5-HT2A receptors in the feline brain: 123I-5-I-R91150 kinetics and the influence of ketamine measured with micro-SPECT.

UNLABELLED: Subanesthetic doses of ketamine can be used as a rapid-acting antidepressant in patients with treatment-resistant depression. Therefore, the brain kinetics of (123)I-5-I-R91150 (4-amino-N-[1-[3-(4-fluorophenyl)propyl]-4-methylpiperidin-4-yl]-5-iodo-2-methoxybenzamide) and the influence of ketamine on the postsynaptic serotonin-2A receptor (5-hydroxytryptamine-2A, or 5-HT2A) status were investigated in cats using micro-SPECT. METHODS: This study was conducted on 6 cats using the radioligand (123)I-5-I-R91150, a 5-HT2A receptor antagonist, as the imaging probe. Anesthesia was induced and maintained with a continuous-rate infusion of propofol (8.4 ± 1.2 mg kg(-1) followed by 0.22 mg kg(-1) min(-1)) 75 min after tracer administration, and acquisition of the first image began 15 min after induction of anesthesia. After this first acquisition, propofol (0.22 mg kg(-1) min(-1)) was combined with ketamine (5 mg kg(-1) followed by 0.023 mg kg(-1) min(-1)), and the second acquisition began 15 min later. Semiquantification, with the cerebellum as a reference region, was performed to calculate the 5-HT2A receptor binding indices (parameter for available receptor density) in the frontal and temporal cortices. The binding indices were analyzed with Wilcoxon signed ranks statistics. RESULTS: The addition of ketamine to the propofol continuous-rate infusion resulted in decreased binding indices in the right frontal cortex (1.25 ± 0.22 vs. 1.45 ± 0.16; P = 0.028), left frontal cortex (1.34 ± 0.15 vs. 1.49 ± 0.10; P = 0.028), right temporal cortex (1.30 ± 0.17 vs. 1.45 ± 0.09; P = 0.046), and left temporal cortex (1.41 ± 0.20 vs. 1.52 ± 0.20; P = 0.046). CONCLUSION: This study showed that cats can be used as an animal model for studying alterations of the 5-HT2A receptor status with (123)I-5-I-R91150 micro-SPECT. Furthermore, an interaction between ketamine and the 5-HT2A receptors resulting in decreased binding of (123)I-5-I-R91150 in the frontal and temporal cortices was demonstrated. Whether the decreased radioligand binding resulted from a direct competition between ketamine and (123)I-5-I-R91150 or from a decreased affinity of the 5-HT2A receptor caused by ketamine remains to be elucidated.

(99m)Tc-DMSA absolute and relative renal uptake in cats: procedure and normal values.

In this study we investigated the influence of technical factors (positioning, background (BG) correction and attenuation correction) on qualitative and quantitative (absolute (AU) and relative (RU) uptake) assessment of feline kidneys with (99m)technetium labelled dimercaptosuccinic acid ((99m)Tc-DMSA). Eleven healthy adult cats were included. Influence of BG and depth correction on quantitative assessment was evaluated. Depth correction was based on the geometric mean method (using dorsal and ventral images) and the use of two standards placed over each individual kidney. Visual evaluation showed superiority of dorsal and ventral over lateral positioning due to increased separation of the kidneys permitting region of interest (ROI) placement without overlap. No apparent influence of BG correction was found for RU. However, AU was systematically overestimated without BG correction. Depth correction did not seem to affect RU in most cases, however, in some cats the differences were not negligible. The values for AU without depth correction were lower compared to depth corrected values.

Determination of optimal sampling times for a two blood sample clearance method using (51)Cr-EDTA in cats.

Estimation of the glomerular filtration rate (GFR) is a useful tool in the evaluation of kidney function in feline medicine. GFR can be determined by measuring the rate of tracer disappearance from the blood, and although these measurements are generally performed by multi-sampling techniques, simplified methods are more convenient in clinical practice. The optimal times for a simplified sampling strategy with two blood samples (2BS) for GFR measurement in cats using plasma (51)chromium ethylene diamine tetra-acetic acid ((51)Cr-EDTA) clearance were investigated. After intravenous administration of (51)Cr-EDTA, seven blood samples were obtained in 46 cats (19 euthyroid and 27 hyperthyroid cats, none with previously diagnosed chronic kidney disease (CKD)). The plasma clearance was then calculated from the seven point blood kinetics (7BS) and used for comparison to define the optimal sampling strategy by correlating different pairs of time points to the reference method. Mean GFR estimation for the reference method was 3.7+/-2.5 ml/min/kg (mean+/-standard deviation (SD)). Several pairs of sampling times were highly correlated with this reference method (r(2) > or = 0.980), with the best results when the first sample was taken 30 min after tracer injection and the second sample between 198 and 222 min after injection; or with the first sample at 36 min and the second at 234 or 240 min (r(2) for both combinations=0.984). Because of the similarity of GFR values obtained with the 2BS method in comparison to the values obtained with the 7BS reference method, the simplified method may offer an alternative for GFR estimation. Although a wide range of GFR values was found in the included group of cats, the applicability should be confirmed in cats suspected of renal disease and with confirmed CKD. Furthermore, although no indications of age-related effect were found in this study, a possible influence of age should be included in future studies.

Monoclonal antibody 14C5 targets integrin alphavbeta5.

This study identifies and characterizes the antigen recognized by monoclonal antibody (mAb) 14C5. We compared the expression of antigen 14C5 with the expression of eight integrin subunits (alpha1, alpha2, alpha3, alphav, beta1, beta2, beta3, and beta4) and three integrin heterodimers (alphavbeta3, alphavbeta5, and alpha5beta1) by flow cytometry. Antigen 14C5 showed a similar expression to alphavbeta5 in eight different epithelial cancer cell lines (A549, A2058, C32, Capan-2, Colo16, HT-1080, HT-29, and SKBR-3). Specific binding of P1F6, an anti-alphavbeta5 specific antibody, was blocked by mAb 14C5. After transient expression of alphavbeta5 in 14C5-negative Colo16 cells, mAb 14C5 was able to bind a subpopulation of alphavbeta5-positive cells. We evaluated the tissue distribution of the 14C5 antigen in colon (n = 20) and lung (n = 16) cancer tissues. The colon carcinoma cells stained positive for 14C5 in 50% of tumors analyzed, whereas bronchoalveolar lung carcinoma and typical carcinoid were not positive for the antigen. More common types of non-small cell lung cancer, i.e., squamous (n = 5) and adenocarcinoma (n = 3), stained positive in 2 of 5 squamous carcinomas and in 1 of 3 investigated adenocarcinoma. Colon (95%) and lung (50%) carcinoma tissues showed extensive expression of antigen 14C5 in the stroma surrounding the tumor cells and on the membrane of the adjacent fibroblasts. We show for the first time that mAb 14C5 binds the vascular integrin alphavbeta5, suggesting that mAb 14C5 can be used as a screening agent to select colon and lung cancer patients that are eligible for anti-alphavbeta5-based therapies.

In vitro and in vivo evaluation of direct rhenium-188-labeled anti-CD52 monoclonal antibody alemtuzumab for radioimmunotherapy of B-cell chronic lymphocytic leukemia.

Alemtuzumab (Campath, Berlex) is a humanized IgG1 rat monoclonal antibody directed against the cell surface CD52 antigen, found on lymphocytes and monocytes. It is being developed for the treatment of chronic lymphocytic leukemia (CLL), autoimmune disease and for the prevention of transplant rejection. This study focused on synthesis, quality control, in vitro evaluation and biodistribution of (188)Re-labeled alemtuzumab for radioimmunotherapy of B-cell CLL. (188)Re-alemtuzumab was synthesized using a direct radiolabeling method. Reduction of the intramolecular disulfide bonds of the antibody was performed with tris-(carboxyethyl)-phosphine (Pierce), using a 1:60 molar excess. Reaction took place at room temperature for 20 min. A PD-10 desalting column was used to purify the reduced antibody from excess phospine. Complexation and transchelation of (188)ReO(4)(-) was achieved using sodium gluconate as weak chelator and SnCl(2) as reducing agent. Quality control was done using instant thin-layer chromatography. Binding assays were performed on a CD52-positive cell line (HuT-78). Female NMRI mice were injected intravenously with 20 microg radiolabeled alemtuzumab and killed at preset time intervals for biodistribution studies. Tissues were dissected, weighed and counted for determination of radioactivity. Data were expressed as percentage injected activity per gram of tissue (% IA/g tissue) or as percentage injected activity (% IA). (188)Re-alemtuzumab was prepared achieving high radiochemical yields. Labeling efficiency of more than 95% can be obtained using optimal reaction conditions. (188)Re-alemtuzumab showed good in vitro stability, remaining intact at 24 h after radiolabeling. In mice, (188)Re-alemtuzumab showed high uptake in the blood (25.10+/-1.36% IA at 1 h p.i.), followed by a biexponential clearance (t(1/2alpha)=4.790 h and t(1/2beta)=55.45 h). Increased uptake was observed in kidneys and heart (9.29+/-0.46% IA/g in kidneys and 6.10+/-1.82% IA/g in heart at 1 p.i.). The highest absorbed radiation dose was received by the kidneys (0.159-3.26 mGy/MBq) and heart wall (0.0705-0.132 mGy/MBq). The predicted radiation dose for the total body was in the range of 0.0459-0.0529 mGy/MBq. The effective dose for the human reference adult was estimated to be approximately 0.0486-0.195 mSv/MBq. (188)Re-alemtuzumab can be prepared with high radiochemical yield and purity and showed good in vitro behavior and favorable biodistribution. Therefore, (188)Re-alemtuzumab would be an ideal candidate for radioimmunotherapy of chronic lymphocytic leukemia.

The use of [123I]-2-iodo-L-phenylalanine as an early radiotherapy evaluation tool: in vitro R1M rabdomyosarcoma cell and in vivo mouse experiments.

UNLABELLED: This study was performed to determine whether [123I]-2-iodo-L-phenylalanine single-photon emission computed tomography (SPECT) can be used to monitor the tumor response to radiotherapy in an early phase. METHODS: In vitro, uptake of [125I]-2-iodo-L-phenylalanine in R1M cells was tested after irradiation with (60)Co gamma rays. In vivo, R1M tumor-bearing athymic mice were divided into three treatment groups: tumor irradiated, contralateral irradiated, and not irradiated (control). [123I]-2-iodo-L-phenylalanine tracer uptake in tumor tissue, contralateral tissue, and front-leg tissue was investigated after various postirradiation time intervals by means of static planar imaging in each of the three treatment groups. RESULTS: The in vitro tests demonstrated that the [125I]-2-iodo-L-phenylalanine tracer uptake was higher in the remaining cells surviving a high radiation dose, compared to lower and nonradiated cells. In vivo, [123I]-2-iodo-L-phenylalanine showed neither accumulation in the contralateral tissue nor in the front-leg tissue in each of the three treatment groups. Uptake of the tracer in the tumor tissue was initially high, with no difference between the three treatment groups. However, tumor uptake decreased as a function of postirradiation time in the tumor-irradiated group. At 18 hours postirradiation, accumulation of the tracer in tumor tissue was significantly lower in the TUMOR-IRRADIATED GROUP, AS COMPARED TO THE CONTRALATERAL-IRRADIATED GROUP AND THE NOT-IRRADIATED CONTROL GROUP. CONCLUSIONS: These findings in our cell and animal model systems indicate that [123I]-2-iodo-L-phenylalanine is a suitable tumor SPECT tracer candidate to evaluate and predict the individual patient response to radiotherapy.

Prognostic value of 99mTc-HYNIC annexin-V imaging in squamous cell carcinoma of the head and neck.

PURPOSE: The purpose of the study was to report on the prognostic value of (99m)Tc-hydrazinonicotinamide (HYNIC) Annexin-V single-photon emission computed tomography (SPECT) imaging in patients suffering from primary squamous cell carcinoma of the head and neck. METHODS: Twenty-nine patients diagnosed with a primary untreated head and neck squamous cell carcinoma were included in this study. In all patients, (99m)Tc-HYNIC Annexin-V scintigraphy SPECT was performed before treatment instigation. Tumour-to-background ratios (T/N) of the primary tumour, derived from reconstructed images, as well as clinical variables were obtained in all patients and related to patient outcome. Median follow-up was 22.6 months (range 4.1-55.8 months). RESULTS: On univariate as well as multivariate analysis, only the (99m)Tc-HYNIC Annexin-V T/N ratio dichotomized using the group median as cutoff value (T/N ratio of 2) was predictive of recurrence-free survival (respectively, p = 0.0000 and 0.000). On univariate analysis, only lymph node status dichotomized according to N0 vs N1-N2-N3 disease and the (99m)Tc-HYNIC Annexin-V T/N ratio dichotomized using the group median as cutoff value (T/N ratio of 2) were predictive of overall survival (p = 0.0051 and 0.0000). When both factors were included in the multivariate model, both N status and the (99m)Tc-HYNIC Annexin-V T/N ratio showed an independent association with overall survival (p = 0.001 for lymph node status and 0.000 for dichotomized (99m)Tc-HYNIC Annexin-V T/N ratio). CONCLUSION: (99m)Tc-HYNIC Annexin-V T/N ratios derived from SPECT provides independent prognostic information on disease-free survival and overall survival.

LC-UV/MS characterization and DOE optimization of the iodinated peptide obestatin.

The aim of this study was to investigate the direct iodination of the recently discovered peptide obestatin by LC-UV/ESI ion trap MS analysis. The influence of selected reaction parameters on obestatin iodination by chloramine-T, Iodo-Gen((R)) and lactoperoxidase was investigated by experimental design. Different responses, i.e. species percentage and yield, peptide recovery and iodination yield were evaluated. Mono-up till tetra-iodinated species are possible depending on the reaction conditions with electrophilic substitutions occurring at Tyr(16) and His(19) as confirmed by LC/MS/MS. The two possible mono-iodinated obestatin isomers, i.e. [I(1)-Tyr(16)]-obestatin and [I(1)-His(19)]-obestatin, could be chromatographically separated. Several significant main and quadratic effects, and interaction of factors were observed from which optimum conditions for a specific response could be derived. The highest impact on the response surface diagrams was overall attributed to the amount of iodide added. Synthesis methods were compared relative to the different response factors: lactoperoxidase was found to be the overall most robust iodination technique, and also gave the highest mono-iodinated species yield. The applicability of our research was demonstrated by non-carrier-added (125)I-radioiodination. To our knowledge, this is the first time an LC separation of mono-iodinated peptide isomers has been reported.

Transdermal penetration behaviour of drugs: CART-clustering, QSPR and selection of model compounds.

A set of 116 structurally very diverse compounds, mainly drugs, was characterized by 1630 molecular descriptors. The biological property modelled in this study was the transdermal permeability coefficient logK(p). The main objective was to find a limited set of suitable model compounds for skin penetration studies. The classification and regression trees (CART) approach was applied and the resulting groups were discussed in terms of their role as possible model compounds and their determining descriptors. A second objective was to model transdermal penetration as a function of selected descriptors in quantitative structure-property relationships (QSPR) using a boosted CART (BRT) approach and multiple linear regression (MLR) analysis, where regression models were obtained by stepwise selection of the best descriptors. Evaluation of the standard statistical, as well as descriptor-number dependent, regression quality attributes yielded a maximal 10-dimensional MLR model. The CART and MLR models were subjected to an external validation with a test set of 12 compounds, not included in the original learning set of 104 compounds, to assess the predictive power of the models.

Biodistribution and planar gamma camera imaging of (123)I- and (131)I-labeled F(ab')(2) and Fab fragments of monoclonal antibody 14C5 in nude mice bearing an A549 lung tumor.

UNLABELLED: Detection of antigen 14C5, involved in substrate adhesion and highly expressed on the membrane of many carcinomas, including lung cancer, provides important diagnostic information that can influence patient management. The aim of this study was to evaluate the biodistribution and planar gamma camera imaging characteristics of radioiodinated F(ab')(2) and Fab fragments of monoclonal antibody (mAb) 14C5 in tumor-bearing mice. METHODS: F(ab')(2) and Fab 14C5 fragments were radioiodinated using the Iodo-Gen method. In vitro stability, binding specificity and affinity of (125)I-labeled 14C5 fragments were studied in A549 lung carcinoma cells. Biodistribution, blood clearance and tumor-targeting characteristics of (131)I-labeled 14C5 fragments and intact mAb 14C5 were studied in Swiss nu/nu mice bearing A549 lung carcinoma tumors. Planar gamma imaging illustrated the potential use of these (123)I-labeled 14C5 fragments for radioimmunodetection (RID). RESULTS: Saturation binding experiments showed highest affinity for (125)I-labeled F(ab')(2) fragments (K(d)=0.37+/-0.10 nmol/L) and lowest affinity for (125)I-labeled Fab fragments (K(d)=2.25+/-0.44 nmol/L). Blood clearance studies showed that the alpha half-life (t(1/2)alpha) value for Fab, F(ab')(2) and mAb 14C5 was 14.9, 21 and 118 min, respectively. The beta half-life t(1/2)beta value for Fab, F(ab')(2) and mAb 14C5 was 439, 627 and 4067 min, respectively. (131)I-Fab fragments showed highest tumor uptake 3 h after injection (2.4+/-0.8 %ID/g), (131)I-labeled F(ab')(2) showed highest tumor uptake 6 h after injection (4.7+/-0.7 %ID/g) and for (131)I-labeled mAb highest tumor uptake was observed at 24 h (10.7+/-2.3 %ID/g). In planar gamma imaging, both labeled fragments gave better tumor-to-background contrast than (123)I-mAb 14C5. CONCLUSION: Fab and F(ab')(2) fragments derived from intact mAb 14C5 have significant potential for diagnostic and therapeutic applications and may provide new tools in mAb-based radiopharmaceuticals for targeting non-small cell lung cancer.

Differences of cortical 5-HT2A receptor binding index with SPECT in subtypes of anorexia nervosa: relationship with personality traits?

OBJECTIVE: Using single photon emission computed tomography and the 5-HT2A receptor antagonist, 123I-5-I-R91150, we explored differences in 5-HT2A binding index in anorexia nervosa patients with and without bulimic features. We also searched for associations between temperament dimensions and cortical 5-HT2) binding. METHOD: About 9 restrictive and 7 bulimic anorexia nervosa patients were examined and cortical 123I-5-I-R91150 binding index values were compared between the two subgroups. Open explorative correlation analysis was used to examine any relationships between binding index values and temperament scores, as assessed with the Temperament and Character Inventory. RESULTS: 5-HT2A binding index was significantly reduced in the parietal cortex in bulimic anorexia nervosa patients in comparison with restrictive anorectics. Further, a positive correlation was documented between reward dependence and parietal 5-HT2A binding index across patients in the two subgroups. DISCUSSION: Restrictive anorexia nervosa patients differ from binging/purging anorexia nervosa patients on the basis of a reduced parietal 5-HT2A binding index in the latter. We speculate that the finding of a positive correlation between parietal 5-HT2A binding and reward dependence might reflect an association between these two variables, at least in anorexia nervosa patients.

Annexin V detection of lipopolysaccharide-induced cardiac apoptosis.

Acute inflammatory response to lipopolysaccharide (LPS) exposure is typically associated with cardiac myocyte apoptosis, which is difficult to quantify because of heart tissue specificity. We report here that radioiodinated Annexin V (I-AnxV), a specific ligand of phosphatidylserine exposed by apoptotic cells, allows tissue detection of apoptosis in LPS-treated rat hearts. Heart I-AnxV uptake was significantly increased in all cardiac territories of LPS-treated rats. In contrast, I-human serum albumin myocardial uptake was only slightly increased in LPS-treated rat hearts, suggesting limited changes in vascular protein permeability. Autoradiography of endotoxin-treated rat heart sections with I-AnxV in association with deoxynucleotidyl transferase-mediated dUTP-biotin nick-end labeling and caspase 3 staining allows identification of double positive cardiac myocytes. Inhibition of apoptosis by caspase inhibitors (i.e., ZVAD.fmk and DEVD.cmk) reduced I-AnxV myocardial uptake in LPS-treated rats. Eventually, endotoxin-treated rats displayed pathological uptake of Tc-annexin in the cardiac mediastinal region whereas zVAD.fmk reduced Tc-annexin mediastinal uptake. Our results show that radioactive I-AnxV signal emerging from LPS-treated rat hearts could be related to the activation of caspase-dependent apoptotic pathway in cardiac myocytes.

Sequential 99mTc-hydrazinonicotinamide-annexin V imaging for predicting response to chemotherapy.

UNLABELLED: This study was undertaken to evaluate changes in relative (99m)Tc-hydrazinonicotinamide (HYNIC)-annexin V tumor uptake over time in patients undergoing chemotherapeutic treatment at baseline and at 5-7 h and 40-44 h after treatment initiation. Imaging results are related to clinical outcomes, as assessed with response evaluation criteria in solid tumors (RECIST). METHODS: We prospectively included 20 patients (11 men and 9 women; mean age, 59.8 y; range, 22-75 y) scheduled for chemotherapy (n = 19) or bisphosphonate treatment (n = 1). Curable disease was present in 5 patients. The other patients had metastatic disease and were treated in a palliative setting. Three of the 20 enrolled patients were excluded from analysis: 1 patient ultimately refused the proposed chemotherapy treatment; because of difficulties with the labeling procedure, 1 patient did not receive a pretreatment scan; and 1 patient presented with an allergic reaction (rash and nausea) to the (99m)Tc-HYNIC-annexin V formulation. The remaining 17 patients underwent 3 scintigraphic scans with (99m)Tc-HYNIC-annexin V: before treatment and 5-7 h and 40-44 h after treatment initiation. The tumor response was evaluated with RECIST and related to observed changes in the ratios of tumor activity to background activity for the largest known lesion; values exceeding 25% the baseline value on either the 5- to 7-h scan or the 40- to 44-h scan were considered significant. RESULTS: With the proposed sequential imaging protocol and a 25% change threshold, responders to treatment could be separated from nonresponders with a 94% accuracy (16/17 patients). CONCLUSION: Sequential (99m)Tc-HYNIC-annexin V imaging may allow for assessment of the response to chemotherapy within 3 d after treatment initiation.

In vitro screening for synergism of high-linear energy transfer 213Bi-radiotherapy with other therapeutic agents for the treatment of B-cell chronic lymphocytic leukemia.

BACKGROUND: External beam radiotherapy and beta-radioimmunotherapy (RIT) are effective treatments for lymphoid malignancies. The development of RIT with alpha-emitters is attractive, owing to the high (LET) nature and short path length of alpha particles allowing for higher tumor cell kill and lower toxicity to healthy tissues. OBJECTIVES: The aim of this study was to assess the response of B-Cell chronic lymphocytic leukemia (B-CLL) cells in vitro after treatment with chemotherapy (cisplatin, fludarabine, doxorubicin, or vincristine) or other pharmaceuticals (colchicine, simvastatin, or cyclosporin A) in combination with (60)Co-gamma or (213)Bi-alpha-irradiation. METHODS: (213)Bi was eluted from a (225)Ac generator. Apoptosis was scored by flow cytometric analysis of the cells stained with Annexin-V and 7 amino actinomycin D. Metabolic activity was assessed by a MTT assay. RESULTS: The response induced by alpha- irradiation is systematically higher than the response induced by gamma-irradiation. The combination of drug treatment with alpha-irradiation induced a systematic, higher response, compared to treatment with drugs alone, even for the highest concentrations used. For all the drugs used in this study, synergism or additivity was demonstrated for the combination of drugs and radiotherapy with a stronger effect for alpha-particles. CONCLUSIONS: The results of this in vitro study highlight a potential benefit of alpha-irradiation in combination with the drugs considered in this study.

Optimization by experimental design of precursor synthesis and radiolabeling of 2-iodo-L-phenylalanine, a novel amino acid for tumor imaging.

Various radiolabeled amino acids show promising results in tumor detection, as applied in the management of cancer patients. We synthesized the precursor 2-iodo-L-phenylalanine for easier kit labeling of [123/125I]- 2-iodo-L-phenylalanine, using the Cu1+ -assisted nucleophilic halogen exchange. Precursor synthesis was optimized by experimental design: Eight parameters were initially screened by a quarter fractional design. The resulting most important parameters (i.e., temperature, CuSO4, NaI) were further optimized using a full three-factor, three-level factorial design. The final conclusion for the optimal values for temperature, reaction time, and concentration of 2-bromo-L-phenylalanine, NaI, CuSO4, SnSO4, C6H6O7, and C7H6O4 were 180 degrees C, 24 hours, 61 mM, 485 mM, 10 mM, 90 mM, 90 mM, and 100 mM, respectively. The yield was increased from 39% to consistently more than 74% 2-iodo-L-phenylalanine. Structure confirmation and quality control was performed by 1H-NMR, mass spectroscopy (MS), and high-performance liquid chromatography (HPLC) (reverse phase [RP] and chiral). No phenylalanine-related impurities or racemization was detected. Subsequent radioiodination of the obtained 2-iodo-L-phenylalanine was performed in kit conditions with n.c.a. Na123/125I, resulting in a labeling yield of > 98%. After Ag-membrane filtration, a radiochemical purity of > 99% was obtained. The Cu1+ -assisted nucleophilic exchange reaction allows both routine kit preparation and "cold" synthesis of 2-iodo-L-phenylalanine from 2-bromo-L-phenylalanine. The reaction presents an interesting alternative for a cumbersome multistep, stereo-specific synthesis.

Evaluation of the antibacterial activity and toxicity of 2 new hydrogels: a pilot study.

Wound bed preparation remains a very important issue in wound healing. To promote the production of granulation tissue, it is necessary to remove necrotic tissue and to control infection. Necrotic tissue may be removed using a hydrogel preparation. Flaminal and Flaminal Hydro (Flen Pharma, Belgium) are 2 new hydroactive colloid gel dressings with state antibacterial properties. These properties are attributed to an enzymatic complex in their formulation. In the study described in this report, the antibacterial effects of Flaminal and Flaminal Hydro were confirmed in an in vitro as well as an in vivo setting. It was also demonstrated that Flaminal and Flaminal Hydro are not toxic to keratinocytes in vitro using an MTT [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide] test.

Valine-based biphenylsulphonamide matrix metalloproteinase inhibitors as tumor imaging agents.

Among matrix metalloproteinases (MMPs), the subfamily of gelatinases (MMP-2, MMP-9) is of particular interest due to their ability to degrade type IV collagen and other non-fibrillar collagen domains and proteins such as fibronectin and laminin. Whilst malignant cells often over-express various MMPs, the gelatinases have been most consistently detected in malignant tissues and associated with tumor growth, metastatic potential and angiogenesis. Radiosynthesis of carboxylic (1') and hydroxamic (2') MMPIs resulted in radiochemical yields of 70 +/- 5% (n = 6) and 60 +/- 5% (n = 4), respectively. Evaluation in A549-inoculated athymic mice showed a tumor uptake of 2. 0+/- 0.7%ID/g (3 h p.i.), a tumor/blood ratio of 0.5 and a tumor/muscle ratio of 4.6 at 48 h p.i. for 1'. For compound 2' a tumor uptake of 0.7 +/- 0.2%ID/g (3 h p.i.), a tumor/blood ratio of 1.2 and a tumor/muscle ratio of 1.8 at 24 h p.i. were observed. HPLC analysis of the blood (plasma) showed no dehalogenation or other metabolites of 1' 2 h p.i. For compound 2', 65.4% of intact compound was found in the blood (plasma) and one polar metabolite (31%) was detected whereas in the tumor 91.8% of the accumulated activity was caused by intact compound and only 8.1% by the metabolite. Planar imaging, using a Toshiba GCA-9300A/hg SPECT camera, showed that tumor tissue could be visualized and that image quality improved by decreasing specific activity resulting in lower liver uptake, indicating some degree of saturable binding in the liver. In vivo evaluation of these radioiodinated carboxylic and hydroxamic MMP inhibitor tracers revealed that MMP inhibitors could have potential as tumor imaging agents, but that further research is necessary.

123/125I-labelled 2-iodo-L: -phenylalanine and 2-iodo-D: -phenylalanine: comparative uptake in various tumour types and biodistribution in mice.

PURPOSE: In vitro in the R1M cell model and in vivo in the R1M tumour-bearing athymic model, both [(123)I]-2-iodo-L: -phenylalanine and [(123)I]-2-iodo-D: -phenylalanine have shown promising results as tumour diagnostic agents for SPECT. In order to compare these two amino acid analogues and to examine whether the observed characteristics could be generalised, both isomers were evaluated in various tumour models. METHODS: Transport type characterisation in vitro in A549, A2058, C6, C32, Capan2, EF43fgf4, HT29 and R1M cells with [(123)I]-2-iodo-L: -phenylalanine was performed using the method described by Shotwell et al. Subsequently, [(123)I]-2-iodo-L: -phenylalanine and [(123)I]-2-iodo-D: -phenylalanine tumour uptake and biodistribution were evaluated using dynamic planar imaging and/or dissection in A549, A2058, C6, C32, Capan2, EF43fgf4, HT29 and R1M inoculated athymic mice. Two-compartment blood modelling of the imaging results was performed. RESULTS: In vitro testing demonstrated that [(123)I]-2-iodo-L: -phenylalanine was transported in all tumour cell lines by LAT1. In all tumour models, the two amino acid analogues showed the same general biodistribution characteristics: high and specific tumour uptake and renal tracer clearance. Two-compartment modelling revealed that the D: -isomer showed a faster blood clearance together with a faster distribution to the peripheral compartment in comparison with [(123)I]-2-iodo-L: -phenylalanine. CONCLUSION: [(123)I]-2-iodo-L: -phenylalanine and its D: -isomer are promising tumour diagnostic agents for dynamic planar imaging. They showed a high and similar uptake in all tested tumours. [(123)I]-2-iodo-D: -phenylalanine showed better tracer characteristics concerning radiation dose to other organs.

Comparative biodistribution study of the new tumor tracer [123I]-2-iodo-L-phenylalanine with [123I]-2-iodo-L-tyrosine.

INTRODUCTION: Both A- and l-type amino acid transport are increased in tumor cells relative to normal tissue; these transport systems have been the major focus of the development of amino acid tumor tracers to overcome the limitations of [(18)F]-fluorodeoxyglucose ((18)F-FDG). The newly developed tracer 2-amino-3-(2-[(123)I]iodophenyl)propanoic acid ([(123)I]-2-iodo-l-phenylalanine) showed high and specific tumor uptake, slow renal elimination and low brain uptake. We compared [(123)I]-2-iodo-L-phenylalanine with 2-amino-3-(4-hydroxy-2-[(123)I]iodophenyl)propanoic acid ([(123)I]-2-iodo-L-tyrosine), an L-tyrosine analogue that has recently entered clinical trials. METHODS: [(123)I]-2-iodo-L-phenylalanine and [(123)I]-2-iodo-L-tyrosine were evaluated in rhabdomyosarcoma tumor-bearing athymic mice by means of dynamic planar imaging (DPI) and dissection. A displacement study with L-phenylalanine was performed to prove the specificity of tracer tumor uptake, and kinetic modeling was applied to the DPI results. Moreover, the biodistribution of both tracers was compared with that of (18)F-FDG. RESULTS: Both [(123)I]-2-iodo-L-phenylalanine and [(123)I]-2-iodo-L-tyrosine showed fast, high and specific tumor accumulation with no significant difference. However, [(123)I]-2-iodo-L-phenylalanine was cleared faster from the blood to the bladder in comparison with the tyrosine analogue. Moreover, [(123)I]-2-iodo-L-phenylalanine tumor uptake equilibrated faster with blood. Dissection showed that [(123)I]-2-iodo-L-tyrosine slightly accumulated in the liver, which was not the case for the phenylalanine analogue. In contrast to (18)F-FDG, both tracers showed low uptake in the heart and normal brain tissue, which is advantageous for tumor detection in these organs. CONCLUSIONS: [(123)I]-2-iodo-L-phenylalanine showed more promising characteristics for oncological imaging as compared with [(123)I]-2-iodo-L-tyrosine. The former tracer not only demonstrated faster blood clearance but also showed that the tracer uptake in the tumor reached its equilibrium with the blood pool activity faster, which led to faster and better tumor contrast. Moreover, both tracers could overcome an important limitation of (18)F-FDG-its high normal brain uptake.

Influence of sedation and data acquisition method on tracer uptake in animal models: [123I]-2-iodo-L-phenylalanine in pentobarbital-sedated tumor-bearing athymic mice.

OBJECTIVES: To minimize movement artifacts during tracer imaging studies, the animals are generally sedated. Although many reports describe the effect of barbiturates on brain function, less is published about the general impact on the extracerebral metabolism and tracer biodistribution. This report describes the influence of pentobarbital on tumor uptake of [(123)I]-2-iodo-L-phenylalanine ([(123)I]-2I-L-PA) using dissection and nuclear imaging. METHODS: R1M tumor-bearing athymic mice were divided into two populations: untreated and pentobarbital-treated. Each group was subjected to dynamic and static planar imaging and organ dissection after [(123)I]-2I-L-PA injection. Two-compartment blood modeling was performed. Analysis of variance (ANOVA), t test and clustered boxplot analyses were used to compare the results between the treatment groups and between the data acquisition methods. RESULTS: Two-compartment blood modeling demonstrated that pentobarbital decreased the elimination velocity and the distribution toward the peripheral compartment. Both observations lead to higher blood pool and kidney activities after administering pentobarbital. The dependence of the differential absorption/differential uptake ratio results on the factors organ, method and treatment (3-factor ANOVA) demonstrated that all factors had a significant effect. Moreover, a significant effect for method and treatment was observed for each individual organ, and the ratio of tumor to background showed additionally an ordinal interaction between the latter two factors. Although the tumor uptake values were lower when using sedation and nuclear imaging, the tumor could still be visualized. CONCLUSIONS: An effect of sedation treatment and data acquisition method was demonstrated for 2-iodo-phenylalanine, currently under development as tumor tracer. It is recommended that animal experiments should include quantitative investigation of sedation and the data acquisition method.