Franz Oberdorfer

FDG uptake, tumor proliferation and expression of glycolysis associated genes in animal tumor models

To determine the influence of tumor cell proliferation and changes in the genetic program in malignant cells on the fluorodeoxyglucose (FDG) uptake we performed PET studies in several animal tumors: spontaneous mammary fibroadenoma, chemically-induced mammary adenocarcinoma and Dunning prostate adenocarcinoma. The expression of the glucose transporter (GLUT1) and of hexokinase (Hk) was measured using 32P-labeled cDNA probes and densitometry. Furthermore the proliferative activity was determined with one-dimensional flow cytometry. The FDG uptake and the proliferation parameters were not correlated. The normalized amounts of GLUT and Hk mRNA were lower in spontaneous fibroadenomas and prostate tumors than in chemically induced mammary. The FDG uptake was correlated to GLUT1 expression with r = 0.83 and to Hk expression with r = 0.77. Multiple regression analysis revealed a relation of FDG uptake to GLUT1 and HK with r = 0.87. Our results show that the FDG uptake in our study was related not to differences in proliferation, but rather to differences in the transcription of glycolysis associated genes.

Bone marrow uptake of fluorine-18-fluorodeoxyglucose following treatment with hematopoietic growth factors: Initial evaluation☆

Hematopoietic growth factors (HGF) such as G-CSF and GM-CSF stimulate cell growth of the bone marrow and thereby mitigate the myelotoxic effect of chemotherapy. Using 18F-fluoro-2-deoxy-D-glucose (FDG) positron emission tomography (PET) for therapy response monitoring of patients with small-cell lung cancer, both an extension and an intensification of thoracic bone marrow uptake were noted in patients treated with HGF (n = 5) compared to those patients without HGF supplementation (n = 11). FDG uptake was a very sensitive marker of stimulated hematopoiesis, and both the extension and the intensification of uptake have to be noted during HGF therapy.

Synthesis and purification of 2-deoxy-2-[18F]fluoro-d-glucose and 2-deoxy-2-[18F]fluoro-d-mannose: characterization of products by 1H- and 19F-NMR spectroscopy

A procedure has been developed that allows the separation of 2-deoxy-2-[18F]fluoro-D-glucose from 2-deoxy-2-[18F]fluoro-D-mannose employing selectively optimized ion-moderated partition chromatography. Both compounds can be obtained with a greater than 98% chemical and radiochemical purity in about one half-life of 18F. Both the alpha- and beta-anomers of both sugars were completely characterized by high-resolution 1H- and 19F-NMR spectroscopy. Various convenient preparation methods for 2-deoxy-2-[18F]fluoro-D-glucose were compared.

Cytosine deaminase gene as a potential tool for the genetic therapy of colorectal cancer

The bacterial enzyme cytosine deaminase (CD) catalyzes the conversion of 5‐fluorocytosine (5‐FC) to the lethal 5‐fluorouracil (5‐FU) and so provides a useful system for selective killing of gene‐modified mammalian tumor cells. Cloning of the CD gene from Escherichia coli and expression in human tumor cell lines enabled these cells to convert 3H‐labeled 5‐FC into 3H‐5‐FU. Two CD‐expressing human tumor cell lines (adenocarcinoma cell line KM12 and glioblastoma cell line T1115) became 200‐fold more sensitive to 5‐FC than the nonexpressing parental cell lines. At least 90% of the cells are killed within 7 days. CD‐expressing cells are able to kill nonexpressing cells when grown in the same culture flask (bystander effect). The CD gene may be used as a suicide system for in situ chemotherapy or as a safety mechanism abrogating the expression of other genes.

Application of α-aminoisobutyric acid, L-methionine, thymidine and 2-fluoro-2-deoxy-D-glucose to monitor effects of chemotherapy in a human colon carcinoma cell line

Up to 4 h after treatment of human SW 707 colon carcinoma cells with the antineoplastic drug 4-amino-N-(2′-aminophenyl)-benzamide (GOE 1734, dinaline), the effects of tumour cell metabolism and proliferation were examined in vitro. Four tracers which can be labelled with isotopes suitable for positron emission tomography (PET) were used for this purpose:α-aminoisobutyric acid (AIB) and methionine to study changes in amino acid transport and protein synthesis, thymidine to observe changes in tumour proliferation and 2-fluoro-2-deoxy-d-glucose (FDG) to estimate glucose metabolism. Dinaline showed an inhibition of the sodium-dependent and -independent uptake of AIB. The methionine uptake was found to increase shortly after therapy. Thymidine incorporation into DNA was impaired and the FDG uptake showed a maximally 2.2-fold enhancement. Inhibition of AIB uptake suggests changes in amino acid transport, whereas increased uptake of methionine and FDG points to an enhancement of protein synthesis and glycolysis caused by repair mechanisms. The cytostatic and antiproliferative effect of dinaline, observed in cell growth curves, could be demonstrated by the impaired thymidine incorporation into DNA. This study demonstrates that in vitro screening with radiotracers suitable for PET can help to clarify effects of new antineoplastic substances on tumour cell metabolism. These data may be applied to choose the appropriate time schedule for monitoring therapeutic effects on tumour tissue.

Metabolic and transcriptional changes in osteosarcoma cells treated with chemotherapeutic drugs

Two cell lines derived from a lung metastasis of a rat osteosarcoma were treated with cisplatin (CDDP) and two phosphonic acid compounds (AMDP, DADP), AMDP-treated cells showed a decrease in FDG uptake, CDDP and DADP resulted in an increase. A block in G2 or in S and G2 phase was seen after CDDP and AMDP treatment. The changes in the cell cycle fractions were not related to the changes in FDG uptake. Furthermore, the transcription of the glucose transporter and hexokinase genes were elevated in CDDP and decreased in AMDP treated cells. However, the changes in FDG uptake were not fully explained by changes at the transcriptional level. The total uptake of thymidine was elevated although the incorporation of thymidine into DNA decreased. In both cell lines the changes in FDG uptake correlated with the changes in thymidine incorporation into DNA (r = 0.95 and r = 0.83, respectively). Cells with an increased FDG uptake showed a weaker growth inhibition than cells with a decrease in FDG uptake.

Preparation of a new 18F-labelled precursor: 1-[18F]fluoro-2-pyridone

Abstract The new 18 F-labelled precursor 1-[ 18 F]fluoro-2-pyridone was prepared by reacting [ 18 F]F 2 with 2-(trimethylsiloxy)pyridine. This compound has potential as an easy to use yet selective labelling reagent in preparations, where fluorine carrier may be tolerated. Pure (>92%) 1-[ 18 F]fluoro-2-pyridone is obtained within 35 min from end of bombardment in yields of up to 48%. Quality control consisted of HPLC combined with 19 F-NMR spectroscopy. The mean specific activity of several preparations was 4 MBq/μmol ( ϵ = 16.6). The precursor may be used directly or purified by sublimation.

A new procedure for the preparation of 11C-labelled methyl iodide

Abstract The use of diphosphorous tetraiodide for the preparation of [ 11 C]methyl iodide from [ 11 C]methanol has been investigated. This method offers several practical advantages over the established procedure using hydrogen iodide. [ 11 C]Methyl iodide, produced by this new route, has been obtained in a radiochemical yield of 80–90% with a radiochemical purity of 99.5%. The specific activity of the product has been determined and was 3.4 Ci/μmol. The reagent P 2 I 4 is reusable for several preparations.

A simple synthesis of 2-deoxy-2-fluoro-d-galactose using xenon difluoride

Abstract Treatment of tri-O-acetyl-D-galactal with xenon difluoride at −20 to +5 0 C in CFCl 3 afforded, after hydrolysis, 2-deoxy-2-fluoro-D-galactose in a smooth and stereoselective reaction. Preparative HPLC on a cation exchange resin delivered the white, crystalline product in yields of 63%.

Downregulation of T cell growth factor production by ornithine decarboxylase and its product putrescine: D,L-α-difluoromethylornithine suppresses general protein synthesis but augments simultaneously the production of interleukin-2

Treatment of EL-4 lymphoma cells with tetradecanoylphorbol-acetate (TPA), a well-known activator of protein kinase C, induces the production of the T cell growth factor interleukin-2 (IL-2) and the expression of IL-2-specific mRNA within 4-8 h. This system is an ideal model for studies on the induction of a differentiated function in a homogeneous lymphoid cell population by a defined signal. TPA induces also an increase of ornithine decarboxylase (ODC) activity and elevates the intracellular concentrations of putrescine and polyamines within 4-8 h. A similar increase of intracellular putrescine and polyamine concentrations can be achieved by administration of 2 mM putrescine to the culture medium. However, putrescine cannot induce the production of IL-2 in the absence of TPA and cannot reconstitute the IL-2 production in cultures with PGE2 or cyclosporine A, i.e., two well-known immunosuppressive substances which inhibit ODC activity. Putrescine has rather a counter-regulatory effect as concluded from the observation that the TPA-induced TCGF production and IL-2-specific mRNA expression are augmented (superinduced) by the ODC inhibitor D,L-alpha-difluoromethylornithine (DFMO) and again suppressed after the administration of putrescine or polyamines to DFMO-treated cultures. The glycolytic activity, general protein synthesis [( 3H]leucine incorporation), and the cell cycle progression from G2/M to G1, in contrast, are inhibited by DFMO and reconstituted by putrescine. This demonstrates that the cells are able to sacrifice to a large extent several vital functions including their general protein synthesis and to devote themselves at the same time to a fulminant production of their functionally most relevant protein IL-2. This process is downregulated by ODC and its product putrescine. A correlation between increased IL-2 production and accumulation of cells in the G2/M phase was also observed in cultures treated with hydroxyurea or with a combination of amethopterin and adenosine.