Spectral entropy monitoring allowed lower sevoflurane concentration and faster recovery in children.

BACKGROUND: Anesthetic titration using spectral entropy monitoring reduces anesthetic requirements and shortens recovery in adult surgical patients. This study was performed to evaluate the effect of entropy monitoring on end-tidal sevoflurane concentration and recovery characteristics in pediatric patients undergoing sevoflurane anesthesia. METHODS: Seventy-eight children (aged 3-12 years) scheduled for a tonsillectomy and/or an adenoidectomy were randomly divided into one of two groups: standard practice (Standard) or entropy-guided (Entropy). In the Standard group, sevoflurane was adjusted to maintain the heart rate and systolic blood pressure (BP) within 20% of the baseline values. In the Entropy group, sevoflurane was adjusted to achieve a state entropy of 40-50. We compared the entropy values, end-tidal sevoflurane concentration and recovery times between groups. RESULTS: During maintenance of anesthesia, the entropy and BP values were higher in the Entropy group (P<0.05). The end-tidal sevoflurane concentration during maintenance was lower in the Entropy group (2.2 (0.3) vol%) compared with the Standard group (2.6 (0.4) vol%) (P<0.05). Recovery times were faster in the Entropy group (P<0.05). CONCLUSIONS: Compared with standard practice, we found that entropy-guided anesthetic administration was associated with a reduced sevoflurane concentration and a slightly faster emergence and recovery in 3-12-year-old children.

Flowering time quantitative trait Loci analysis of oilseed brassica in multiple environments and genomewide alignment with Arabidopsis.

Most agronomical traits exhibit quantitative variation, which is controlled by multiple genes and are environmentally dependent. To study the genetic variation of flowering time in Brassica napus, a DH population and its derived reconstructed F(2) population were planted in 11 field environments. The flowering time varied greatly with environments; 60% of the phenotypic variation was attributed to genetic effects. Five to 18 QTL at a statistically significant level (SL-QTL) were detected in each environment and, on average, two new SL-QTL were discovered with each added environment. Another type of QTL, micro-real QTL (MR-QTL), was detected repeatedly from at least 2 of the 11 environments; resulting in a total of 36 SL-QTL and 6 MR-QTL. Sixty-three interacting pairs of loci were found; 50% of them were involved in QTL. Hundreds of floral transition genes in Arabidopsis were aligned with the linkage map of B. napus by in silico mapping; 28% of them aligned with QTL regions and 9% were consistent with interacting loci. One locus, BnFLC10, in N10 and a QTL cluster in N16 were specific to spring- and winter-cropped environments respectively. The number of QTL, interacting loci, and aligned functional genes revealed a complex genetic network controlling flowering time in B. napus.

Effect of remifentanil on oculocardiac reflex in paediatric strabismus surgery.

BACKGROUND: Rapidly acting narcotics enhance the degree of bradycardia due to the oculocardiac reflex (OCR) elicited by extraocular muscle (EOM) tension during strabismus surgery. We evaluated and compared the effects of remifentanil and sevoflurane on OCR during paediatric strabismus surgery. METHODS: One hundred and twenty children, 1-9 years old, undergoing elective strabismus surgery, were randomly assigned to receive sevoflurane or remifentanil. No anticholinergic prophylaxis was administered. Anaesthesia was induced using ketamine 1.0 mg/kg or midazolam 0.15 mg/kg with 66% N(2)O in O(2). Laryngeal mask airways were placed with rocuronium 0.5 mg/kg. Anaesthesia was maintained with sevoflurane 2.0-3.0 vol% with 66% N(2)O in O(2) or remifentanil 0.75 mug/kg over 1 min and followed by the continuous infusion of remifentanil 0.5 mug/kg/min with 66% N(2)O in O(2). Heart rate (HR) and blood pressure (BP) were measured and compared. OCR was defined as a reduction in HR of >20% induced by traction of an EOM. RESULTS: During anaesthesia, HR and BP were maintained at a lower level in the remifentanil group than in the sevoflurane group (each, P<0.05). The mean percent change in HR (-23.3+/-17.0% vs. -11.2+/-13.0%; P<0.05) and the incidence of OCR (58.3% vs. 28.3%; P<0.05) following traction of an EOM were higher in the remifentanil group than in the sevoflurane group. CONCLUSIONS: Remifentanil enhanced the degree of bradycardia due to OCR as compared with sevoflurane during paediatric strabismus surgery.

Laparoscopic wedge resection for gastric GIST: long-term follow-up results.

AIM: Gastrointestinal stromal tumor (GIST) is the most common mesenchymal neoplasm of the gastrointestinal tract. Recently, many investigations have been conducted on various aspects of laparoscopic surgery for gastric GIST. However, no study has provided long-term follow up results of laparoscopic surgery for gastric GIST. The aims of this study were to assess the feasibility and safety of laparoscopic surgery for gastric GIST and to evaluate the oncologic validity of the procedure. MATERIALS AND METHODS: Between January 1998 and August 2005, 51 patients with submucosal tumor of the stomach were treated by laparoscopic surgery at our institution. Of 51 patients, 23 patients were confirmed as gastric GIST by immunohistochemistry (CD 117, c-kit gene product). Patients' clinicopathologic characteristics, operative outcomes, postoperative complications, and follow-up findings were analyzed retrospectively. RESULTS: The mean age of patients was 59.7 years, and 12 patients were women. Twelve patients (47%) presented with epigastric pain. The mean tumor size was 4.2+/-2.1 cm, and most tumors were located in the upper stomach (52.2%). The mean operative time was 104.3 min. No case of open conversion, reoperation and operative mortality occurred in the present study. Most patients had very low and low risk (60.6%), while only two patients had high risk malignancy. During a median follow-up period of 61 months (range, 7-98 months), there have been no recurrences or metastases. CONCLUSION: Laparoscopic wedge resection for gastric GIST is safe, and oncologically and technically feasible in the hands of an experienced laparoscopic gastric surgeon.

New insight of amino acid-based dialysis solutions.

Malnutrition is a major complication of peritoneal dialysis (PD) and is associated with increased morbidity and mortality. Daily losses of proteins and amino acids (AAs) into dialysate contribute to this problem. Previous metabolic balance study demonstrated that treatment with 1.1% AA-based dialysis solution is safe and may improve protein malnutrition in continuous ambulatory peritoneal dialysis (CAPD) patients ingesting low protein intake. Other prospective studies also showed that AA solution can provide nutritional benefit for malnourished PD patients resulting in a significant improvement in some biochemical and/or anthropometric nutritional parameters. However, there are other studies showing no particular improvement in nutritional parameters after long-term use of AA solution. This may be related to the differences in the study design, sample size, methods used to assess nutritional status, and other factors such as dietary intake and comorbidities of study subjects. Published data will be reviewed to further emphasize the nutritional benefit of long-term use of AA solution in malnourished PD patients along with a brief discussion on the various reasons that may partly explain the different study results. We will also present the results of a longitudinal observational study evaluating changes in nutritional parameters following use of one exchange of 1.1% AA solution in malnourished Korean PD patients. A significant improvement of somatic protein status such as lean body mass (LBM) and hand grip strength was observed. No significant change in serum albumin level was noted. Patients with a positive estimated coefficient for LBM in the fitted regression model to the repeated observations over 1 year were classified as responders and patients with neutral or negative coefficient were considered as non-responders. Thirty-one out of 43 malnourished patients (72%) showed nutritional benefit based on the change of LBM. Hand grip strength and back lift strength were significantly higher in responders at baseline. Other baseline parameters did not differ between the two groups.

Role of peripheral sigma-1 receptors in ischaemic pain: Potential interactions with ASIC and P2X receptors.

BACKGROUND: The role of peripheral sigma-1 receptors (Sig-1Rs) in normal nociception and in pathologically induced pain conditions has not been thoroughly investigated. Since there is mounting evidence that Sig-1Rs modulate ischaemia-induced pathological conditions, we investigated the role of Sig-1Rs in ischaemia-induced mechanical allodynia (MA) and addressed their possible interaction with acid-sensing ion channels (ASICs) and P2X receptors at the ischaemic site. METHODS: We used a rodent model of hindlimb thrombus-induced ischaemic pain (TIIP) to investigate their role. Western blot was performed to observe changes in Sig-1R expression in peripheral nervous tissues. MA was measured after intraplantar (i.pl.) injections of antagonists for the Sig-1, ASIC and P2X receptors in TIIP rats or agonists of each receptor in naïve rats. RESULTS: Sig-1R expression significantly increased in skin, sciatic nerve and dorsal root ganglia at 3 days post-TIIP surgery. I.pl. injections of the Sig-1R antagonist, BD-1047 on post-operative days 0-3 significantly attenuated the development of MA during the induction phase, but had no effect on MA when given during the maintenance phase (days 3-6 post-surgery). BD-1047 synergistically increased amiloride (an ASICs blocker)- and TNP-ATP (a P2X antagonist)-induced analgesic effects in TIIP rats. In naïve rats, i.pl. injection of Sig-1R agonist PRE-084 alone did not produce MA; but it did induce MA when co-administered with either an acidic pH solution or a sub-effective dose of αßmeATP. CONCLUSION: Peripheral Sig-1Rs contribute to the induction of ischaemia-induced MA via facilitation of ASICs and P2X receptors. Thus, peripheral Sig-1Rs represent a novel therapeutic target for the treatment of ischaemic pain.

Mucins secreted by cell lines derived from colorectal mucinous carcinoma and adenocarcinoma.

Mucinous (colloid) carcinoma and well- to moderately-differentiated adenocarcinoma of the colon differ in the pattern and the amount of mucin secretion and perhaps in their behaviour and clinical outcome. To ascertain why these differences exist and to elucidate the mechanisms of tumour progression, we examined two model human cell lines derived from colorectal mucinous carcinoma (C1a) and moderately differentiated adenocarcinoma (HM3) which show typical pathological and mucin staining patterns of the respective type of carcinomas to nude mouse tumour xenografts. Specifically, we sought to determine if there were quantitative and qualitative differences in mucin synthesis, in mucin gene expression and in biological properties between the two model cell lines. Northern blot analysis showed that MUC2 mRNA levels were significantly higher in C1a cells compared with HM3 cells, while those of MUC3, -5 and -6 mRNA were lower. C1a cells secreted approximately five times more radiolabelled apomucin and 1.5 times more glycosylated apomucin than HM3 cells. When the carbohydrate side-chain length of secreted mucins by these cell lines were examined by beta-elimination followed by P4 column chromatography, C1a mucins had mostly short carbohydrate side-chains, while HM3 cells had predominantly longer side-chains. Western blot analysis of the cell homogenate showed higher expression of MUC2 apomucin and mucin-associated carbohydrate antigens, such as T, Tn and sialyl Tn, with decreased sialyl Le(x) expression in C1a cells compared with HM3. Immunohistochemical analysis of 35 colorectal adenocarcinoma and 25 mucinous colorectal carcinoma tissues also demonstrated increased MUC2 apomucin, T, Tn and sialyl Tn antigens in the mucinous cancer specimens. Examination of the biological properties of these cell lines showed that C1a cells had significantly higher in vitro invasive activity in assays of invasion and collagenase activity and significantly lower E-selectin binding and liver colonisation activities in nude mice. These results indicate that colorectal mucinous carcinoma cells differ considerably from colorectal adenocarcinoma cells, both qualitatively and quantitatively, in the pattern of mucin gene expression and in the synthesis and secretion of mucin. In addition, biological studies showed that mucinous carcinoma cells have a greater degree of invasiveness, but less liver colonising activity. These results suggest that the biological and mucin characteristics of mucinous carcinoma cells contribute to extensive local invasion through tissue stroma as the predominant mechanism of tumour progression, while the biological and mucin characteristics of well- to moderately-differentiated colorectal adenocarcinoma contribute to progression via distant metastasis formation.

Quantification of serotonin transporters in nonhuman primates using [(123)I]ADAM and SPECT.

UNLABELLED: We reported recently a highly selective radioligand, 2-([2-([dimethylamino]methyl)phenyl]thio)-5-[(123)I]iodophenylamine (ADAM), for SPECT imaging of serotonin transporters (SERT). In this article we describe the kinetic modeling of [(123)I]ADAM and its ability to quantitatively and reproducibly measure the concentrations of SERT in the nonhuman primate brain. We also investigate simplified models of tracer behavior that do not require invasive arterial blood sampling. METHODS: Three female baboons each underwent 3 [(123)I]ADAM SPECT studies. The studies consisted of a dynamic sequence of seventy-two 5-min scans after injection of 330 +/- 50 MBq (mean +/- SD) [(123)I]ADAM. Rapid arterial blood samples were obtained and corrected for the presence of labeled metabolites. Dynamic imaging and metabolite-corrected plasma data were analyzed using graphic analysis to give the distribution volumes (DVs) of different brain regions. DV ratios (DVRs) of target to cerebellum were derived and compared against a kinetic reference tissue model and simple target-to-background ratio. RESULTS: Averaged over all 9 scans, the mean DV in the midbrain was 4.86 +/- 1.06 mL/mL and the mean DV in the cerebellum was 2.25 +/- 0.48 mL/mL. The mean test-retest repeatability of the midbrain DV was 14.5%. The reference tissue model gave a mean midbrain DVR of 2.01 +/- 0.17 and correlated strongly with the DVR calculated from the full kinetic model (correlation coefficient [R(2)] = 0.94; P < 0.001), but with much improved repeatability (test-retest, 5.4%; intersubject variability, 5.2%). Similarly, the simple ratio method gave strong correlations with the full kinetic model (R(2) = 0.89; P < 0.001) and a test-retest of 7.6%. CONCLUSION: Accurate, repeatable quantification of SERT in the nonhuman primate brain is possible using kinetic modeling of dynamic [(123)I]ADAM SPECT scans. Simplified models, which do not require arterial blood sampling, gave accurate results that correlated strongly with the full kinetic model. The test-retest reliability of the simplified reference region models was excellent. Quantification of SERT is possible using full kinetic modeling and also with simpler reference region methods.

Mechanisms related to [18F]fluorodeoxyglucose uptake of human colon cancers transplanted in nude mice.

UNLABELLED: [18F]Fluorodeoxyglucose ([18F]FDG), a glucose analogue, has been widely used for tumor imaging. To investigate the mechanisms related to [18F]FDG uptake by tumors, an experiment involving nude mice was performed. METHODS: Human colon cancer cell lines SNU-C2A, SNU-C4 and SNU-C5 were transplanted to nude mice. Using immunohistochemical staining and Western blot, the expression of glucose transporter (Glut) isoforms (Glut-1 through -5) in xenografted tumors was analyzed. For the analysis of messenger ribonucleic acid (mRNA) expression, reverse-transcription polymerase chain reaction and Northern blot were used and the enzyme activity of hexokinase in cancer tissues was measured by continuous spectrophotometric rate determination. RESULTS: [18F]FDG uptake in SNU-C4 and SNU-C5 cells was higher than in normal colon cells. Among these cells and xenografted tumors, SNU-C5 showed the highest level of [18F]FDG uptake, followed by SNU-C4 and SNU-C2A. An immunostaining experiment showed intense staining of Glut-1 in SNU-C5 tumors but somewhat faint staining in SNU-C4. SNU-C5 tumors also showed positive staining with Glut-3, although this was not the case with SNU-C2A and SNU-C4. Western blot analysis showed the expression of Glut-1 and Glut-3 in all tumors. Experiments involving Northern blot analysis and reverse-transcription polymerase chain reaction confirmed the overexpression of Glut-1 mRNA in all tumors, with the highest level in SNU-C5. The level of Glut-3 mRNA was also elevated in SNU-C5 tumors but not in SNU-C2A and SNU-C4. The enzyme activity of hexokinase did not vary among different tumors. CONCLUSION: Gluts, especially Glut-1, are responsible for [18F]FDG uptake in a nude mouse model of colon cancer rather than hexokinase activity. Increased numbers of glucose transporters at the plasma membrane of cancer cells is attributed to an increased level of transcripts of glucose transporter genes and may be a cause of increased [18F]FDG uptake, at least in colon cancer tumors.

Biological properties and expression of mucins in 5-fluorouracil resistant HT29 human colon cancer cells.

We have previously reported that HT29 human colon cancer cells selected by adaptation to 5-fluorouracil (5FU) (HT29-5FU cells) express increased levels of a major intestinal mucin MUC2 mRNA compared with parental HT29 cells. In this study, we examined in detail the changes in synthesis and secretion of mucin that occur in these cells and accompanying changes in the expression of cancer associated mucin related carbohydrate antigens and cell lineage associated biochemical markers. We further investigated their relationship to biological properties of cells. Northern blot analysis revealed a markedly increased level of MUC2 mRNA but no significant change in the mRNA levels of other mucins in HT29-5FU cells compared with parental HT29 cells. Labeling with radiolabeled precursors demonstrated increased synthesis and secretion of mucin glycoproteins by HT29-5FU cells. Immunoblot analysis showed a higher expression of mucin associated carbohydrate antigens such as T, Tn, sialyl Tn, sialyl Lea, sialyl Lex and non-O-acetylated sialic acid concomitant with significant increases in the expression of goblet cell lineage marker, MUC2 apomucin and a panepithelial cell marker, carcinoembryonic antigen. HT29-5FU cells showed significantly higher adhesion to E-selectin and to matrigel and in vitro invasive properties and significantly increased liver colonization capacity in nude mice following splenic vein injection. Nude mouse xenograft tumors produced by HT29-5FU cells showed a greater degree of differentiation, consisting of mucin secreting glands than those produced by parental HT29 cells. These results indicate that predominantly colonic type mucin, MUC2, has been selectively induced in HT29-5FU cells and that altered regulation of mucin genes associated with altered synthesis and secretion of mucin glycoproteins and the degree of differentiation in cancer cells may be responsible for the altered biological properties of these cells.

Nicotine enhances morphine- and beta-endorphin-induced antinociception at the supraspinal level in the mouse.

The effect of nicotine administered supraspinally on antinociception induced by supraspinally administered opioids was examined in ICR mice. The intracerebroventricular (i.c.v.) injection of nicotine alone at doses from 1 to 12 micrograms produced only a minimal inhibition of the tail-flick response. Morphine (0.2 micrograms), beta-endorphin (0.1 microgram), D-Pen2.5-enkephalin (DPDPE; 0.5 microgram), trans-3,4-dichloro-N-methyl-N-[2-(1-pyrrolidinyl) cyclohexyl] benzeocetamide (U50, 488H; 6 micrograms) caused only slight inhibition of the tail-flick response. Nicotine dose dependently enhanced inhibition of the tail-flick response induced by i.c.v. administered morphine (0.2 microgram) or beta-endorphin (0.1 microgram). The degree of enhancing effect of nicotine toward beta-endorphin-induced inhibition of the tail-flick response was greater than toward morphine-induced inhibition of the tail-flick response. However, i.c.v. administered nicotine at the same doses was not effective in enhancing the inhibition of the tail-flick response induced by DPDPE (0.5 microgram) or U50, 488H (6 micrograms) administered i.c.v. Our results suggest that stimulation of supraspinal nicotinic receptors may enhance antinociception induced by morphine (a mu-opioid receptor agonist) and beta-endorphin (an epsilon-opioid receptor agonist) administered supraspinally. However, the activation of nicotinic receptors at supraspinal sites may not be involved in enhancing the antinociception induced by DPDPE (a delta-opioid receptor agonist) or U50, 488H (a kappa-opioid receptor agonist) administered supraspinally.

Intrathecally injected nicotine enhances the antinociception induced by morphine but not beta-endorphin, D-Pen2,5-enkephalin and U50,488H administered intrathecally in the mouse.

The effect of nicotine injected intrathecally (i.t.) on the inhibition of the tail-flick response induced by morphine, beta-endorphin, D-Pen2,5-enkephalin (DPDPE), or [(trans-3,4-dichloro-N-methyl-N-[2-(1-pyrrolidinyl)cyclohexyl] benzeocetamide)] (U50,488H) administered i.t. was studied in ICR mice. The i.t. injection of nicotine alone at doses from 1 to 12 microg produced only a minimal inhibition of the tail-flick response. Morphine (0.2 microg), beta-endorphin (0.1 microg), DPDPE (0.5 microg) or U50,488H (6 microg) caused only slight inhibition of the tail-flick response. Nicotine injected i.t. dose dependently enhanced the inhibition of the tail-flick response induced by i.t. administered morphine (0.2 microg). However, i.t. injected nicotine at the same doses was not effective in enhancing the inhibition of the tail-flick response induced by beta-endorphin, DPDPE, or U50,488H administered i.t. Our results suggest that stimulating nicotinic receptors located in the spinal cord may enhance the antinociception induced by morphine administered spinally. However, the activation of nicotinic receptors at the spinal level may not be involved in modulating the antinociception induced by beta-endorphin, DPDPE, and U50,488H administered spinally.

Effects of intrathecal injection of nimodipine, omega-conotoxin GVIA, calmidazolium, and KN-62 on the antinociception induced by cold water swimming stress in the mouse.

The present study was designed to determine if spinal calcium channels, calmodulin, and calcium/calmodulin-dependent protein kinase II were involved in the production of antinociception induced by cold water swimming stress (CWSS). The effects of intrathecal (i.t.) injection of nimodipine, omega-conotoxin GVIA, calmidazolium, or (S)-5-isoquinolinesulfonic acid, 4-[2-[(5-isoquinolinyl-sulfonyl)methylamino]-3-oxo-3-(4-phenyl-1-piperaz inyl)-propyl]phenyl ester (KN-62) on CWSS-induced antinociception were studied in ICR mice. The antinociception was assessed by the tail-flick test. CWSS produced inhibition of the tail-flick response. Various doses of nimodipine (10-40 ng), omega-conotoxin GVIA (5-40 ng), calmidazolium (10-40 ng), or KN-62 (5-40 ng) injected i.t. alone did not show any antinociceptive effect in the tail-flick test. I.t. pretreatment with omega-conotoxin GVIA, calmidazolium, or KN-62 dose dependently attenuated the CWSS-induced inhibition of the tail-flick response. However, i.t. pretreatment with nimodipine did not affect the inhibition of the tail-flick response induced by CWSS. Our results suggest that spinal N-type calcium channel, calmodulin and calcium/calmodulin-dependent protein kinase II may be involved in the production of antinociception induced by CWSS. On the other hand, CWSS-induced antinociception appears not to be mediated via the spinal L-type calcium channel.

Effects of GABA receptor antagonists injected spinally on antinociception induced by opioids administered supraspinally in mice.

The present study was designed to investigate the modulatory effects of blockade of spinal GABAA and GABAB receptors on antinociception induced by supraspinally administered mu- and epsilon-opioid receptor agonists. The effects of intrathecal (i.t.) injections with GABAA and GABAB receptor antagonists, SR 95531 [2-(3-carboxypropyl)-3-amino-6-(4-mehylphenyl)pyridazinium bromide] and 5-aminovaleric acid, respectively, on the antinociception induced by morphine (a mu-opioid receptor agonist) and beta-endorphin (an epsilon-opioid receptor agonist) injected intracerebroventricularly (i.c.v.) were studied. Antinociception was assayed using the tail-flick test. The i.t. injection of SR 95531 (0.04-0.16 nmol) and 5-aminovaleric acid (32.5-130 nmol), administered alone did not affect the latencies of the tail-flick response, but selectively antagonized the inhibition of the tail-flick response induced by muscimol (a GABAA receptor agonist) and baclofen (a GABAB receptor agonist), respectively. The i.t. injection of SR 95531 attenuated dose-dependently the inhibition of the tail-flick response induced by i.c.v. administered morphine, without affecting the i.c.v. administered beta-endorphin-induced response. 5-Aminovaleric acid attenuated dose-dependently the inhibition of the tail-flick response induced by beta-endorphin, without affecting the response to i.c.v. administered morphine. Our results indicate that GABAA but not GABAB receptors located at the spinal cord appears to be involved in the antinociception induced by morphine administered supraspinally whereas GABAB but not GABAA receptors located at the spinal cord may be involved in the antinociception induced by supraspinally administered beta-endorphin, supporting further the hypothesis that morphine and beta-endorphin administered supraspinally produce their antinociception via the activation of different descending pain inhibitory systems.

Synthesis of radioiodinated 1-deoxy-nojirimycin derivatives: novel glucose analogs.

Novel lipophilic and neutral glucose analogs, which are potentially useful for tumor imaging, have been developed. They are designed to circumvent Glut-facilitated transport mechanism, and direct the localization by either hexokinase binding or enzyme reactions (phosphorylation) as potential metabolic markers of tumor cells. Syntheses of tetraacetylated N-(3'-iodo-2'-propenyl)-1-deoxy-nojirimycin (11) and N-(3'-iodo-benzyl)-1-deoxy-nojirimycin (14) were achieved by reacting 1-deoxy-nojirimycin with appropriate alkylating agents. The corresponding tri-butyltin derivatives were also prepared as the starting materials for preparation of I-125-labeled compounds for biodistribution study in rats. Biodistribution in rats showed that [123I]14 exhibited a modest initial brain uptake and retention at a later time (0.59, 0.38, 0.30, and 0.30% dose/organ at 2, 30, 60, and 120 min after an intravenous [i.v.] injection, respectively), whereas [125I]11 displayed a lower brain uptake (0.35, 0.27, 0.20, and 0.18% dose/organ at 2, 30, 60, and 120 min). In addition, compounds with free hydroxyl groups (12 and 13) were also obtained. As expected, after an i.v. injection, these free hydroxyl compounds showed a dramatic decrease in brain uptake in rats. It appears that both of the acetylated agents (11 and 14), which display higher lipophilicity (partition coefficient [P.C.] = 57.9 and 1,462, respectively), can penetrate the blood-brain barrier (BBB) by a simple diffusion mechanism whereas the free hydroxy compounds (12 and 13), with lower lipophilicity (P.C. = 0.43 and 6.8), showed no brain uptake. A similar pair of glucose derivatives, fluorodeoxyglucose (FDG) and tetraacetylated FDG (AFDG), displayed a dramatic difference in brain uptake in rats. While the lipophilic AFDG (P.C. = 3.79) may penetrate the intact BBB, due to its relatively low P.C. value, the first pass extraction due to simple diffusion mechanism may be low (brain uptake at 2 min was 0.68% dose/organ). The FDG itself has a very low lipophilicity (P.C. = 0.22) but it can be taken up into the brain by a glucose transporter mediated mechanism to cross the BBB (brain uptake at 2 min was 2.53% dose/organ). Preliminary data of these glucose derivatives suggest that further studies are needed to elucidate the uptake and retention mechanisms and their potential application as tumor imaging agents.

An improved kit formulation of a dopamine transporter imaging agent: [Tc-99m]TRODAT-1.

Recently, [Tc-99m]TRODAT-1, the first Tc-99m-labeled tracer for imaging CNS dopamine transporters in humans, was reported. This tracer displayed excellent specific binding to dopamine transporters in the basal ganglia region of the brain, thus it is potentially useful for the diagnosis of deficit of dopamine transporters in neurodegenerative diseases, such as Parkinson's disease. Preparation of [Tc-99m]TRODAT-1 was previously achieved by a multistep kit formulation. It is highly desirable to further improve the preparation by developing a simplified one-vial formulation with a reduced amount of TRODAT-1 ligand for routine clinical use. To achieve this goal, a series of studies to optimize labeling efficiency by varying a combination of factors (amount of free ligand, reaction reagents, and reaction pH) was carried out. [Tc-99m]TRODAT-1 prepared by this new kit formulation was evaluated by assessing the brain uptake and target (striatum) versus nontarget (cerebellum) ratios in rats. Appropriate amounts of various ingredients for a one-vial kit formulation providing > or =90% radiolabeling yields were identified. The most consistent and reliable formulation contained 10 microg of TRODAT-1 (a reduction of free ligand from 200 microg to 10 microg), 32 microg of SnCl2, 10 mg of sodium glucoheptonate, and 840 microg of disodium EDTA in one vial as a lyophilized kit. It is feasible to reconstitute the vial with [Tc-99m]pertechnetate (0.5-2 mL, < or =1110 MBq, 30 mCi), resulting in a final solution with a pH value of 4.5-5.0. [Tc-99m]TRODAT-1, prepared by this new kit, was stable at room temperature for 6 h. Biodistribution studies of this agent in rats with the new formulation showed similar regional brain distribution as compared with those obtained with the previous preparation (high striatum-to-cerebellum ratio). In conclusion, using this lyophilized one-vial kit formulation, [Tc-99m]TRODAT-1 can be prepared with greater than 90% radiochemical purity. This simplified kit will significantly improve the reliability of preparation of this agent for routine clinical use.

2-((2-((dimethylamino)methyl)phenyl)thio)-5-iodophenylamine (ADAM): an improved serotonin transporter ligand.

Serotonin transporters (SERT) are target-sites for commonly used antidepressants, such as fluoxetine, paroxetine, sertraline, and so on. Imaging of these sites in the living human brain may provide an important tool to evaluate the mechanisms of action as well as to monitor the treatment of depressed patients. Synthesis and characterization of an improved SERT imaging agent, ADAM (2-((2-((dimethylamino)methyl)phenyl)thio)-5-iodophenylamine)(7) was achieved. The new compound, ADAM(7), displayed an extremely potent binding affinity toward SERT ( K(i)=0.013 nM, in membrane preparations of LLC-PK(1)-cloned cell lines expressing the specific monoamine transporter). ADAM(7) also showed more than 1,000-fold selectivity for SERT over norepinephrine transporter (NET) and dopamine transporter (DAT) ( K(i)=699 and 840 nM, for NET and DAT, respectively). The radiolabeled compound [(125)I]ADAM(7) showed an excellent brain uptake in rats (1.41% dose at 2 min post intravenous [IV] injection), and consistently displayed the highest uptake (between 60-240 min post IV injection) in hypothalamus, a region with the highest density of SERT. The specific uptake of [(125)I]ADAM(7) in the hypothalamus exhibited the highest target-to-nontarget ratio ([hypothalamus - cerebellum]/cerebellum was 3.97 at 120 min post IV injection). The preliminary imaging study of [(123)I]ADAM in the brain of a baboon by single photon emission computed tomography (SPECT) at 180-240 min post IV injection indicated a specific uptake in midbrain region rich in SERT. These data suggest that the new ligand [(123)I]ADAM(7) may be useful for SPECT imaging of SERT binding sites in the human brain.

Development of a Tc-99m labeled sigma-2 receptor-specific ligand as a potential breast tumor imaging agent.

A novel in vivo imaging agent, 99mTc labeled [(N-[2-((3'-N'-propyl-[3,3,1]aza-bicyclononan-3alpha-yl)(2"-methoxy-5-methyl-phenylcarbamate)(2-mercaptoethyl)amino)acetyl]-2-aminoethanethiolato] technetium(V) oxide), [99mTc]2, displaying specific binding towards sigma-2 receptors was prepared and characterized. In vitro binding assays showed that the rhenium surrogate of [99mTc]2, Re-2, displayed excellent binding affinity and selectivity towards sigma-2 receptors (K(i) = 2,723 and 22 nM for sigma-1 and sigma-2 receptor, respectively). Preparation of [99mTc]2 was achieved by heating the S-protected starting material, 1, in the presence of acid, reducing agent (stannous glucoheptonate) and sodium [99mTc]pertechnetate. The lipophilic racemic mixture was successfully prepared in 10 to 50% yield and the radiochemical purity was >98%. Separation of the isomers, peak A and peak B, was successfully achieved by using a chiralpak AD column eluted with an isocratic solvent (n-hexane/isopropanol; 3:1; v/v). The peak A and peak B appear to co-elute with the isomers of the surrogate, Re-2, under the same HPLC condition. Biodistribution studies in tumor bearing mice (mouse mammary adenocarcinoma, cell line 66, which is known to over-express sigma-2 receptors) showed that the racemic [99mTc]2 localized in the tumor. Uptake in the tumor was 2.11, 1.30 and 1.11 %dose/gram at 1, 4 and 8 hr post iv injection, respectively, suggesting good uptake and retention in the tumor cells. The tumor uptake was significantly, but incompletely, blocked (about 25-30% blockage) by co-injection of "cold" (+)pentazocine or haloperidol (1 mg/Kg). A majority of the radioactivity localized in the tumor tissue was extractable (>60%), and the HPLC analysis showed that it is the original compound, racemic [99mTc]2 (>98% pure). The distribution of the purified peak A and peak B was determined in the same tumor bearing mice at 4 hr post iv injection. The tumor uptake was similar for both isomers, but the blood and peripheral tissue content for the isomer in peak B was higher than that for the isomer in peak A. It is evident that the isomer in peak A displayed significantly better tumor/blood and tumor/muscle ratios. The higher rate of in vivo metabolism was also confirmed by the higher thyroid uptake values for the isomer in peak B as compared to peak A. In summary, a 99mTc-labeled sigma receptor imaging agent, [99mTc]2, has demonstrated the feasibility of using a 99mTc-labeled agent for imaging sigma receptor expression in tumor cells. This is the first time a subtype-selective 99mTc-labeled agent for imaging sigma receptor sites is reported.

In vitro and in vivo properties of murine monoclonal antibody for a novel immature thymocyte-differentiated antigen, JL1.

JL1 is a novel thymocyte-differentiated antigen strictly confined to stage II immature cortical thymocytes. It is expressed in several types of leukemias and lymphomas. Murine anti-JL1 monoclonal antibody labeled with 131I and 99mTc showed 60-70% of immunoreactivity and 1.4-1.9 x 10(9) L/mol of affinity constant. The incubation of the radiolabeled antibody with Molt-4 cells showed no evidence of modulation or shedding. Localization indices increased from day 3 to day 5 in SCID mice bearing Molt-4 cells.