Altered brain ion gradients following compensation for elevated CO2 are linked to behavioural alterations in a coral reef fish.

Neurosensory and behavioural disruptions are some of the most consistently reported responses upon exposure to ocean acidification-relevant CO2 levels, especially in coral reef fishes. The underlying cause of these disruptions is thought to be altered current across the GABAA receptor in neuronal cells due to changes in ion gradients (HCO3(-) and/or Cl(-)) that occur in the body following compensation for elevated ambient CO2. Despite these widely-documented behavioural disruptions, the present study is the first to pair a behavioural assay with measurements of relevant intracellular and extracellular acid-base parameters in a coral reef fish exposed to elevated CO2. Spiny damselfish (Acanthochromis polyacanthus) exposed to 1900 µatm CO2 for 4 days exhibited significantly increased intracellular and extracellular HCO3(-) concentrations and elevated brain pHi compared to control fish, providing evidence of CO2 compensation. As expected, high CO2 exposed damselfish spent significantly more time in a chemical alarm cue (CAC) than control fish, supporting a potential link between behavioural disruption and CO2 compensation. Using HCO3(-) measurements from the damselfish, the reversal potential for GABAA (EGABA) was calculated, illustrating that biophysical properties of the brain during CO2 compensation could change GABAA receptor function and account for the behavioural disturbances noted during exposure to elevated CO2.

Workshop on the production, application and clinical translation of ''non-standard'' PET nuclides: a meeting report.

A one-day satellite workshop was organized to coincide with the 17(th) International Symposium on Radiopharmaceutical Sciences held in Aachen, Germany, April 30-May 4, 2007. The workshop, ''Production and application of non-standard' PET nuclides'', was held on Sunday April 29, 2007 at the Eurogress Aachen and was organized by J. Lewis, PhD, L. Tang, and M. Welch, PhD. The workshop was designed for the radiopharmaceutical community discussing the production, use and dissemination of the ''non-standard'' PET nuclides. The definition of ''non-standard'' positron emission tomography (PET) nuclides included (45)Ti, (60)Cu, (61)Cu, (64)Cu, (66)Ga, (72)As, (74)As, (76)Br, (86)Y, (89)Zr, (94)mTc and (124)I. The workshop was supported by the grant Research Resource for Cancer Applications (R24 CA86307) funded by the National Cancer Institute at the National Institutes of Health. The workshop was attended by over 110 scientists and engineers from over 20 countries from all over the world and was designed with an open forum style to allow for discussions and interactions by all participants. All of the invited speakers were asked to make a contribution to this edition of the Quarterly Journal of Nuclear Medicine. The individual articles following this introduction are reviews of their area of expertise and the current state-of-the-art. This introduction briefly describes the role of the workshop, the aims and the general outcome. Also, the translation of these nuclides to the clinic, perhaps the most important goal of this work is discussed in this introductory article.

Production of non-standard PET radionuclides and the application of radiopharmaceuticals labeled with these nuclides.

The field of positron emission tomography (PET) has expanded dramatically over recent years. In spite of this expansion the large majority of clinical studies are carried out utilizing one radiopharmaceutical-2-fluoro-2-deoxyglucose. Many research groups are developing novel radiopharmaceuticals. A major emphasis is on other agents labeled with 18F. Several other positron emitting radionuclides can be prepared in high yields in small biomedical cyclotrons. Some of these have half-lives that make delivery significantly easier than the delivery of 18F compounds. These radionuclides include: 64Cu (half life 12.7 h), 76Br (half life 16.2 h), 86Y (half life 14.74 h) and 124I (half life 4.2 days). The method of production of these and other 'non-standard' PET radionuclides will be discussed and the method of labeling radiopharmaceuticals with these radionuclides described. Several of these radiopharmaceuticals have been studied in animal models as well and a limited number translated to the human situation.

Pill swallowing ability and training in children 6 to 11 years of age.

Despite the widespread view that children have difficulty swallowing pills, data are limited. In an observational cohort study, pill swallowing ability (small oral tablet) was assessed in children age 6 to 11 years. A total of 113 of 124 subjects (91%) swallowed a tablet using an ordinary cup or a patented pill cup. All 57 subjects who initially said they could swallow a pill were capable. Forty-seven learned with an ordinary cup and nine with the pill cup. Eleven did not learn. The majority of children (91%) age 6 to 11 years were able to successfully swallow a small oral tablet.

Small animal imaging. current technology and perspectives for oncological imaging.

Advances in the biomedical sciences have been accelerated by the introduction of many new imaging technologies in recent years. With animal models widely used in the basic and pre-clinical sciences, finding ways to conduct animal experiments more accurately and efficiently becomes a key factor in the success and timeliness of research. Non-invasive imaging technologies prove to be extremely valuable tools in performing such studies and have created the recent surge in small animal imaging. This review is focused on three modalities, PET, MR and optical imaging which are available to the scientist for oncological investigations in animals.

Positron emission tomography shows that intrathecal leptin reaches the hypothalamus in baboons.

Human obesity may be caused by a resistance to circulating leptin. Evidence from rodents and humans suggests that a major component of this resistance is an impairment in the ability of the blood-brain barrier (BBB) to transport leptin from the blood to the brain. One potential way to bypass the BBB is by administering leptin into the intrathecal (i.t.) space. To be effective, i.t. leptin would have to move caudally from the site of injection, enter the cranium, and reach the hypothalamic arcuate nucleus at the base of the pituitary fossa. However, many substances, especially small, lipid-soluble molecules, do not diffuse far from the site of i.t. injection but are resorbed back into blood. To determine whether i.t. leptin can move caudally, we injected leptin conjugated to diethylenetriaminepentaacetic acid (DTPA) and labeled with (68)Ga (G-Ob) into the lumbar space of three baboons. We also studied unconjugated DTPA labeled with (68)Ga, which did not move up the spinal cord but rapidly appeared in blood after i.t. injection. In contrast, G-Ob steadily moved toward the cranium and had reached the hypothalamus 91 and 139 min after i.t. injection in two baboons. We estimated the concentration of leptin in the hypothalamic region to be at least 8 ng/ml, which is about 40 times higher than cerebrospinal fluid levels in normal weight humans and about 4 times higher than the highest level ever recorded after the peripheral administration of leptin. In a third baboon, the leptin neither moved caudally nor appeared in the blood. We conclude that leptin administered i.t. can reach the hypothalamus in therapeutic concentrations, although there is considerable individual variation.

Evaluation of radiolabeled type IV collagen fragments as potential tumor imaging agents.

The objective of this study was to examine radiopharmaceuticals that target the alpha3beta1 integrin to determine if these agents target tumors for diagnostic imaging and/or targeted radiotherapy of cancer. Prior studies had shown that residues 531-542 from the alpha1 chain of type IV collagen bind a variety of tumor cell alpha3beta1 integrins. A peptide mimic of this sequence containing all D-amino acids (designated D-Hep-III) was synthesized by solid-phase methods. The tetraazamacrocyclic chelator, TETA, was conjugated to the peptide while it was resin-bound. TETA-D-Hep-III and D-Hep-III were radiolabeled with 64Cu and 125I, respectively, in high specific activity and radiochemical purity. Heterologous competitive binding assays between D-Hep-III and either 125I-D-Hep-III or 64Cu-TETA-D-Hep-III indicated low micromolar affinity of D-Hep-III. The biodistribution of each radiolabeled analogue of D-Hep-III was carried out in rats and tumor-bearing mice. Both analogues were rapidly cleared from the blood in normal rats, with the kidneys receiving the highest accumulation of each. SKOV3 human ovarian tumor cells, known to strongly express alpha3beta1, were xenografted in SCID mice. Localization of 125I-D-Hep III and 64Cu-TETA-D-Hep III in the xenografts were low (<2% ID/g), and in the case of 125I-D-Hep III, not inhibited by a competitive dose of D-Hep III. The low tumor accumulation is likely not due to receptor down-regulation, but rather due to the weak affinity of the radioligands for the alpha3beta1 integrin.

Copper-62 ATSM as a hypoxic tissue tracer in myocardial ischemia.

Copper-62 labeled diacetyl-bis(N4-methylthiosemicarbazone) (62Cu-ATSM) has been proposed as a generator produced positron-emitting tracer for hypoxic tissue imaging. To clarify the usefulness of 62Cu-ATSM for myocardial ischemia, 62Cu-ATSM PET was performed in 7 patients with coronary artery disease. Increased myocardial uptake of 62Cu-ATSM was observed (myocardium/blood ratio: 3.09) in one patient with unstable angina, who had increased 18F-fluorodeoxyglucose (18F-FDG) uptake under the fasting condition. The other 6 patients, who were clinically stable, did not have increased 62Cu-ATSM uptake, although abnormal 18F-FDG uptake was seen in 4 patients. This preliminary study suggests that 62Cu-ATSM is a promising PET tracer for hypoxic imaging in acute ischemia.

Molecular modeling of bifunctional chelate peptide conjugates. 1. Copper and indium parameters for the AMBER force field.

In this work we describe the development of parameters for In(III) and Cu(II) for the AMBER force field as found in the modeling package MacroModel. These parameters were developed using automated procedures from a combination of crystallographic structures and ab initio calculations. The new parameters were added in the form of AMBER substructures containing specific metal-ligand parameters to the existing force field. These new parameters have produced results in good agreement with experiment without requiring additional changes to the existing AMBER parameters. These parameters were then utilized to examine the conformational effects caused by the conjugation of InDTPA (DTPA = diethylenetriaminepentaacetic acid) and CuDOTA (DOTA = 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid) to the cyclic octapeptide octreotide.

Comparison of animal models for the evaluation of radiolabeled androgens.

Biodistribution of two 18F-labeled androgens and an 124I/125I-labeled androgen were studied in five androgen receptor (prostate) animal models with or lacking sex hormone binding globulin (SHBG). As models for androgen-receptor positive ovarian cancer, xenografts of three human ovarian cancer cell lines were tested in SCID mice. SHBG in the prostate model systems significantly affects the metabolism, clearance, and distribution of the radiolabeled androgens in several tissues, but ovarian cancer animal models were disappointing.

PET imaging of hypoxia.

Hypoxia in tumors has been related to poor response to conventional therapies. This paper will discuss the methods, both invasive and non-invasive, used to determine hypoxia levels within tumors. PET imaging with two lead compounds 18F-fluoromisonidazole (18FMISO) and Cu(II)-diacetyl-bis(N4-methylthiosemicarbazone (Cu-ATSM) and their relative effectiveness in delineating hypoxic regions will be discussed. The advantages of Cu-ATSM-PET over existing imaging agents will be discussed along with its potential application as a direct- and/or surrogate marker for the determination of oncological hypoxia in vivo.

Recent developments in food-matrix Reference Materials at NIST.

Since 1996, the National Institute of Standards and Technology (NIST) has developed several food-matrix Standard Reference Materials (SRMs) characterized for nutrient concentrations. These include SRM 1544 Fatty Acids and Cholesterol in a Frozen Diet Composite, SRM 1546 Meat Homogenate, SRM 1548a Typical Diet, SRM 1566b Oyster Tissue, SRM 1846 Infant Formula, and SRM 2383 Baby Food Composite. Three additional materials--SRM 1946 Lake Superior Fish Tissue, SRM 2384 Baking Chocolate, and SRM 2385 Spinach--are in preparation. NIST also recently assigned values for proximate (fat, protein, etc.), individual fatty acid, and total dietary fiber concentrations in a number of existing SRMs and reference materials (RMs) that previously had values assigned for their elemental composition. NIST has used several modes for assignment of analyte concentrations in the food-matrix RMs, including the use of data provided by collaborating laboratories, alone and in combination with NIST data. The use of data provided by collaborating food industry and contract laboratories for the analysis of food-matrix RMs has enabled NIST to provide assigned values for many analytes that NIST does not have the resources or analytical expertise to measure.

Effect of administration route on FES uptake into MCF-7 tumors.

We have observed that intraperitoneal administration of [(18)F]fluoroestradiol (FES), a radiolabeled estrogen receptor ligand, results in higher abdominal organ uptake and slower blood clearance than intravenous administration in female mice. In SCID mice bearing MCF-7 human tumors SC, IP administration resulted in tumor uptake that was only about one third that obtained with IV administration. Thus, the route of administration of a radiopharmaceutical for imaging or radiotherapy of a tumor in the abdomen, an ovarian tumor, for example, could have a profound effect on the efficiency and selectivity of delivery of the agent to the tumor.

Metabolic flare: indicator of hormone responsiveness in advanced breast cancer.

PURPOSE: The purpose of this study was to investigate whether positron emission tomography (PET) with the glucose analog [(18)F]fluorodeoxyglucose (FDG) and the estrogen analog 16 alpha-[(18)F]fluoroestradiol-17 beta (FES), performed before and after treatment with tamoxifen, could be used to detect hormone-induced changes in tumor metabolism (metabolic flare) and changes in available levels of estrogen receptor (ER). In addition, we investigated whether these PET findings would predict hormonally responsive breast cancer. PATIENTS AND METHODS: Forty women with biopsy-proved advanced ER-positive (ER(+)) breast cancer underwent PET with FDG and FES before and 7 to 10 days after initiation of tamoxifen therapy; 70 lesions were evaluated. Tumor FDG and FES uptake were assessed semiquantitatively by the standardized uptake value (SUV) method. The PET results were correlated with response to hormonal therapy. RESULTS: In the responders, the tumor FDG uptake increased after tamoxifen by 28.4% +/- 23.3% (mean +/- SD); only five of these patients had evidence of a clinical flare reaction. In nonresponders, there was no significant change in tumor FDG uptake from baseline (mean change, 10.1% +/- 16.2%; P =.0002 v responders). Lesions of responders had higher baseline FES uptake (SUV, 4.3 +/- 2.4) than those of nonresponders (SUV, 1.8 +/- 1.3; P =.0007). All patients had evidence of blockade of the tumor ERs 7 to 10 days after initiation of tamoxifen therapy; however, the degree of ER blockade was greater in the responders (mean percentage decrease, 54.8% +/- 14.2%) than in the nonresponders (mean percentage decrease, 19.4% +/- 17.3%; P =.0003). CONCLUSION: The functional status of tumor ERs can be characterized in vivo by PET with FDG and FES. The results of PET are predictive of responsiveness to tamoxifen therapy in patients with advanced ER(+) breast cancer.

The certification of SRM 1546--Meat Homogenate, a new reference material for nutrients in a high protein, high fat matrix.

In response to reference material needs expressed by the food industry and government regulators, the National Institute of Standards and Technology (NIST) has developed a new Standard Reference Material (SRM) consisting of a canned meat product with certified and reference values for a large number of constituents. SRM 1546 Meat Homogenate consists of a mixture of finely ground pork and chicken prepared and canned by a commercial process. NIST determined the concentration levels of cholesterol, sodium, calcium, iron, and seven fatty acids in this SRM using well defined methods and procedures. These analytes as well as 34 other constituents or properties were determined in an interlaboratory comparison exercise involving 21 laboratories, most of which are associated with the National Food Processors Association (NFPA) Food Industry Analytical Chemists Subcommittee (FIACS). From statistical analysis of the data, NIST assigned certified concentrations for the eleven analytes measured at NIST and reference concentrations for the proximates, six additional fatty acids, seven minerals, and seven water-soluble vitamins. Information values without uncertainties are provided for the concentrations of six additional constituents for which the uncertainties could not adequately be assessed. SRM 1546 will provide laboratories with a means to evaluate the accuracy of the methods they use to assign nutrient levels to processed meats and similar products.

Tumor uptake of copper-diacetyl-bis(N(4)-methylthiosemicarbazone): effect of changes in tissue oxygenation.

UNLABELLED: We showed previously that, in vitro, copper-diacetyl-bis(N(4)-methylthiosemicarbazone) (Cu-ATSM) uptake is dependent on the oxygen concentration (pO2). We also showed that, in vivo, Cu-ATSM uptake is heterogeneous in animal tumors known to contain hypoxic fractions. This study was undertaken to confirm the pO2 dependence of this selective uptake in vivo by correlating Cu-ATSM uptake with measured tumor pO2. METHODS: Experiments were performed with the 9L gliosarcoma rat model using a needle oxygen electrode to measure tissue pO2. Using PET and electronic autoradiography, Cu-ATSM uptake was measured in tumor tissue under various pO2 levels. The oxygen concentration within implanted tumors was manipulated by chemical means or by altering the inhaled oxygen content. RESULTS: A good correlation between low pO2 and high Cu-ATSM accumulation was observed. Hydralazine administration in animals caused a decrease in the average tumor pO2 from 28.61 +/- 8.74 mm Hg to 20.81 +/- 7.54 mm Hg in untreated control animals breathing atmospheric oxygen. It also caused the tumor uptake of Cu-ATSM to increase by 35%. Conversely, in animals breathing 100% oxygen, the average tumor pO2 increased to 45.88 +/-15.9 mm Hg, and the tumor uptake of Cu-ATSM decreased to 48% of that of the control animals. PET of animals treated in a similar fashion yielded time-activity curves showing significantly higher retention of the tracer in hypoxic tissues than in oxygenated tissues. CONCLUSION: These data confirm that Cu-ATSM uptake in tissues in vivo is dependent on the tissue pO2, and that significantly greater uptake and retention occur in hypoxic tumor tissue. Therefore, the possible use of Cu-ATSM PET as a prognostic indicator in the management of cancer is further validated.

Fluorine-substituted ligands for the peroxisome proliferator-activated receptor gamma (PPARgamma): potential imaging agents for metastatic tumors.

The peroxisome proliferator-activated receptor gamma (PPARgamma), a primary regulator of lipid metabolism, is present in many tumor cell lines and animal tumor systems and, in some cases, can mediate effective antitumor therapy with potent synthetic ligands. In an approach to image tumors with positron-emission tomography (PET) based on their content of PPARgamma, we have synthesized two fluorine-substituted analogues of a high affinity ligand from the phenylpropanoic acid class. The analogue having the highest affinity for PPARgamma was labeled with the positron-emitting radionuclide fluorine-18. In tissue distribution studies in normal rats and in SCID mice bearing human breast tumor xenografts, this compound did not show evidence of receptor-mediated uptake. The prospects for using PPARgamma as a target for imaging tumors may be limited by the low receptor concentrations in tumors and by the pharmacokinetic behavior of this class of ligands, which appears to be more favorable for therapy than for imaging.

Characterization of acetate metabolism in tumor cells in relation to cell proliferation: acetate metabolism in tumor cells.

To reveal the metabolic fate of acetate in neoplasms that may characterize the accumulation patterns of [1-(11)C]acetate in tumors depicted by positron emission tomography. Four tumor cell lines (LS174T, RPMI2650, A2780, and A375) and fibroblasts in growing and resting states were used. In uptake experiments, cells were incubated with[1-(14)C]acetate for 40 min. [(14)C]CO(2) was measured in the tight-air chamber, and the metabolites in cells were identified by thin layer chromatography and paper chromatography. The glucose metabolic rate of each cell line was measured with [2,6-(3)H]2-deoxy-glucose (DG), and the growth activity of each cell line was estimated by measuring the incorporation of [(3)H]methyl thymidine into DNA. Compared with resting fibroblasts, all four tumor cell lines showed higher accumulation of (14)C activity from [1-(14)C]acetate. These tumor-to-normal ratios of [1-(14)C]acetate were larger than those of DG. Tumor cells incorporated (14)C activity into the lipid-soluble fraction, mostly of phosphatidylcholine and neutral lipids, more prominently than did fibroblasts. The lipid-soluble fraction of (14)C accumulation in cells showed a positive correlation with growth activity, whereas the water-soluble and CO(2) fractions did not. These findings suggest that the high tumor-to-normal ratio of [1-(14)C]acetate is mainly due to the enhanced lipid synthesis, which reflects the high growth activity of neoplasms. This in vitro study suggests that [1-(11)C]acetate is appropriate for estimating the growth activity of tumor cells.

A novel approach to overcome hypoxic tumor resistance: Cu-ATSM-guided intensity-modulated radiation therapy.

PURPOSE: Locoregional tumor control for locally advanced cancers with radiation therapy has been unsatisfactory. This is in part associated with the phenomenon of tumor hypoxia. Assessing hypoxia in human tumors has been difficult due to the lack of clinically noninvasive and reproducible methods. A recently developed positron emission tomography (PET) imaging-based hypoxia measurement technique which employs a Cu(II)-diacetyl-bis(N(4)-methylthiosemicarbazone) (Cu-ATSM) tracer is of great interest. Oxygen electrode measurements in animal experiments have demonstrated a strong correlation between low tumor pO(2) and excess (60)Cu-ATSM accumulation. Intensity-modulated radiation therapy (IMRT) allows selective targeting of tumor and sparing of normal tissues. In this study, we examined the feasibility of combining these novel technologies to develop hypoxia imaging (Cu-ATSM)-guided IMRT, which may potentially deliver higher dose of radiation to the hypoxic tumor subvolume to overcome inherent hypoxia-induced radioresistance without compromising normal tissue sparing. METHODS AND MATERIALS: A custom-designed anthropomorphic head phantom containing computed tomography (CT) and positron emitting tomography (PET) visible targets consisting of plastic balls and rods distributed throughout the "cranium" was fabricated to assess the spatial accuracy of target volume mapping after multimodality image coregistration. For head-and-neck cancer patients, a CT and PET imaging fiducial marker coregistration system was integrated into the thermoplastic immobilization head mask with four CT and PET compatible markers to assist image fusion on a Voxel-Q treatment-planning computer. This system was implemented on head-and-neck cancer patients, and the gross tumor volume (GTV) was delineated based on physical and radiologic findings. Within GTV, regions with a (60)Cu-ATSM uptake twice that of contralateral normal neck muscle were operationally designated as ATSM-avid or hypoxic tumor volume (hGTV) for this feasibility study. These target volumes along with other normal organs contours were defined and transferred to an inverse planning computer (Corvus, NOMOS) to create a hypoxia imaging-guided IMRT treatment plan. RESULTS: A study of the accuracy of target volume mapping showed that the spatial fidelity and imaging distortion after CT and PET image coregistration and fusion were within 2 mm in phantom study. Using fiducial markers to assist CT/PET imaging fusion in patients with carcinoma of the head-and-neck area, a heterogeneous distribution of (60)Cu-ATSM within the GTV illustrated the success of (60)Cu-ATSM PET to select an ATSM-avid or hypoxic tumor subvolume (hGTV). We further demonstrated the feasibility of Cu-ATSM-guided IMRT by showing an example in which radiation dose to the hGTV could be escalated without compromising normal tissue (parotid glands and spinal cord) sparing. The plan delivers 80 Gy in 35 fractions to the ATSM-avid tumor subvolume and the GTV simultaneously receives 70 Gy in 35 fractions while more than one-half of the parotid glands are spared to less than 30 Gy. CONCLUSION: We demonstrated the feasibility of a novel Cu-ATSM-guided IMRT approach through coregistering hypoxia (60)Cu-ATSM PET to the corresponding CT images for IMRT planning. Future investigation is needed to establish a clinical-pathologic correlation between (60)Cu-ATSM retention and radiation curability, to understand tumor re-oxygenation kinetics, and tumor target uncertainty during a course of radiation therapy before implementing this therapeutic approach to patients with locally advanced tumor.

Standardization of cardiac troponin I assays: round Robin of ten candidate reference materials.

BACKGROUND: Cardiac troponin I (cTnI) results vary 100-fold among assays. As a step toward standardization, we examined the performance of 10 candidate reference materials (cRMs) in dilution studies with 13 cTnI measurement systems. METHODS: Solutions of 10 cTnI cRMs, each characterized by NIST, were shipped to the manufacturers of 13 cTnI measurement systems. Manufacturers used their respective diluents to prepare each cRM in cTnI concentrations of 1, 10, 25, and 50 microg/L. For the purpose of ranking the cRMs, the deviation of each cTnI measurement from the expected response was assessed after normalization with the 10 microg/L cTnI solution. Normalized deviations were examined in five formats. Parameters from linear regression analysis of the measured cTnI vs expected values were also used to rank performance of the cRMs. RESULTS: The three cRMs demonstrating the best overall rankings were complexes of troponins C, I, and T. The matrices for these three cRMs values differed; one was reconstituted directly from the lyophilized form submitted by the supplier; one was submitted in liquid form, lyophilized at NIST, and subsequently reconstituted; and the third was evaluated in the liquid form received from the supplier. The cRM demonstrating the fourth best performance was a binary complex of troponins C and I supplied in lyophilized form and reconstituted before distribution. CONCLUSIONS: The cRMs demonstrating the best performance characteristics in 13 cTnI analytical systems will be included in subsequent activities of the cTnI Standardization Committee of the AACC.