physical properties, ¹H and ¹³C nuclear magnetic resonance spectroscopy, gas chromatography–mass spectrometry, and ultraviolet–visible spectroscopy of all SC compounds are described. Additionally, AB-CHMINACA, AB-FUBINACA, AB-PINACA, and 5F-AB-PINACA were found to act as high potency agonists at CB₁ (EC₅₀ = 2.1–11.6 nM) and CB₂ (EC₅₀ = 5.6–21.1 nM) receptors in fluorometric assays, while the corresponding 2-alkyl-2H-indazole regioisomers demonstrated low potency (micromolar) agonist activities at both receptors. Taken together, these data suggest that 2-alkyl-2H-indazole regioisomers of AB-CHMINACA, AB-FUBINACA, AB-PINACA, and 5F-AB-PINACA are likely to be encountered by forensic chemists and toxicologists as the result of improper purification during the clandestine synthesis of 1-alkyl-1H-indazole regioisomers, and can be distinguished by differences in gas chromatography–mass spectrometry fragmentation pattern.     

Quantitative <Emphasis Type="Italic">T</Emphasis><Subscript>1</Subscript> and <Emphasis Type="Italic">T</Emphasis><Subscript>2</Subscript>* carotid atherosclerotic plaque imaging using a three-dimensional multi-echo phase-sensitive inversion recovery sequence: a feasibility study

Magnetic resonance imaging (MRI) is widely used to detect carotid atherosclerotic plaques. Although it is important to evaluate vulnerable carotid plaques containing lipids and intra-plaque hemorrhages (IPHs) using T₁-weighted images, the image contrast changes depending on the imaging settings. Moreover, to distinguish between a thrombus and a hemorrhage, it is useful to evaluate the iron content of the plaque using both T₁-weighted and T₂*-weighted images. Therefore, a quantitative evaluation of carotid atherosclerotic plaques using T₁ and T₂* values may be necessary for the accurate evaluation of plaque components. The purpose of this study was to determine whether the multi-echo phase-sensitive inversion recovery (mPSIR) sequence can improve T₁ contrast while simultaneously providing accurate T₁ and T₂* values of an IPH. T₁ and T₂* values measured using mPSIR were compared to values from conventional methods in phantom and in vivo studies. In the phantom study, the T₁ and T₂* values estimated using mPSIR were linearly correlated with those of conventional methods. In the in vivo study, mPSIR demonstrated higher T₁ contrast between the IPH phantom and sternocleidomastoid muscle than the conventional method. Moreover, the T₁ and T₂* values of the blood vessel wall and sternocleidomastoid muscle estimated using mPSIR were correlated with values measured by conventional methods and with values reported previously. The mPSIR sequence improved T₁ contrast while simultaneously providing accurate T₁ and T₂* values of the neck region. Although further study is required to evaluate the clinical utility, mPSIR may improve carotid atherosclerotic plaque detection and provide detailed information about plaque components.     

New cannabimimetic indazole derivatives, N-(1-amino-3-methyl-1-oxobutan-2-yl)-1-pentyl-1H-indazole-3-carboxamide (AB-PINACA) and N-(1-amino-3-methyl-1-oxobutan-2-yl)-1-(4-fluorobenzyl)-1H-indazole-3-carboxamide (AB-FUBINACA) identified as designer drugs in illegal products

Two new cannabimimetic indazole derivatives, N-(1-amino-3-methyl-1-oxobutan-2-yl)-1-pentyl-1H-indazole-3-carboxamide (AB-PINACA, 1) and N-(1-amino-3-methyl-1-oxobutan-2-yl)-1-(4-fluorobenzyl)-1H-indazole-3-carboxamide (AB-FUBINACA, 2), have been identified as designer drugs in illegal products. These identifications were based on liquid chromatography–mass spectrometry, gas chromatography–mass spectrometry, high-resolution mass spectrometry, and nuclear magnetic resonance spectroscopy. Because there have been neither chemical nor pharmacological data about compound 1 until now, this is the first report of this compound. Compound 2 was reported as a potent cannabinoid CB₁ receptor modulator when synthesized by Pfizer in 2009; but this is the first report of its detection in illegal products.     

The prognostic and predictive value of sstr<Subscript>2</Subscript>-immunohistochemistry and sstr<Subscript>2</Subscript>-targeted imaging in neuroendocrine tumors

Purpose

Our aim was to assess the prognostic and predictive value of somatostatin receptor 2 (sstr₂) in neuroendocrine tumors (NETs).

Methods

We established a tissue microarray and imaging database from NET patients that received sstr₂-targeted radiopeptide therapy with yttrium-90-DOTATOC, lutetium-177-DOTATOC or alternative treatment. We used univariate and multivariate analyses to identify prognostic and predictive markers for overall survival, including sstr₂-imaging and sstr₂-immunohistochemistry.

Results

We included a total of 279 patients. In these patients, sstr₂-immunohistochemistry was an independent prognostic marker for overall survival (HR: 0.82, 95 % CI: 0.67 – 0.99, n?=?279, p?=?0.037). In DOTATOC patients, sstr₂-expression on immunohistochemistry correlated with tumor uptake on sstr₂-imaging (n?=?170, p?<?0.001); however, sstr₂-imaging showed a higher prognostic accuracy (positive predictive value: +27 %, 95 % CI: 3 – 56 %, p?=?0.025). Sstr₂-expression did not predict a benefit of DOTATOC over alternative treatment (p?=?0.93).

Conclusions

Our results suggest sstr₂ as an independent prognostic marker in NETs. Sstr₂-immunohistochemistry correlates with sstr₂-imaging; however, sstr₂-imaging is more accurate for determining the individual prognosis.

     

<Emphasis Type="Italic">V</Emphasis>O<Subscript>2Max</Subscript> and <Emphasis Type="Italic">V</Emphasis>O<Subscript>2AT</Subscript>: athletic performance and field role of elite soccer players

Purpose

Maximal oxygen consumption (VO_2max) and oxygen consumption at anaerobic threshold (VO_2AT) are commonly measured parameters to test elite soccer players; however, studies relating metabolic parameters of professional soccer players with performance and best fitting to the field role are scarce. Our aim was to study the relations of VO_2max and VO_2AT with the field role of elite soccer players to generate a robust dataset with a solid statistical analysis.

Method

Over a 12-year period we performed 953 field evaluation tests of VO_{2 max} and VO_2AT on 450 elite soccer players of 13 professional teams by incremental, continuous and exhausting test modified from Conconi’s test. Statistical analysis was performed by one-way ANOVA followed—when appropriate—by Tukey post hoc test. Effect size was evaluated by the Cohen D test and η partial squared test. Statistical significance was set for p < 0.05.

Results

VO_2max mean values increased at the beginning of season by 1.073 ± 0.06 respect to pre-season then decreased again up to pre-season levels and remained stable, while VO_2AT did not change during time. VO_2max differences were also related to players’ field role. VO_2max mean value for Goalkeeper was 50.85 ± 4.2, for central Defender was 57.58 ± 4.3, for Winger-sides back was 60.53 ± 5.02, for Midfielder was 59.53 ± 5.08, for Forward was 56.52 ± 4.14. On the contrary, as expected, VO_2AT percentage variations among the roles were not significant.

Conclusions

VO_2max is the choice parameter to consider for the metabolic compliance of athletes to the field role requirements, consequently influencing training programs, recovery and injury prevention strategies.

     

Two new-type cannabimimetic quinolinyl carboxylates, QUPIC and QUCHIC, two new cannabimimetic carboxamide derivatives, ADB-FUBINACA and ADBICA, and five synthetic cannabinoids detected with a thiophene derivative α-PVT and an opioid receptor agonist AH-7921 identified in illegal products

We identified two new-type cannabimimetic quinolinyl carboxylates, quinolin-8-yl 1-pentyl-(1H-indole)-3-carboxylate (QUPIC, 1) and quinolin-8-yl 1-(cyclohexylmethyl)-1H-indole-3-carboxylate (QUCHIC, 2); and two new cannabimimetic carboxamide derivatives, N-(1-amino-3,3-dimethyl-1-oxobutan-2-yl)-1-(4-fluorobenzyl)-1H-indazole-3-carboxamide (ADB-FUBINACA, 3) and N-(1-amino-3,3-dimethyl-1-oxobutan-2-yl)-1-pentyl-1H-indole-3-carboxamide (ADBICA, 4), as designer drugs in illegal products. Compound 3 was reported to have a potent affinity for cannabinoid CB₁ receptor by Pfizer in 2009, but this is the first report of its detection in illegal products. No chemical or pharmacological data for compounds 1, 2, and 4 have appeared until now, making this the first report on these compounds. We also detected synthetic cannabinoids, APICA N-(5-fluoropentyl) analog (5), APINACA N-(5-fluoropentyl) analog (6), UR-144 N-(5-chloropentyl) analog (7), JWH-122 N-(5-chloropentyl) analog (8), and AM-2201 4-methoxynaphthyl analog (4-MeO-AM-2201, 9) herein as newly distributed designer drugs in Japan. It is of interest that compounds 1 and 2 were detected with their synthetic component, 8-quinolinol (10). A stimulant thiophene analog, α-pyrrolidinovalerothiophenone (α-PVT, 11), and an opioid receptor agonist, 3,4-dichloro-N-([1-(dimethylamino)cyclohexyl]methyl)benzamide (AH-7921, 12), were also detected as new types of designer drugs coexisting with several synthetic cannabinoids and cathinone derivatives in illegal products.     

Transjugular intrahepatic portosystemic shunt creation: three-dimensional roadmap versus CO<Subscript>2</Subscript> wedged hepatic venography

Objectives

The blind portal vein puncture remains the most challenging step during transjugular intrahepatic portosystemic shunt (TIPS) creation. We performed a prospective randomised clinical trial to compare three-dimensional (3D) roadmap with CO₂ wedged hepatic vein portography for portal vein puncture guidance.

Methods

Between March 2017 and May 2017, 30 patients were enrolled and randomly allocated to the study group (3D roadmap) or the control group (CO₂ wedged hepatic vein portography).

Results

Technical success of TIPS procedures was achieved in all 30 patients. The mean number of needle passes was significantly lower in the study group (2.0 ± 1.0) compared to the control group (3.7 ± 2.5; p = 0.021). A total of six (40%) patients in the study group and three (20%) in the control group required only one puncture for the establishment of TIPS. There were no significant differences in total fluoroscopy time (p = 0.905), total procedure time (p = 0.199) and dose-area product (p = 0.870) between the two groups.

Conclusions

3D roadmap is a safe and technically feasible means for portal vein puncture guidance during TIPS creation, equivalent in efficacy to CO₂ wedged hepatic vein portography. This technique could reduce the number of needle passes, thereby simplifying the TIPS procedure.

Key Points

? 3D roadmap can be used to guide portal vein puncture. ? Compared with CO ₂ venography, 3D roadmap reduced the number of needle passes. ? 3D roadmap has a potential to simplify the TIPS procedure.

     

High-resolution T<Subscript>2</Subscript>-weighted cervical cancer imaging: a feasibility study on ultra-high-field 7.0-T MRI with an endorectal monopole antenna

Objectives

We studied the feasibility of high-resolution T₂-weighted cervical cancer imaging on an ultra-high-field 7.0-T magnetic resonance imaging (MRI) system using an endorectal antenna of 4.7-mm thickness.

Methods

A feasibility study on 20 stage IB1–IIB cervical cancer patients was conducted. All underwent pre-treatment 1.5-T MRI. At 7.0-T MRI, an external transmit/receive array with seven dipole antennae and a single endorectal monopole receive antenna were used. Discomfort levels were assessed. Following individualised phase-based B₁ ⁺ shimming, T₂-weighted turbo spin echo sequences were completed.

Results

Patients had stage IB1 (n = 9), IB2 (n = 4), IIA1 (n = 1) or IIB (n = 6) cervical cancer. Discomfort (ten-point scale) was minimal at placement and removal of the endorectal antenna with a median score of 1 (range, 0–5) and 0 (range, 0–2) respectively. Its use did not result in adverse events or pre-term session discontinuation. To demonstrate feasibility, T₂-weighted acquisitions from 7.0-T MRI are presented in comparison to 1.5-T MRI. Artefacts on 7.0-T MRI were due to motion, locally destructive B₁ interference, excessive B₁ under the external antennae and SENSE reconstruction.

Conclusions

High-resolution T₂-weighted 7.0-T MRI of stage IB1–IIB cervical cancer is feasible. The addition of an endorectal antenna is well tolerated by patients.

Key Points

? High resolution T ₂ -weighted 7.0-T MRI of the inner female pelvis is challenging ? We demonstrate a feasible approach for T ₂ -weighted 7.0-T MRI of cervical cancer ? An endorectal monopole receive antenna is well tolerated by participants ? The endorectal antenna did not lead to adverse events or session discontinuation

     

Correlation of <Superscript>18</Superscript>F-FDG and <Superscript>11</Superscript>C-methionine uptake on PET/CT with Ki-67 immunohistochemistry in newly diagnosed intracranial meningiomas

Objective

We evaluated the uptake of 2-deoxy-2-¹⁸F-fluoro-d-glucose (FDG) and l-[methyl-¹¹C]-methionine (MET) in patients with newly diagnosed intracranial meningiomas and correlated the results with tumor proliferation.

Methods

Data from 22 patients with newly diagnosed intracranial meningioma (12 grade I and 10 grade II) who underwent both FDG and MET brain PET/CT studies were retrospectively analyzed. The PET images were evaluated by a qualitative method and semiquantitative analysis using standardized uptake value (SUV) (SUV_max and SUV_peak) and tumor-to-reference tissue ratio (T_max/N ratio and T_peak/N ratio). Proliferative activity as indicated by the Ki-67 index was estimated in tissue specimens.

Results

MET PET/CT showed a higher detection rate of meningioma than did FDG PET/CT (100 vs. 46%, respectively). The T_max/N ratio and T_peak/N ratio on MET PET/CT were significantly higher than those on FDG PET/CT (p?<?0.001 and p?<?0.001, respectively). There was a significant difference between grades I and II with respect to FDG SUVmax (p?=?0.003), FDG SUV_peak (p?=?0.003), FDG T_max/N ratio (p?=?0.02), FDG T_peak/N ratio (p?=?0.006), MET SUV_max (p?=?0.002), MET SUV_peak (p?=?0.002), MET T_max/N ratio (p?=?0.002), and MET T_peak/N ratio (p?=?0.002). There was a significant correlation between Ki-67 index and FDG PET/CT for SUV_max (p?=?0.02), SUV_peak (p?=?0.005), and T_peak/N ratio (p?=?0.05) and between Ki-67 index and MET PET/CT for SUV_max (p?=?0.004), SUV_peak (p?=?0.007), T_max/N ratio (p?=?0.002), and T_peak/N ratio (p?=?0.004).

Conclusion

MET PET/CT showed a high sensitivity compared with FDG PET/CT for detection of newly diagnosed WHO grades I and II intracranial meningiomas. Both FDG and MET uptake were found to be useful for evaluating tumor proliferation in meningiomas.

     

Pixel-by-pixel precise delay correction for measurement of cerebral hemodynamic parameters in H<Subscript>2</Subscript><Superscript>15</Superscript>O PET study

Objective

A new method of delay time estimation was proposed to measure precise cerebral blood flow (CBF) and arterial-to-capillary blood volume (V ₀) using ¹⁵O-water PET.

Methods

Nineteen patients with unilateral arterial stenoocclusive lesions were studied to evaluate hemodynamic status before treatment. The delay time of each pixel was calculated using least squares fitting with an arterial blood input curve adjusted to the internal carotid artery counts at the skull base. Pixel-by-pixel delay estimation provided a delay map image that could be used for precise calculation of CBF and V ₀ using a one-tissue compartment model, and the values from this method were compared with those from the slice-by-slice correction method.

Results

The affected side showed a longer delay time than the contralateral cerebral hemisphere. Although the mean cortical CBF values were not different between the two methods, the slice-by-slice delay correction overestimated CBF in the hypo perfused area. The scatter plot of V ₀ pixel values showed significant difference between the two correction methods where the slice-by-slice delay correction significantly overestimated V ₀ in the whole brain (P?<?0.05).

Conclusion

Pixel-by-pixel delay correction provides delay images as well as better estimation of CBF and V ₀, thus offering useful and beneficial information for the treatment of cerebrovascular disease.

     

Comparative evaluation of the algorithms for parametric mapping of the novel myocardial PET imaging agent <Superscript>18</Superscript>F-FPTP

Objective

(¹⁸F-fluoropentyl)triphenylphosphonium salt (¹⁸F-FPTP) is a new promising myocardial PET imaging tracer. It shows high accumulation in cardiomyocytes and rapid clearance from liver. We performed compartmental analysis of ¹⁸F-FPTP PET images in rat and evaluated two linear analyses: linear least-squares (LLS) and a basis function method (BFM) for generating parametric images. The minimum dynamic scan duration for kinetic analysis was also investigated and computer simulation undertaken.

Methods

¹⁸F-FPTP dynamic PET (18 min) and CT images were acquired from rats with myocardial infarction (MI) (n = 12). Regions of interest (ROIs) were on the left ventricle, normal myocardium, and MI region. Two-compartment (K ₁ and k ₂; 2C2P) and three-compartment (K ₁–k ₃; 3C3P) models with irreversible uptake were compared for goodness-of-fit. Partial volume and spillover correction terms (V _a and α = 1 ? V _a ) were also incorporated. LLS and BFM were applied to ROI- and voxel-based kinetic parameter estimations. Results were compared with the standard ROI-based nonlinear least-squares (NLS) results of the corresponding compartment model. A simulation explored statistical properties of the estimation methods.

Results

The 2C2P model was most suitable for describing ¹⁸F-FPTP kinetics. Average K ₁, k ₂, and V _a values were, respectively, 6.8 (ml/min/g), 1.1 (min^?1), and 0.44 in normal myocardium and 1.4 (ml/min/g), 1.1 (min^?1), and 0.32, in MI tissue. Ten minutes of data was sufficient for the estimation. LLS and BFM estimations correlated well with NLS values for the ROI level (K ₁: y = 1.06x + 0.13, r ²  = 0.96 and y = 1.13x + 0.08, r ²  = 0.97) and voxel level (K ₁: y = 1.22x ? 0.30, r ²  = 0.90 and y = 1.26x + 0.00, r ²  = 0.92). Regional distribution of kinetic parametric images (αK ₁, K ₁, k ₂, V _a) was physiologically relevant. LLS and BFM showed more robust characteristics than NLS in the simulation.

Conclusions

Fast kinetics and highly specific uptake of ¹⁸F-FPTP by myocardium enabled quantitative analysis with the 2C2P model using only the initial 10 min of data. LLS and BFM were feasible for estimating voxel-wise parameters. These two methods will be useful for quantitative evaluation of ¹⁸F-FPTP distribution in myocardium and in further studies with different conditions, disease models, and species.

     

In vitro metabolism of synthetic cannabinoid AM1220 by human liver microsomes and <Emphasis Type="Italic">Cunninghamella elegans</Emphasis> using liquid chromatography coupled with high resolution mass spectrometry

Purpose

Identifying intake of synthetic cannabinoids generally requires the metabolism data of the drugs so that appropriate metabolite markers can be targeted in urine testing. However, the continuous appearance of new cannabinoids during the last decade has made it difficult to keep up with all the compounds including {1-[(1-methylpiperidin-2-yl)methyl]-1H-indol-3-yl}(naphthalen-1-yl)methanone (AM1220). In this study, metabolism of AM1220 was investigated with human liver microsomes and the fungus Cunninghamella elegans.

Methods

Metabolic stability of AM1220 was analysed by liquid chromatography–tandem mass spectrometry in multiple reaction monitoring mode after 1 µM incubation in human liver microsomes for 30 min. Tentative structure elucidation of metabolites was performed on both human liver microsome and fungal incubation samples using liquid chromatography–high-resolution mass spectrometry.

Results

Half-life of AM1220 was estimated to be 3.7 min, indicating a high clearance drug. Nine metabolites were detected after incubating human liver microsomes while seven were found after incubating Cunninghamella elegans, leading to 11 metabolites in total (five metabolites were common to both systems). Demethylation, dihydrodiol formation, combination of the two, hydroxylation and dihydroxylation were the observed biotransformations.

Conclusions

Three most abundant metabolites in both human liver microsomes and Cunninghamella elegans were desmethyl, dihydrodiol and hydroxy metabolites, despite different isomers of dihydrodiol and hydroxy metabolites in each model. These abundant metabolites can potentially be useful markers in urinalysis for AM1220 intake.

     

Fully parametric imaging with reversible tracer <Superscript>18</Superscript>F-FLT within a reasonable time

PET enables quantitative imaging of the rate constants K ₁, k ₂, k ₃, and k ₄, with a reversible two tissue compartment model (2TCM). A new method is proposed for computing all of these rates within a reasonable time, less than 1 min. A set of differential equations for the reversible 2TCM was converted into a single formula consisting of differential and convolution terms. The validity was tested on clinical data with ¹⁸F-FLT PET for patients with glioma (n = 39). Parametric images were generated with the formula that was developed. Parametric values were extracted from regions of interest (ROIs) for glioma from the images generated, and they were compared with those obtained with the non-linear fitting method. We performed simulation studies for testing accuracy by generating simulated images, assuming clinically expected ranges of the parametric values. The computation time was about 20 s, and the quality of the images generated was acceptable. The values obtained for K ₁ for grade IV tumor were 0.24 ± 0.23 and 0.26 ± 0.25 ml^?1 min^?1 g^?1 for the image-based and ROI-based methods, respectively. The values were 0.21 ± 0.12 and 0.21 ± 0.12 min^?1 for k ₂, 0.13 ± 0.07 and 0.13 ± 0.07 min^?1 for k ₃, and 0.052 ± 0.020 and 0.054 ± 0.021 min^?1 for k ₄. The differences between the methods were not significant. Regression analysis showed correlations of r = 0.94, 0.86, 0.71, and 0.52 for these parameters. Simulation demonstrated that the accuracy was within acceptable ranges, namely, the correlations were r = 0.99, r = 0.97, r = 0.99, and r = 0.91 for K ₁, k ₂, k ₃, and k ₄, respectively, between estimated and assumed values. This results suggest that parametric images can be obtained fully within reasonable time, accuracy, and quality.     

<Superscript>11</Superscript>C-L-methyl methionine dynamic PET/CT of skeletal muscle: response to protein supplementation compared to L-[ring <Superscript>13</Superscript>C<Subscript>6</Subscript>] phenylalanine infusion with serial muscle biopsy

Objective

The objective of this study was to determine if clinical dynamic PET/CT imaging with ¹¹C-L-methyl-methionine (¹¹C-MET) in healthy older women can provide an estimate of tissue-level post-absorptive and post-prandial skeletal muscle protein synthesis that is consistent with the more traditional method of calculating fractional synthesis rate (FSR) of muscle protein synthesis from skeletal muscle biopsies obtained during an infusion of L-[ring ¹³C₆] phenylalanine (¹³C₆-Phe).

Methods

Healthy older women (73?±?5 years) completed both dynamic PET/CT imaging with ¹¹C-MET and a stable isotope infusion of ¹³C₆-Phe with biopsies to measure the skeletal muscle protein synthetic response to 25 g of a whey protein supplement. Graphical estimation of the Patlak coefficient K_i from analysis of the dynamic PET/CT images was employed as a measure of incorporation of 11 C-MET in the mid-thigh muscle bundle.

Results

Post-prandial values [mean?±?standard error of the mean (SEM)] were higher than post-absorptive values for both K_i (0.0095?±?0.001 vs. 0.00785?±?0.001 min^?1, p?<?0.05) and FSR (0.083?±?0.008 vs. 0.049?±?0.006%/h, p?<?0.001) in response to the whey protein supplement. The percent increase in K_i and FSR in response to the whey protein supplement was significantly correlated (r?=?0.79, p?=?0.015).

Conclusions

Dynamic PET/CT imaging with ¹¹C-MET provides an estimate of the post-prandial anabolic response that is consistent with a traditional, invasive stable isotope, and muscle biopsy approach. These results support the potential future use of ¹¹C-MET imaging as a non-invasive method for assessing conditions affecting skeletal muscle protein synthesis.

     

Monitoring early response to chemoradiotherapy with <Superscript>18</Superscript>F-FMISO dynamic PET in head and neck cancer

Purpose

There is growing recognition that biologic features of the tumor microenvironment affect the response to cancer therapies and the outcome of cancer patients. In head and neck cancer (HNC) one such feature is hypoxia. We investigated the utility of ¹⁸F-fluoromisonidazole (FMISO) dynamic positron emission tomography (dPET) for monitoring the early microenvironmental response to chemoradiotherapy in HNC.

Experimental design

Seventy-two HNC patients underwent FMISO dPET scans in a customized immobilization mask (0–30 min dynamic acquisition, followed by 10 min static acquisitions starting at ～95 min and ～160 min post-injection) at baseline and early into treatment where patients have already received one cycle of chemotherapy and anywhere from five to ten fractions of 2 Gy per fraction radiation therapy. Voxelwise pharmacokinetic modeling was conducted using an irreversible one-plasma two-tissue compartment model to calculate surrogate biomarkers of tumor hypoxia (k ₃ and Tumor-to-Blood Ratio (TBR)), perfusion (K ₁ ) and FMISO distribution volume (DV). Additionally, Tumor-to-Muscle Ratios (TMR) were derived by visual inspection by an experienced nuclear medicine physician, with TMR > 1.2 defining hypoxia.

Results

One hundred and thirty-five lesions in total were analyzed. TBR, k ₃ and DV decreased on early response scans, while no significant change was observed for K ₁ . The k ₃ -TBR correlation decreased substantially from baseline scans (Pearson’s r = 0.72 and 0.76 for mean intratumor and pooled voxelwise values, respectively) to early response scans (Pearson’s r = 0.39 and 0.40, respectively). Both concordant and discordant examples of changes in intratumor k ₃ and TBR were identified; the latter partially mediated by the change in DV. In 13 normoxic patients according to visual analysis (all having lesions with TMR = 1.2), subvolumes were identified where k ₃ indicated the presence of hypoxia.

Conclusion

Pharmacokinetic modeling of FMISO dynamic PET reveals a more detailed characterization of the tumor microenvironment and assessment of response to chemoradiotherapy in HNC patients than a single static image does. In a clinical trial where absence of hypoxia in primary tumor and lymph nodes would lead to de-escalation of therapy, the observed disagreement between visual analysis and pharmacokinetic modeling results would have affected patient management in <20% cases. While simple static PET imaging is easily implemented for clinical trials, the clinical applicability of pharmacokinetic modeling remains to be investigated.

     

In vitro and in vivo pharmacokinetics and metabolism of synthetic cannabinoids CUMYL-PICA and 5F-CUMYL-PICA

CUMYL-PICA [1-pentyl-N-(2-phenylpropan-2-yl)-1H-indole-3-carboxamide] and 5F-CUMYL-PICA [1-(5-fluoropentyl)-N-(2-phenylpropan-2-yl)-1H-indole-3-carboxamide] are recently identified recreationally used/abused synthetic cannabinoids, but have uncharacterized pharmacokinetic profiles and metabolic processes. This study characterized clearance and metabolism of these compounds by human and rat liver microsomes and hepatocytes, and then compared these parameters with in vivo rat plasma and urine sampling. It also evaluated hypothermia, a characteristic cannabimimetic effect. Incubation of CUMYL-PICA and 5F-CUMYL-PICA with rat and human liver microsomes suggested rapid metabolic clearance, but in vivo metabolism was prolonged, such that parent compounds remained detectable in rat plasma 24 h post-dosing. At 3 mg/kg (intraperitoneally), both compounds produced moderate hypothermic effects. Twenty-eight metabolites were tentatively identified for CUMYL-PICA and, coincidentally, 28 metabolites for 5F-CUMYL-PICA, primarily consisting of phase I oxidative transformations and phase II glucuronidation. The primary metabolic pathways for both compounds resulted in the formation of identical metabolites following terminal hydroxylation or dealkylation of the N-pentyl chain for CUMYL-PICA or of the 5-fluoropentyl chain for 5F-CUMYL-PICA. These data provide evidence that in vivo elimination of CUMYL-PICA, 5F-CUMYL-PICA and other synthetic cannabinoids is delayed compared to in vitro modeling, possibly due to sequestration into adipose tissue. Additionally, the present data underscore the need for careful selection of metabolites as analytical targets to distinguish between closely related synthetic cannabinoids in forensic settings.     

Identification of the synthetic cannabinoid <Emphasis Type="Italic">N</Emphasis>-(2-phenyl-propan-2-yl)-1-(4-cyanobutyl)-1<Emphasis Type="Italic">H</Emphasis>-indazole-3-carboxamide (CUMYL-4CN-BINACA) in a herbal mixture product

We were the first to detect N-(2-phenylpropan-2-yl)-1-(4-cyanobutyl)-1H-indazole-3-carboxamide (common name CUMYL-4CN-BINACA) as a new synthetic cannabinoid, on the illegal market in Bursa, Turkey. To elucidate the chemical structure, the dried herbal mixture was extracted with methanol. The extract was purified by column chromatography. Pure compound was analyzed by gas chromatography–mass spectrometry (GC–MS), attenuated total reflection Fourier-transform infrared spectroscopy (FT-IR), and one- and two-dimensional nuclear magnetic resonance (NMR) spectroscopy. The GC–MS, FT-IR and ¹H and ¹³C NMR spectra of the compound coincided well with the reference data. All protons and carbons were assigned by their couplings and correlations observed in ¹H-¹H correlation spectroscopy, ¹H-¹³C heteronuclear multiple bond correlation, and ¹H-¹³C heteronuclear single quantum coherence spectra. On the basis of the spectral data, the compound was identified as CUMYL-4CN-BINACA. Herewith, we report analytical characteristics of CUMYL-4CN-BINACA enabling its (and possible analogues thereof) determination in criminal seizures.