Cytosolic/microsomal redox pathway: a reductive retention mechanism of a PET-oncology tracer, Cu-pyruvaldehyde-bis(N 4-methylthiosemicarbazone) (Cu-PTSM)

OBJECTIVE: To clarify the retention mechanism of a PET imaging agent Cu-pyruvaldehyde-bis(N4-methylthiosemicarbazone) (Cu-62-PTSM) in tumor cells, reductive metabolism of non-radioactive Cu-PTSM in five cultured tumor cell lines, a tumor specimen and non-tumor tissues in vitro was evaluated by electron spin resonance spectrometry (ESR). RESULTS: In the brain, mitochondrial electron transport enzyme reduced Cu-PTSM specifically. On the other hand, Cu-PTSM was not reduced in tumor mitochondria. The mitochondrial electron transport enzyme in tumor cells was not damaged, but NADH was considered to be depleted. In compensation for that, the tumor cells acquired complementary reduction activity in the microsome/cytosol. The reduction was enzymatic and NADH-dependent, possibly similar to the activation mechanism of bioreductive anticancer drugs. CONCLUSION: Cu-PTSM and its derivatives are considered to be used as a marker for microsome/cytosol redox ability in PET oncology, although the physiological role of the redox enzyme system in tumor cells has not been clarified. The change in electron (NADH) flow in tumor cells might be a mechanism supporting aerobic glycolysis in tumor cells.     

Retention mechanism of hypoxia selective nuclear imaging/radiotherapeutic agent Cu-diacetyl-bis(N 4-methylthiosemicarbazone) (Cu-ATSM) in tumor cells

The retention mechanism of the novel imaging/radiotherapeutic agent, Cu-diacetyl-bis(N4-methylthiosemicarbazone) (Cu-ATSM) in tumor cells was clarified in comparison with that in normal tissue in vitro. With Cu-ATSM and reversed phase HPLC analysis, the reductive metabolism of Cu-ATSM in subcellular fractions obtained from Ehrlich ascites tumor cells was examined. As a reference, mouse brain was used. To determine the contribution of enzymes in the retention mechanisms, and specific inhibitor studies were performed. In subcellular fractions of tumor cells, Cu-ATSM was reduced mainly in the microsome/cytosol fraction rather than in the mitochondria. This finding was completely different from that found in normal brain cells. The reduction process in the microsome/cytosol was heat-sensitive and enhanced by adding exogenous NAD(P)H, an indication of enzymatic reduction of Cu-ATSM in tumor cells. Among the known bioreductive enzymes, NADH-cytochrome b5 reductase and NADPH-cytochrome P450 reductase in microsome played a major role in the reductive retention of Cu-ATSM in tumors. This enzymatic reduction was enhanced by the induction of hypoxia. Radiocopper labeled Cu-ATSM provides useful information for the detection of hypoxia as well as the microsomal bioreductive enzyme expression in tumor.     

Evaluation of copper(II)-pyruvaldehyde bis (N-4-methylthiosemicarbazone) for tissue blood flow measurement usina a trapped tracer model

Copper(II)-pyruvaldehyde bis (N-4-methylthiosemicarbazone) (Cu-PTSM) labelled with ^62,64Cu is a promising radiotracer for the study of blood flow using positron emission tomography (PET). We have investigated the application of a simple trapped tracer model to measurements of tissue ⁶⁴Cu-PTSM uptake combined with continuous arterial sampling. A dual-tracer method was used to compare blood flow estimated by ⁶⁴Cu-PTSM with values derived from measurements using cobalt-57 microspheres in the rat. Prolonged retention of ⁶⁴Cu-PTSM following intravenous administration was initially confirmed in both normal tissues and tumours. After intraventricular ⁶⁴Cu-PTSM infusion, cumulative arterial ⁶⁴Cu activity increased progressively, and after extraction in n-octanol was found to plateau to levels corresponding with those reached following administration of ⁵⁷CO microspheres. Rapid and species-dependent rates of ⁶⁴Cu-PTSM decomposition to non-extractable ⁶⁴Cu complexes were found in rat and human blood in vitro (70%±6% and 43%±5% respectively at 16 min), demonstrating the need for immediate processing of arterial samples. Close agreement was found between blood flow estimated by ⁶⁴Cu-PTSM and ⁵⁷CO microsphere methods in tissues of low to moderate flow: muscle (0.01, 0.08, 0.07 ml/min per gram; mean difference, mean ⁶⁴Cu, mean ⁵⁷Co), brain (0.09, 0.52, 0.43 ml/min per gram) and kidney (–0.16, 2.29, 2.45 ml/min per gram). Estimates of cardiac output also compared favourably between the two methods (5.7, 59.8, 54.1 ml/min). We conclude that a simple tissue trapping model may be suitable for the derivation of blood flow estimates using ^62,64Cu-PTSM, PET imaging and continuous arterial blood sampling. Correspondence to: P. Carnochan     

Studies on the67Ga uptake mechanism by Ehrlich ascites tumor cells

In order to obtain more information concerning the mechanism of Gallium (⁶⁷Ga) accumulation in malignant tissue, an investigation was carried out using Ehrlich ascites tumor cells. Chlorpromazine, a phospholipid stabilizer decreased the uptake of⁶⁷Ga and Calcium 45 (⁴⁵Ca) by the cells at low dose, but increased them at high dose. On the other hand, the uptake of both radionuclides by the cells was inhibited by ruthenium red, a Ca ATPase inhibitor in a dose dependent manner. The time course of⁶⁷Ga and⁴⁵Ca uptake were quite different from each other. Moreover, the subcellular distribution patterns of⁶⁷Ga and⁴⁵Ca were also different from each other;⁶⁷Ga accumulated in the lysosomal fraction and⁴⁵Ca mainly in the mitochondrial fraction. These results suggest that there may be a commonality for⁶⁷Ga and⁴⁵Ca uptake by the tumor cells, whereas the behaviour of these two radionuclides in the cells is dissimilar.     

Studies on the 67Ga uptake mechanism by Ehrlich ascites tumor cells

In order to obtain more information concerning the mechanism of Gallium (67Ga) accumulation in malignant tissue, an investigation was carried out using Ehrlich ascites tumor cells. Chlorpromazine, a phospholipid stabilizer decreased the uptake of 67Ga and Calcium 45 (45Ca) by the cells at low dose, but increased them at high dose. On the other hand, the uptake of both radionuclides by the cells was inhibited by ruthenium red, a Ca ATPase inhibitor in a dose dependent manner. The time course of 67Ga and 45Ca uptake were quite different from each other. Moreover, the subcellular distribution patterns of 67Ga and 45Ca were also different from each other; 67Ga accumulated in the lysosomal fraction and 45Ca mainly in the mitochondrial fraction. These results suggest that there may be a commonality for 67Ga and 45Ca uptake by the tumor cells, whereas the behaviour of these two radionuclides in the cells is dissimilar.     

Effect of neuraminidase and x-rays (2 Gy and 8 Gy) on microvilli and membrane invaginations of Ehrlich ascites tumor cells in monolayer culture

A monolayer culture (Eagle basal medium plus 10% of fetal calf serum) of Ehrlich ascites tumor cells was exposed to X-radiation with 2 Gy and 8 Gy and treated with Vibrio cholerae neuraminidase alone or combined with sublethal X-ray irradiation (2 Gy). Pictures of the Ehrlich ascites tumor cells taken with the electron microscope were investigated in order to find out any cell surface modifications due to membrane invaginations and microvilli. The results showed that the rate of microvilli as well as that of membrane invaginations became higher with the increasing X-ray dose (2 Gy; 8 Gy). Following to neuraminidase treatment there was a considerable augmentation of membrane invaginations as compared to control cells, whereas the number of microvilli was slightly reduced. As it has been already described before, the influence of neuraminidase produced an increased endocytosis activity and a strengthening of the cytoskeleton. Combined treatment with neuraminidase and sublethal X-radiation (2 Gy) caused a higher rate of membrane invaginations than each method alone; the number of microvilli was slightly increased by combined treatment. The conclusion is drawn that these structure modifications are due to reparation processes induced by radiation on the one hand and to an enzymic action of neuraminidase on the cell surface on the other hand.     

An experimental study on differential diagnosis of tumor from inflammation by using 125I labeled Pisum sativum agglutinin

We have reported that Pisum sativum agglutinin (PSA), a plant lectin which recognizes mannosyl residues, accumulates markedly in Ehrlich solid tumor (EST) and suggested the possibility of applying PSA to tumor imaging radiopharmaceuticals. In the present work, an inflammation was induced by implantation of cotton thread in the left rear leg skeletal muscle of ddY mice and Ehrlich ascites tumor cells were inoculated into the right rear leg. ⁶⁷Gacitrate accumulated in the tumor tissue and the inflammatory lesion to almost equal extents. On the other hand, ¹²⁵IPSA preferentially accumulated in tumor tissues in mice bearing both tumor and inflammation. The results suggest that differential diagnosis of tumor from inflammation using radiolabeled PSA may be possible.     

Subcellular distribution of 111In and 169Yb in tumor and liver

Subcellular distribution of ¹¹¹In and ¹⁶⁹Yb was quantitatively determined to evaluate the role of the lysosome in accumulation of these nuclides in malignant tumor tissue and in the liver using three different tumor models and the host liver. In Yoshida sarcoma and Ehrlich tumor, most of the radioactivity of these nuclides was localized in the supernatant fraction, and only a small amount of radioactivity was localized in the mitochondrial fraction, which contains lysosomes. In the liver, most of the radioactivity was concentrated in the mitochondrial fraction. The radioactivity of this fraction increased with time after the administration of these nuclides and reached approximately 50% of the total radioactivity within 24 h. In the case of hepatoma AH109A, radioactivity of the mitochondrial fraction increased with time after administration, and about 30% of the total radioactivity was concentrated in this fraction after 24 h. It is concluded that the lysosome does not play a major role in the tumor concentration of these nuclides, although it may play an important role in their liver concentration. In the case of hepatoma AH109A, it is presumed that lysosome plays a considerably important role in the tumor concentration of these nuclides, hepatoma AH109A possessing some residual features of the liver.     

Analysis of the radiorespirometric pattern after administration of [U-14C] glucose to Ehrlich ascites tumor-bearing mice

Ehrlich ascites tumor-bearing mice were subjected to ¹⁴CO₂ radiorespirometric analysis after administration of [U-¹⁴C]glucose, and the results were compared with the levels of host liver glycolytic enzyme activities and the uptake of the radioactivity into the liver. After IP administration of [U-¹⁴C]glucose, there was a progressive decline in respiratory ¹⁴CO₂ after the transplantation of the Ehrlich ascites tumor cells. The peak time (PT) was about 10 min on day 1, but thereafter was increasingly delayed, and could not be determined on day 13. Peak height (PH) and yield value (YV) were both considerably decreased, and again the magnitudes of the changes increased with the time after transplantation of the tumor cells. Glycolytic enzyme activities in the host liver were at normal levels 13 days after transplantation of the tumor cells. The uptake of the radioactivity into the liver after IP administration of [U-¹⁴C]glucose began to decline from day 5 and was 50% the value in normal mice 13 days after transplantation of the tumor cells. These results indicate that the radiorespirometric patterns with [U-¹⁴C]glucose reflects hepatic biochemical changes rather well.     

Application of lectins to tumor imaging radiopharmaceuticals

We investigated the in vitro binding of ¹²⁵I-lectins to Ehrlich ascites tumor (EAT) cells and in vivo uptake of ¹²⁵I-lectins in Ehrlich solid tumor (EST) bearing mice. In in vitro binding assays, phaseolus vulgaris agglutinin (PHA), pisum sativum agglutinin (PSA), and concanavalia agglutinin (Con A) showed a high affinity for EAT cells. The in vivo biodistribution of ¹²⁵I-lectins showed ¹²⁵I-PSA to be significantly taken up into EST tissues 24 h postinjection. After IV injection of ¹²⁵I-PSA, uptake of the radioactivity into the tumor tissues reached a maximum at 6 h, and thereafter decreased. Rapid clearance of the radioactivity from blood and its exretion into kidney soon after injection of ¹²⁵I-PSA were observed. When compared with the biodistribution of ⁶⁷Ga-citrate in EST bearing mice 24 h postinjection, tumor to liver (T/B), tumor to muscle (T/M), and tumor to blood (T/B) ratios were superior for ¹²⁵I-PSA. At 6 h postinjection, the T/B-ratio of ¹²⁵I-PSA was 2.5, and this value may be sufficient to enable discernable diagnostic images. Our results suggest that PSA might be a useful tumor imaging radiopharmaceutical.     

Optimization of tumor radiotherapy. Part VI: Modification of tumor glucose metabolism for increasing the bioavailability of 2-deoxy-D-glucose (2-DG) in a murine tumor model.

AIM: Differential radiomodification induced by 2-deoxy-D-glucose (2-DG) is proving to be a feasible modality for optimizing tumor radiotherapy. Our earlier work on Ehrlich ascites tumor cells has shown that pretreatment with hematoporphyrin derivatives increases the uptake and phosphorylation of 2-DG. Moreover, the alteration induced in bioenergetic profile was more drastic and less reversible. The promising combination of hematoporphyrin derivatives and 2-DG has been further evaluated in the Ehrlich ascites tumor bearing mice for determining the effects on radiotherapeutic response. MATERIALS AND METHODS: Solid tumors (average volume = 0.9 +/- 0.1 cm3) implanted in Swiss-albino strain "A" mice were focally irradiated (10 Gy) using 60Co teletherapy. Drugs were administered intravenously. Tumor bioenergetics was assessed by 31P MR spectroscopy. RESULTS: The uptake and phosphorylation of 2-DG was observed to be increased following pretreatment with hematoporphyrin derivatives. Upon hematoporphyrin derivatives + 2-DG treatment followed by irradiation, the intracellular pH reduced and a remarkable increase in glycerophosphorylcholine and inorganic phosphate levels was observed. CONCLUSION: The present study demonstrates the potential of hematoporphyrin derivative pretreatment in increasing the bioavailability of 2-DG in a mice Ehrlich ascites tumor model. The finding may have interesting clinical implications in the form of increased manifestation of the radiation-induced damage in the case of use of these drugs as a potential adjuvant in radiotherapy of tumors.     

Comparisons of labeling efficiency,biological activity and biodistribution among 125I-, 67Ga-DTPA-and 67Ga-DFO-lectins

The labeling efficiency, biological activity and biodistribution of ¹²⁵I labeled and ⁶⁷Ga chelating agent conjugated lectins were investigated. Pisum sativum agglutinin (PSA) and Lens culinaris agglutinin (LCH) were efficiently labeled with ⁶⁷Ga using bifunctional chelating agents such as diethylenetriaminepentaacetic acid (DTPA) and deferoxamine (DFO), whereas labeling with ¹²⁵I was significantly less efficient. The agglutinating activity of these lectins towards Ehrlich ascites tumor (EAT) cells was retained on conjugation with DFO, but not with DTPA. The in vitro binding ratio of ⁶⁷Ga-DFO-lectins for EAT cells was almost the same as that of ¹²⁵I-lectins. However, the value was significantly decreased in the case of ⁶⁷Ga-DTPA-lectins. In the biodistribution study of radiolabeled lectins in Ehrlich solid tumor (EST) bearing mice, the accumulation of radioactivity in tumor tissue was very much less with ⁶⁷Ga-DTPA-lectins than with ¹²⁵I-lectins. However, the concentration was significantly elevated in the case of ⁶⁷Ga-DFO-lectins. While, these lectins accumulated in liver, spleen, lung, and kidney to a greater extent than ⁶⁷Ga citrate, the tumor to organ ratios became very low. These low tumor to organ ratios, in contrast to ⁶⁷Ga citrate, will certainly inhibit the tumor delineation, and therefore it seems that in spite of a high accumulation ratio of ⁶⁷Ga-DFO-lectins in tumor tissue, these agents are not useful in tumor detection.     

Synthesis and evaluation of copper radiopharmaceuticals with mixed bis(thiosemicarbazone) ligands

Four “mixed” bis(thiosemicarbazone) derivatives of pyruvaldehyde were synthesized that incorporate two dissimilar thiosemicarbazone functions. The corresponding [⁶⁷Cu]copper(II) complexes were prepared and evaluated as possible copper radiopharmaceuticals. The pyruvaldehyde-based mixed bis(thiosemicarbazone) ligands, CH₃C[=NNHC(S)NHMe]CH[=NNHC(S)NHEt] (1), CH₃C[=NNHC(S)NHMe]CH[=NNHC(S)NEt₂] (2), CH₃C[=NNHC(S)NHMe]CH[=NNHC(S)-cyclo-N(CH₂)₅] (3), and CH₃C[=NNHC(S)NHMe]CH[=NNHC(S)-cyclo-N(CH₂)₆] (4), were obtained by reaction of the appropriate thiosemicarbazide derivative with pyruvaldehyde-2-N⁴-methylthiosemicarbazone (CH₃C[=NNHC(S)NHMe]CHO). The ⁶⁷Cu-labeled copper(II) complexes of ligands 1-4 were prepared and screened in a rat model to assess the potential of each chelate as a ⁶²Cu-radiopharmaceutical for imaging with positron emission tomography. The ⁶⁷Cu-complexes of ligands 1–4 exhibit significant uptake into the brain and heart 1 min following intravenous administration to rats. For the ⁶⁷Cu-complexes of ligands 2, 3, and 4, the cerebral and myocardial uptake of ⁶⁷Cu is two-to-threefold lower at 2 h than at 1 min postinjection, due to significant biological clearance of these ⁶⁷Cu-chelates. However, the ⁶⁷Cu-complex of 1 affords cerebral and myocardial uptake and retention comparable to that of [⁶⁷Cu]Cu-PTSM in this model. Although the kinetics of this new agent appear attractive, ultrafiltration studies using solutions of dog and human serum albumin reveal that the ⁶⁷Cu-complex of ligand 1, like Cu-PTSM, interacts more strongly with human albumin than dog albumin. Thus, this new agent would appear to offer no advantage over Cu-PTSM as a ⁶²Cu-labeled tracer for evaluation of regional tissue perfusion.     

An experimental study on differential diagnosis of tumor from inflammation by using 125I labeled Pisum sativum agglutinin

We have reported that Pisum sativum agglutinin (PSA), a plant lectin which recognizes mannosyl residues, accumulates markedly in Ehrlich solid tumor (EST) and suggested the possibility of applying PSA to tumor imaging radiopharmaceuticals. In the present work, an inflammation was induced by implantation of cotton thread in the left rear leg skeletal muscle of ddY mice and Ehrlich ascites tumor cells were inoculated into the right rear leg. 67Ga-citrate accumulated in the tumor tissue and the inflammatory lesion to almost equal extents. On the other hand, 125I-PSA preferentially accumulated in tumor tissues in mice bearing both tumor and inflammation. The results suggest that differential diagnosis of tumor from inflammation using radiolabeled PSA may be possible.     

A comparison of PET imaging characteristics of various copper radioisotopes

Purpose PET radiotracers which incorporate longer-lived radionuclides enable biological processes to be studied over many hours, at centres remote from a cyclotron. This paper examines the radioisotope characteristics, imaging performance, radiation dosimetry and production modes of the four copper radioisotopes, ⁶⁰Cu, ⁶¹Cu, ⁶²Cu and ⁶⁴Cu, to assess their merits for different PET imaging applications. Methods Spatial resolution, sensitivity, scatter fraction and noise-equivalent count rate (NEC) are predicted for ⁶⁰Cu, ⁶¹Cu, ⁶²Cu and ⁶⁴Cu using a model incorporating radionuclide decay properties and scanner parameters for the GE Advance scanner. Dosimetry for ⁶⁰Cu, ⁶¹Cu and ⁶⁴Cu is performed using the MIRD model and published biodistribution data for copper(II) pyruvaldehyde bis(N⁴-methyl)thiosemicarbazone (Cu-PTSM). Results ⁶⁰Cu and ⁶²Cu are characterised by shorter half-lives and higher sensitivity and NEC, making them more suitable for studying the faster kinetics of small molecules, such as Cu-PTSM. ⁶¹Cu and ⁶⁴Cu have longer half-lives, enabling studies of the slower kinetics of cells and peptides and prolonged imaging to compensate for lower sensitivity, together with better spatial resolution, which partially compensates for loss of image contrast. ⁶¹Cu-PTSM and ⁶⁴Cu-PTSM are associated with radiation doses similar to [¹⁸F]-fluorodeoxyglucose, whilst the doses for ⁶⁰Cu-PTSM and ⁶²Cu-PTSM are lower and more comparable with H₂¹⁵O. Conclusion The physical and radiochemical characteristics of the four copper isotopes make each more suited to some imaging tasks than others. The results presented here assist in selecting the preferred radioisotope for a given imaging application, and illustrate a strategy which can be extended to the majority of novel PET tracers.     

Cu(II) bis(thiosemicarbazone) radiopharmaceutical binding to serum albumin: further definition of species dependence and associated substituent effects

IntroductionThe pyruvaldehyde bis(N⁴-methylthiosemicarbazonato)copper(II) (Cu-PTSM) and diacetyl bis(N⁴-methylthiosemicarbazonato)copper(II) (Cu-ATSM) radiopharmaceuticals exhibit strong, species-dependent binding to the IIA site of human serum albumin (HSA), while the related ethylglyoxal bis(thiosemicarbazonato)copper(II) (Cu-ETS) radiopharmaceutical appears to exhibit only nonspecific binding to HSA and animal serum albumins.MethodsTo further probe the structural basis for the species dependence of this albumin binding interaction, we examined protein binding of these three radiopharmaceuticals in solutions of albumin and/or serum from a broader array of mammalian species (rat, sheep, donkey, rabbit, cow, pig, dog, baboon, mouse, cat and elephant). We also evaluated the albumin binding of several copper(II) bis(thiosemicarbazone) chelates offering more diverse substitution of the ligand backbone.ResultsCu-PTSM and Cu-ATSM exhibit a strong interaction with HSA that is not apparent with the albumins of other species, while the binding of Cu-ETS to albumin is much less species dependent. The strong interaction of Cu-PTSM with HSA does not appear to simply correlate with variation, relative to the animal albumins, of a single amino acid lining HSA's IIA site. Those agents that selectively interact with HSA share the common feature of only methyl or hydrogen substitution at the carbon atoms of the diimine fragment of the ligand backbone.ConclusionsThe interspecies variations in albumin binding of Cu-PTSM and Cu-ATSM are not simply explained by unique amino acid substitutions in the IIA binding pocket of the serum albumins. However, the specific affinity for this region of HSA is disrupted when substituents bulkier than a methyl group appear on the imine carbons of the copper bis(thiosemicarbazone) chelate.     

Analysis of the Radiorespirometric pattern after administration of [U-14C] glucose to Ehrlich ascites tumor-bearing mice

Ehrlich ascites tumor-bearing mice were subjected to 14CO2 radiorespirometric analysis after administration of [U-14C]glucose, and the results were compared with the levels of host liver glycolytic enzyme activities and the uptake of the radioactivity into the liver. After IP administration of [U-14C]glucose, there was a progressive decline in respiratory 14CO2 after the transplantation of the Ehrlich ascites tumor cells. The peak time (PT) was about 10 min on day 1, but thereafter was increasingly delayed, and could not be determined on day 13. Peak height (PH) and yield value (YV) were both considerably decreased, and again the magnitudes of the changes increased with the time after transplantation of the tumor cells. Glycolytic enzyme activities in the host liver were at normal levels 13 days after transplantation of the tumor cells. The uptake of the radioactivity into the liver after IP administration of [U-14C]glucose began to decline from day 5 and was 50% the value in normal mice 13 days after transplantation of the tumor cells. THese results indicate that the radiorespirometric patterns with [U-14C]glucose reflects hepatic biochemical changes rather well.     

Heterogeneity of regional nitrogen 13-labeled ammonia tracer distribution in the normal human heart: Comparison with rubidium 82 and copper 62-labeled PTSM

Background

Recent reports on¹³N-labeled ammonia (¹³N-ammonia) positron emission tomographic (PET) imaging have suggested a relative reduction of measured tracer activity in the posterolateral wall. Such inhomogeneity of tracer distribution could potentially affect accuracy for detection of disease. The aim of this study was to compare the regional distribution of¹³N-ammonia with⁸²Rb and⁶²Cu-labeled PTSM (⁶²Cu-PTSM) to identify tracer-specific patterns that may be important in the clinical interpretation of cardiac flow studies.

Methods and Results

Twenty-eight healthy volunteers underwent PET imaging at rest with either¹³N-ammonia (n=14),⁸²Rb (n=8), or⁶²Cu-PTSM (n=6). Eight subjects given¹³N-ammonia also underwent imaging after adenosine. Activity measured in the posterolateral wall on transaxial images was significantly lower than in the septum for¹³N-ammonia, both at rest (p<0.005) and after adenosine (p<0.05). No differences were detected for⁸²Rb or⁶²Cu-PTSM. The septum/posterolateral wall activity ratios for¹³N-ammonia,⁸²Rb, and⁶²Cu-PTSM were 1.15±0.07, 1.00±0.06, and 0.97±0.08, respectively (p<0.001). Regional analysis of image data showed the percent of maximal activity data for¹³N-ammonia in the lateral wall to be less than that of other regions (p<0.001) and in the inferior wall to be greater than in the anterior and lateral walls (p<0.001). For⁶²Cu-PTSM, activity in the inferior wall was greater than that in other regions (p<0.005). No regional differences were detected for⁸²Rb.

Conclusions

The relatively increased wall activity with¹³N-ammonia and⁶²Cu-PTSM is most likely due to cross-contamination of activity from the liver. The significant reduction in activity in the lateral wall with¹³N-ammonia, which persists after adenosine, is most likely related to regional heterogeneity in¹³N-ammonia retention and may reflect regional differences in metabolic-trapping mechanisms for¹³N-ammonia. Further investigation is required to elucidate the underlying mechanism of this phenomenon. Reduced tracer retention in the lateral wall segment as a normal variant must be considered when evaluating clinical¹³N-ammonia PET studies.     

The prevention of Ehrlich ascites tumor using intraperitoneal colloidal 198Au. Dose vs. size of inoculum.

Previous work has shown that radioactive colloids can cure Ehrlich ascites tumor. In this experiment, the dose of colloidal 198Au needed has been correlated with the number of tumor cells inoculated. On Day 1, varying numbers of Ehrlich cells (10(2), 10(3), 10(4), 10(5), 10(6)) were injected intraperitoneally into 310 BALB/c male mice. Two hours later, varying doses (10, 20, 40, 80, and 160 muCi) of 198Au were injected intraperitoneally. Controls were injected with normal saline. Generally, the lower cell inoculums and higher doses of 198Au yielded better survival rates.     

Kinetics of radioiodinated species in subcellular fractions from rat hearts following administration of iodine-123-labelled 15-(p-iodophenyl)-3-(R,S)-methylpentadecanoic acid (123I-BMIPP)

It is recognized that iodine-123-labelled 15-(p-iodophenyl)-3-(R,S)-methylpentadecanoic acid (¹²³IBMIPP) slowly washes out of the myocardium. The mechanism for the washout was investigated in normal rat hearts by analyses of the subcellular distribution and lipid classes based on the BMIPP metabolism. Rat hearts were excised at 1–120 min after intravenous injection of¹²³I-BMIPP. After counting the radioactivity, the hearts were digested with Nagarse and homogenized, and then fractionated into the cytosolic, mitochondrial, microsomal and crude nuclear fractions by centrifugations. The radioactivity of each fraction was counted, and the lipid classes were analysed by radio-thin-layer chromatographic and high-performance liquid chromatographic methods. The heart uptake of 1231-BMIPP was maximal at 5 min (6.81%±0.36% ID/g), and 41% of the radioactivity disappeared within 120 min. The myocardial radioactivity was immediately distributed into the cytosolic, mitochondrial, microsomal and crude nuclear fractions. The distribution (%) of each fraction was almost identical from 5 min through 120 min. The cytosolic fraction was always the major site of radioactivity deposition (60%), and the time-activity curve of the cytosolic fraction paralleled that of the whole heart throughout the 120-min study period. In the cytosolic fraction, most of the radioactivity was incorporated into the triglyceride class, and the rest was present in the free fatty acid, phospholipid (phosphatidylcholine) and diglyceride classes. In the mitochondrial fraction, the radioactivity was mostly incorporated into the phospholipid class (phosphatidylethanolamine), followed by free fatty acids. The final metabolite of¹²³I-BMIPP,¹²³I-p-iodophenylacetic acid (¹²³I-PIPA), initially appeared in the mitochondrial fraction as early as 1 min, and subsequently in the cytosolic fraction at 5 min. Another intermediary metabolite,¹²³I-p-iodophenyldodecanoic acid (¹²³I-PIPC₁₂), was found only in the mitochondrial fraction after 5 min. In conclusion, the slow washout kinetics of¹²³I-BMIPP from the myocardium mainly reflects the turnover rate of the triglyceride pool in the cytosol. The BMIPP metabolism, i.e. initial -oxidation followed by subsequent cycles of -oxidation, was confirmed in vivo. The participation of the mitochondria in the metabolism was also proven.