The Enantioselective Synthesis of Eburnamonine, Eucophylline, and 16'-epi-Leucophyllidine.

A synthetic approach to the heterodimeric bisindole alkaloid leucophyllidine is disclosed herein. An enantioenriched lactam building block, synthesized through palladium-catalyzed asymmetric allylic alkylation, served as the precursor to both hemispheres. The eburnamonine-derived fragment was synthesized through a Bischler-Napieralski/hydrogenation approach, while the eucophylline-derived fragment was synthesized by Friedländer quinoline synthesis and two sequential C-H functionalization steps. A convergent Stille coupling and phenol-directed hydrogenation united the two monomeric fragments to afford 16'-epi-leucophyllidine in 21 steps from commercial material.

Novel delta opioid receptor agonists with oxazatricyclodecane structure showing potent agonistic activities.

We recently reported oxazatricyclodecane derivatives 1 as δ opioid receptor (DOR) agonists having a novel chemotype, but their DOR agonistic activities were relatively low. Based on the working hypothesis that the dioxamethylene moiety in 1 may be an accessory site and that it may interfere with the sufficient conformational change of the receptor required for exerting the full agonistic responses, we designed and synthesized new oxazatricyclodecane derivatives 2-4 lacking the dioxamethylene moiety. As we expected, the designed compounds 2-4 showed pronouncedly improved agonistic activities for the DOR. Compound 2a with the 17-cyclopropylmethyl substituent was a potent agonist with the highest selectivity for the DOR and was expected to be a lead compound for novel and selective DOR agonists.

Design and synthesis of novel δ opioid receptor agonists with an azatricyclodecane skeleton for improving blood-brain barrier penetration.

We designed and synthesized novel δ opioid receptor (DOR) agonists 3a-i with an azatricyclodecane skeleton, which was a novel structural class of DOR agonists. Among them, 3b exhibited high values of binding affinity and potent agonistic activity for the DOR that were approximately equivalent to those of 2 which bore an oxazatricyclodecane skeleton. In vitro assays using the blood-brain barrier (BBB) permeability test kit supported the idea that 3b achieved an excellent BBB permeability by converting an oxygen atom of 2 to a carbon atom (methylene group) in the core skeleton. As a result, 3b showed potent antinociceptive effects.

Prognostic value of pretreatment 18F-FDG PET/CT parameters including visual evaluation in patients with head and neck squamous cell carcinoma.

OBJECTIVE: The purpose of this study was to determine whether pretreatment quantitative and visual parameters seen on PET/CT using (18)F-FDG add prognostic information for clinical staging in patients with head and neck cancer. MATERIALS AND METHODS: We enrolled 108 patients with histologically proven oral, oropharyngeal, hypopharyngeal, and laryngeal squamous cell carcinomas who underwent FDG PET/CT before treatment and, later, definitive therapy in our study. PET/CT parameters-maximum standardized uptake value (SUVmax), metabolic tumor volume (MTV), total lesion glycolysis (TLG), and uptake pattern (sphere-shaped or ring-shaped)-were recorded. The prognostic value of these parameters was evaluated using univariate and multivariate Cox regression analyses. RESULTS: In the univariate analysis, all of the FDG PET/CT parameters--SUVmax (> 10 g/mL) of the primary tumor, MTV (> 20 cm(3)), TLG (> 70 g), and uptake pattern (ring-shaped)--were significantly associated with negative effects on disease-specific survival (DSS) and disease-free survival (DFS). In the multivariate analysis, the MTV and uptake pattern remained associated with DSS after corrections for the Union for International Cancer Control (UICC) stage and definitive therapy (p = 0.023 and < 0.001, respectively). Another multivariate model that included MTV as a continuous variable, uptake pattern, and UICC stage showed that the uptake pattern remained significantly associated with DSS, whereas the association between DSS and MTV was not significant (p < 0.001 and = 0.332, respectively). CONCLUSION: Our data indicate that the pretreatment PET/CT parameters had prognostic value. In particular, a qualitative factor, uptake pattern, provided better prognostic information to the clinical staging of head and neck squamous cell carcinomas than the other PET/CT parameters.

A multicenter brain perfusion SPECT study evaluating idiopathic normal-pressure hydrocephalus on neurological improvement.

PURPOSE: This study was designed to investigate the specific cerebral blood flow (CBF) pattern in patients with idiopathic normal-pressure hydrocephalus (iNPH) and a predictive value for shunt responsiveness in a multicenter study (Study of Idiopathic Normal-Pressure Hydrocephalus on Neurological Improvement: SINPHONI). METHODS: Eighty-four iNPH patients underwent shunt operations using MRI selection criteria from the SINPHONI and were subjected to CBF single photon emission computed tomography (SPECT). The perfusion patterns on SPECT were classified: anterior-dominant CBF reduction type (A type), posterior-dominant CBF reduction type (P type), and mixed or diffuse CBF reduction type (M type). The predictive value of the CBF pattern for favorable shunt outcome was evaluated. RESULTS: Favorable outcomes were obtained in 76% (64/84) of patients, and shunt responsiveness was achieved in 85% (71/84) of patients. Areas of severely reduced relative CBF were demonstrated around the corpus callosum and in the sylvian fissure area, which included the effects of dilatations of the ventricles and sylvian fissures and relatively increased perfusion in the medial and lateral frontal, parietal, and occipital areas at high convexity. Forty-nine (58%) cases were A type, 25 (30%) cases were M type, and 10 (12%) cases were P type. A, M, and P type cases exhibited 83, 84, and 90% positive predictive values for shunt responsiveness, respectively. Mean modified Rankin scale and Mini-Mental State Examination scores of the A type group were significantly better than those of other groups. CONCLUSION: The iNPH patients showed various patterns of CBF reduction, but there was no significant difference in the predictive value among the three patterns, though CBF reduction patterns may suggest a severe condition of iNPH.

Drug design and synthesis of a novel kappa opioid receptor agonist with an oxabicyclo[2.2.2]octane skeleton and its pharmacology.

A conformational analysis of kappa opioid receptor agonists, TRK-820 and U-50,488H indicated an active conformation of TRK-820 in which the C-ring was in the boat form with the 14-OH interacting with the amide nitrogen. Based on the obtained active conformation of TRK-820, we designed and synthesized a novel kappa agonist KNT-63 with oxabicyclo[2.2.2]octane skeleton. KNT-63 showed profound antinociceptive effects via the kappa receptor which were as potent as that of TRK-820.

Renal circulatory effects of acetazolamide in patients with essential hypertension.

BACKGROUND: Reports indicate that acetazolamide (ACZ) induces the vasodilation of all vessels in animal models, as well as in small and medium kidney vessels in animal models. However, the effect of ACZ on the renal circulation of patients with essential hypertension remains unknown. In this study we examined the effects of a carbonic anhydrase inhibitor, acetazolamide (ACZ), on the renal circulation of patients with essential hypertension. METHODS: We directly infused 1000 mg of ACZ into the main renal arteries of 10 patients with essential hypertension who had undergone cardiac catheterization. We then evaluated the effects of ACZ upon heart rate, renal artery blood pressure (BP), renal artery cross-sectional area, renal Doppler blood flow velocity, renal blood flow (RBF), and renal vascular resistance (RVR). RESULTS: The infusion of ACZ was not associated with any significant changes in heart rate or in systolic or diastolic BP. However, the velocity-time integral was increased by 11.1% +/- 7.2%, from 17.6 +/- 1.8 to 20.0 +/- 3.7 cm (P = .009); RBF was increased by 39% +/- 21%, from 300 +/- 43 to 422 +/- 96 mL/min/m(2) (P = .002); and RVR was reduced by 38% +/- 20% from 24,351 +/- 2,291 to 17,651 +/- 2,731 dynes.sec.cm(-5) (P < .01). In contrast the cross-sectional area of the renal artery did not change. CONCLUSIONS: The results of the present study demonstrated that ACZ has a potent vasodilatory effect on the renal circulation of patients with essential hypertension, leading to an obvious decrease in RVR and an increase in RBF.

Circulating CD34-positive cells provide an index of cerebrovascular function.

BACKGROUND: Increasing evidence points to a role for circulating endothelial progenitor cells, including populations of CD34- and CD133-positive cells present in peripheral blood, in maintenance of the vasculature and neovascularization. Immature populations, including CD34-positive cells, have been shown to contribute to vascular homeostasis, not only as a pool of endothelial progenitor cells but also as a source of growth/angiogenesis factors at ischemic loci. We hypothesized that diminished numbers of circulating immature cells might impair such physiological and reparative processes, potentially contributing to cerebrovascular dysfunction. METHODS AND RESULTS: The level of circulating immature cells, CD34-, CD133-, CD117-, and CD135-positive cells, in patients with a history of atherothrombotic cerebral ischemic events was analyzed to assess possible correlations with the degree of carotid atherosclerosis and number of cerebral infarctions. There was a strong inverse correlation between numbers of circulating CD34- and CD133-positive cells and cerebral infarction. In contrast, there was no correlation between the degree of atherosclerosis and populations of circulating immature cells. Analysis of patients with cerebral artery occlusion revealed a significant positive correlation between circulating CD34- and CD133-positive cells and regional blood flow in areas of chronic hypoperfusion. CONCLUSIONS: These results suggest a possible contribution of circulating CD34- and CD133-positive cells in maintenance of the cerebral circulation in settings of ischemic stress. Our data demonstrate the utility of a simple and precise method to quantify circulating CD34-positive cells, the latter providing a marker of cerebrovascular function.

Cerebral hemodynamic evaluation using perfusion-weighted magnetic resonance imaging: comparison with positron emission tomography values in chronic occlusive carotid disease.

BACKGROUND AND PURPOSE: Perfusion-weighted magnetic resonance imaging (PWI) is a reliable and semiquantitative method for estimating cerebral hemodynamics. We sought to evaluate the potential of PWI for assessing cerebral blood flow (CBF) and metabolism compared with positron emission tomography (PET) in patients with chronic occlusive carotid disease. METHODS: In 24 patients with chronic unilateral occlusive carotid disease, time-to-peak (TTP) delay (TTP-D) measured by PWI was compared with CBF, cerebral blood volume (CBV), and oxygen extraction fraction (OEF) obtained by PET. TTP indicates the time from the start of PWI to the bolus peak. TTP-D indicates the difference in TTP values between the occlusive and contralateral hemispheres. TTP-D was compared between patients with normal and reduced CBF/CBV and also between patients with normal and elevated OEF. RESULTS: TTP-D in patients with reduced CBF/CBV was significantly longer than that in patients with normal CBF/CBV (3.4+/-1.8 versus 1.4+/-0.7 seconds; P<0.001). In the patients with reduced CBF/CBV, TTP-D correlated with OEF significantly (r=0.710, P<0.0001). TTP-D in patients with elevated OEF was significantly longer than that in patients with normal OEF (4.8+/-2.2 versus 2.0+/-0.9 seconds; P<0.01). In all 5 patients with TTP-D >or=4 seconds, OEF was elevated markedly. CONCLUSIONS: TTP-D >or=4 seconds is considered to indicate a high risk of hemodynamic failure. The measurement of TTP-D by PWI appears to provide important clinical information for evaluating cerebral hemodynamics in chronic occlusive carotid disease.

A theoretical model of oxygen delivery and metabolism for physiologic interpretation of quantitative cerebral blood flow and metabolic rate of oxygen.

The coupling of cerebral blood flow (CBF) and metabolic rate of oxygen (CMRO2) during physiologic and pathophysiologic conditions remains the subject of debate. In the present study, we have developed a theoretical model for oxygen delivery and metabolism, which describes the diffusion of oxygen at the capillary-tissue interface and the nonlinear nature of hemoglobin (Hb) affinity to oxygen, allowing a variation in simple-capillary oxygen diffusibility, termed "effective oxygen diffusibility (EOD)." The model was used to simulate the relationship between CBF and CMRO2, as well as oxygen extraction fraction (OEF), when various pathophysiologic conditions were assumed involving functional activation, ischemia, hypoxia, anemia, or hypo- and hyper-capnic CBF variations. The simulations revealed that, to maintain CMRO2 constant, a variation in CBF and Hb required active change in EOD. In contrast, unless the EOD change took place, the brain allowed small but significant nonlinear change in CMRO2 directly dependent upon oxygen delivery. Application of the present model to quantitative neuroimaging of CBF and CMRO2 enables us to evaluate the biologic response at capillary level other than Hb- and flow-dependent properties of oxygen transport and may give us another insight regarding the physiologic control of oxygen delivery in the human brain.

Visualization of interscapular brown adipose tissue using (99m)Tc-tetrofosmin in pediatric patients.

UNLABELLED: Brown adipose tissue (BAT) is a site of nonshivering thermogenesis in mammals. The mitochondria of BAT operate in an uncoupled mode and increase fatty acid oxidation to produce heat at birth. Thus, the BAT of human infants and children contains more active mitochondria than that of adults. We surmised that because (99m)Tc-tetrofosmin can be absorbed by functional mitochondria in the myocardium and in tumor cells, it could reveal mitochondrial function in BAT. METHODS: Between January 1999 and December 2002, we retrospectively analyzed 385 consecutive studies of (99m)Tc-tetrofosmin uptake in pediatric patients with cardiac disorders. All patients with symmetric (99m)Tc-tetrofosmin accumulation within the neck and shoulder region according to planar images were selected, and the features of the uptake were analyzed. RESULTS: Increased symmetric (99m)Tc-tetrofosmin uptake in the interscapular BAT was a typical profile of 65 of the 385 patients (17%). The frequency of (99m)Tc-tetrofosmin uptake was significantly higher in winter than in spring or summer (P < 0.05) and prominent in newborns. The frequency peaked between 0 and 2 y of age and then declined with age. CONCLUSION: Gamma-camera imaging with (99m)Tc-tetrofosmin can reveal interscapular BAT distribution in infants and children in terms of mitochondrial activity.

Gated myocardial SPECT to predict response to beta-blocker therapy in patients with idiopathic dilated cardiomyopathy.

UNLABELLED: Because of the difficulty of predicting the response of patients with idiopathic dilated cardiomyopathy (IDC) to beta-blocker therapy, this study was performed to evaluate whether gated myocardial SPECT (gated SPECT) could be useful for predicting that response. METHODS: We performed gated SPECT with (99m)Tc-sestamibi on 38 patients with IDC before treatment with a beta-blocker and standard medication. Perfusion abnormalities, left ventricular (LV) function, and spheric distortion were assessed by a quantitative software program. RESULTS: We classified patients into 2 groups according to improvement in LV function after 4 mo of therapy. The groups consisted of 16 poor responders whose LV ejection fraction (LVEF) increased less than 10% and 22 good responders whose LVEF increased by 10% or more. The patient characteristics before therapy, including LV volume and LVEF, did not significantly differ between the 2 groups, but the size of the myocardial perfusion defect and spheric distortion were significantly greater in poor responders than in good responders. CONCLUSION: Gated SPECT, by allowing simultaneous assessment of perfusion, function and geometry, might be useful for predicting the response of patients with IDC to beta-blocker therapy.

Quantitative analysis of lung perfusion in patients with primary pulmonary hypertension.

UNLABELLED: The aim of this study was to evaluate quantitatively the heterogeneity of lung perfusion scans in patients with primary pulmonary hypertension (PPH) and to compare it with the severity of disease. METHODS: Lung perfusion scans were obtained on 22 patients with PPH and 12 age-matched control subjects. The perfused area rates (PARs) were calculated by dividing the lung area in each 10% threshold width from 10% to 100% of maximal counts by total lung area. The total absolute difference in the PAR between each patient and the mean control value was assumed as the perfusion index of the lung (P index). The P index was compared with hemodynamic parameters and the right ventricular ejection fraction (RVEF), including 7 patients who received long-term vasodilator therapy. RESULTS: The P index correlated significantly with mean pulmonary arterial pressure (P < 0.001) and RVEF (P < 0.05). In patients with vasodilator therapy, the P index was improved significantly after therapy (P < 0.05) and was associated with a reduction in pulmonary arterial pressure. CONCLUSION: Quantitative assessment of lung perfusion irregularity might provide useful information about the severity of disease and the effect of therapy in addition to the routine visual representation.

Topographic distribution of misery perfusion in relation to internal and superficial borderzones.

BACKGROUND AND PURPOSE: Whether misery perfusion (MP) commonly accompanies brain borderzones (BZs) in patients with major cerebral artery occlusion remains unclear. We elucidated topographic patterns of chronic hemodynamic failure in such patients. METHODS: Twenty-four patients with unilateral occlusion or severe stenosis (>75% in diameter) of the internal carotid artery (ICA) or middle cerebral arterial (MCA) trunk with minimal or no infarct underwent PET with (15)O-labeled gas inhalation. Mean cerebral blood flow (CBF), cerebral blood volume (CBV), cerebral metabolic rate of oxygen, oxygen extraction fraction (OEF), and CBV/CBF ratio were determined in the superficial BZs, internal BZ, and MCA territory excluding BZs. Values in BZs were standardized and compared with those in controls. Topographic distributions of regions with OEF greater than that in controls were determined. RESULTS: Values in patients and controls were not significantly different. Topographic distributions included matched perfusion in 10 patients, MP in only the ipsilateral internal BZ in four, MP in both ipsilateral internal and superficial BZs in two, MP in the ipsilateral MCA territory including BZs in one, MP in the ipsilateral MCA territory including BZs and contralateral BZs in two, and MP in the ipsilateral MCA territories including BZs in five. CONCLUSION: Only 25% of the patients had MP localized in affected BZs Although localized MP more frequently accompanied the internal BZ than other regions, no patient had elevated OEF in the superficial BZ alone. These results are inconsistent with clinical observations that 80% of BZ infarctions develop superficially. Thus, hemodynamic mechanisms may not cause most superficial BZ infarctions.

Acetazolamide challenge test using semiquantitative 123 I-IMP SPECT for detection of cerebral misery perfusion.

The accuracy of the acetazolamide (ACZ) challenge test using semiquantitative SPECT in detecting Stage II hemodynamic failure, i.e. cerebral misery perfusion, in patients so diagnosed with PET has yet to be determined. This study was carried out in 53 patients who had a unilateral occlusion or severe stenosis of their cerebral artery. Asymmetry index (AI) was used to determine relative CBF distribution on each SPECT image. DeltaAI (regional vasodilatory capacity) values were compared with the values of several PET parameters. We also repeated SPECT and PET studies in 15 patients after a mean interval of 2.6 years from entry. The correlation between changes in DeltaAI and PET data were examined. The DeltaAI value closely correlated with the ipsilateral OEF value (r=-0.64, p<0.001). Sensitivity-specificity curve analysis revealed that the optimal cut-off value for detecting Stage II hemodynamic failure was -13.5% of DeltaAI, at which point the diagnostic accuracy was 82%. In follow-up studies, changes in DeltaAI correlated significantly with changes in OEF values (r=-0.68, p<0.01). Semiquantitative SPECT examination with ACZ challenge detects Stage II hemodynamic failure with a diagnostic accuracy of 82%. An improvement in reduced vasodilatory capacity as determined by SPECT coincides with a reduction in OEF values.

Cerebral hemodynamics and metabolism in patients with cerebral arteriovenous malformations: an evaluation using positron emission tomography scanning.

OBJECT: The purpose of this study was to evaluate cerebral hemodynamic and metabolic features in patients with arteriovenous malformations (AVMs) by using positron emission tomography (PET) scanning. METHODS: Twenty-four patients with supratentorial cerebral AVMs participated in PET studies in which 15O inhalation steady-state methods were used. The authors recorded the values of regional cerebral blood flow (rCBF), regional cerebral blood volume (rCBV), the regional oxygen extraction fraction (rOEF), and the regional cerebral metabolic rate of O2 (rCMRO2) at three designated regions of interest (ROIs) in each patient. These ROIs included perilesional (ROI-p), ipsilateral remote (ROI-i), and contralateral symmetrical (ROI-c) brain regions. To identify the factors that exert a direct effect on the hemodynamics of brains affected by AVM, we also separated the lesions according to their size and flow type shown on angiograms, and grouped the patients according to the presence or absence of progressive neurological deficits. We then compared the PET parameters at different ROIs in individual patients and evaluated the mean values obtained for all 24 patients according to AVM flow type and size, and the presence or absence of progressive neurological deficits. CONCLUSIONS: Overall, mean rCBV and rOEF values were significantly higher in ROI-p than in ROI-c (p = 0.00046 and p = 0.015, respectively). No significant differences were seen between the ROI-i and ROI-c with respect to rCBF, rCBV, and rOEF. Mean rCMRO2 values were similar in the three ROIs; however, the mean rCBF was significantly lower in the ROI-p than in the ROI-c in patients with high-flow AVMs (p = 0.019), large AVMs (p = 0.017), and progressive neurological deficits (p = 0.021). Furthermore, the mean rOEF values were significantly higher in the ROI-p than in the ROI-c in patients with high-flow AVMs (p = 0.005), large AVMs (p = 0.019), and progressive neurological deficits (p = 0.017). The PET studies revealed hemodynamic impairment characterized by decreased rCBF and increased rOEF and rCBV values in the ROI-p of patients with large, high-flow AVMs regardless of whether they exhibited progressive neurological deficits.

Acetazolamide assisted Tc-99m MAG3 renography to assess renal blood flow reserve.

OBJECTIVE: The present study examines whether or not baseline and acetazolamide (ACZ) Tc-99m MAG3 renography can assess renal blood flow reserve. METHODS: Renography proceeded for 50 min after sequential injections of 370 MBq Tc-99m MAG3 for baseline renography and 10 min after a 1,000 mg injection of ACZ for ACZ renography. Effective renal plasma flow of renal cortex (cERPF) in each kidney and the percentage change in cERPF of those parameters (deltaERPF) were obtained before and after the administration of ACZ in 10 subjects without hypertension or diabetes (normal group), in 10 with essential hypertension (hypertensive group) and in 10 who had Type 2 diabetes with hypertension (diabetic group). A placebo test was performed in the 10 without hypertension or diabetes using distilled water instead of ACZ (placebo group). RESULTS: The placebo test performed in the 10 without hypertension or diabetes using distilled water instead of ACZ indicated that the parameter variance between the two types of renogram was below 3.2%. The cERPF of baseline and ACZ Tc-99m MAG3 renography and deltaERPF in the normal, hypertensive and diabetic groups were 89 +/- 10 and 110 +/- 10 ml/min, 89 +/- 14 and 117 +/- 22 ml/min, 100 +/- 23 and 112 +/- 23 ml/min, respectively, and 24.5 +/- 13.5%, 26.0 +/- 9.7% and 12.3 +/- 11.1%, respectively. The difference in the cERPF value was significant in the normal and hypertensive groups whereas this did not change in the diabetic group before or after ACZ administration. CONCLUSIONS: We suggested that the deltaERPF determined by baseline and ACZ Tc-99m MAG3 renography is a useful parameter for assessing renal blood flow reserve.

Evaluation of a commercial PET tomograph-based system for the quantitative assessment of rCBF, rOEF and rCMRO2 by using sequential administration of 15O-labeled compounds.

The purpose of this study was to develop a reliable and practical strategy that generates quantitative CBF and OEF maps accurately from PET data sets obtained with 15O-tracers. Sequential sinogram data sets were acquired after the administration of 15O-tracers, and combined single-frame images were obtained. The delay time between sampled input function and the brain was estimated from the H2(15)O study with the whole brain and the arterial time-activity curves (TACs). The whole-brain TACs were obtained from the reconstructed images (image-base method) and the sinogram data (sinogram-base method). Six methods were also evaluated for the dead-time and decay correction procedures in the process of generating a single-frame image from the dynamic sinogram. The estimated delay values were similar with both the sinogram-based and image-based methods. A lumped correction factor to a previously added single-frame sinogram caused an underestimation of CBF, OEF and CMRO2 by 16% at maximum, as compared with the correction procedure for a short sinogram. This suggested the need for a dynamic acquisition of a sinogram with a short interval. The proposed strategy provided an accurate quantification of CBF and OEF by PET with 15O-tracers.

[Acute brain infarction clearly demonstrated by using the three-dimensional stereotactic surface projections (3D-SSP) of 99mTc-HMPAO SPECT].

We report a 66-year-old ambidextrous man who suddenly developed aphasia seven days after coronary artery bypass grafting surgery. CT scan performed 30 minutes after onset failed to depict acute ischemic lesion. MR imaging could not be performed because of intrathoracic wires. An emergent angiogram demonstrated filling defect in the right precentral artery territory. Transaxial images of 99mTc-HMPAO SPECT showed hypoperfusion areas in the right frontal and left temporo-parietal regions. Using the three-dimensional stereotactic surface projections (3D-SSP) of 99mTc-HMPAO SPECT, we could clearly detect a sharply demarcated ischemic area which was later confirmed to be an infracted lesion in the follow-up CT. 3D-SSP, thus, appears to be useful for identifying acute infarction in such cases that MRI is impermissible.

Human nitrilase-like protein does not catalyze the hydrolysis of vildagliptin.

Nitrilase, which is found in plants and many types of bacteria, is known as the enzyme that catalyzes hydrolysis of a wide variety of nitrile compounds. While human nitrilase-like protein (NIT), which is a member of the nitrilase superfamily, has two distinct isozymes, NIT1 and NIT2, their function has not been well understood. In this study, we investigated whether human NIT1 and NIT2 are involved in the hydrolysis of drugs using vildagliptin as a substrate. We performed Western blot analysis using human liver samples to examine protein expression of human NIT in the liver, finding that human NIT1 and NIT2 were highly expressed in the liver cytosol. We established stable single expression systems of human NIT1 and NIT2 in HEK293 cells to clarify the contribution of human NIT to hydrolysis of vildagliptin. Although the formation of a carboxylic acid metabolite of vildagliptin (M20.7) was observed in human liver samples, M20.7 was not formed by incubating vildagliptin with HEK293 cells expressing human NIT1 or NIT2. This suggests that human NIT1 or NIT2 is not involved in the metabolism of vildagliptin. Further investigation using other drugs is needed to clarify the contribution of human NIT to drug metabolism.