Biomedical subjects

A Altmann

Publications and source records attributed to A Altmann.

At least 19 recordsLinked to original sources

PubMed2004

Antibody persistence, PRP-specific immune memory, and booster responses in infants immunised with a combination DTPa-HBV-IPV/Hib vaccine.

A new single-injection combination vaccine against six diseases has been developed to accommodate the growing number of recommended paediatric vaccines. A pentavalent liquid diphtheria, tetanus, acellular pertussis (3-component), hepatitis B, and inactivated polio (types 1-3) combined vaccine (DTPa-HBV-IPV) is extemporaneously mixed with a lyophilized Haemophilus influenza type B (Hib) conjugate vaccine (polyribosyl-ribitol phosphate (PRP)-T) and given as a single-injection. A cohort of 368 healthy infants was initially studied to evaluate the immunogenicity and reactogenicity of this hexavalent combination given as a primary course at 2, 4, and 6 months of age. At 15 months of age, from this cohort, 219 children received a booster dose of a licensed DTPa/Hib (PRP-T) vaccine to assess the booster response, while 70 received a challenge dose of unconjugated PRP (PRP) vaccine (to evaluate Hib-specific memory) plus a separate DTPa vaccine. Seven to 10 days following plain PRP challenge, anti-PRP geometric mean antibody concentrations (GMCs) had increased 13-fold to 5.67 microg/ml, and thirty days after conjugated PRP booster vaccination, anti-PRP antibody GMCs increased 102-fold. Both responses are indicative of immune memory. Vaccination was well tolerated following all primary and booster doses, although 10.5% of booster recipients experienced >50-mm local swelling at the site of DTPa vaccination. We conclude that DTPa-HBV-IPV/Hib is safe and immunogenic for primary vaccination, and that Hib-specific memory is induced by primary vaccination.

Antibodies, Bacterial↗

PubMed2003

Enhanced iodide transport after transfer of the human sodium iodide symporter gene is associated with lack of retention and low absorbed dose.

Transfer of the sodium iodide symporter (hNIS) has been proposed as a new principle of cancer gene therapy. Using clinically relevant doses of (131)I for the treatment of NIS-expressing prostate carcinoma cells, we investigated the kinetics and the absorbed doses obtained in these tumors. hNIS-expressing cell lines accumulated up to 200 times more iodide when compared to wild-type cells. However, a rapid efflux of the radioactivity (80%) occurred during the first 20 min after replacement of the medium. In rats, the hNIS-expressing tumors accumulated up to 20 times more iodide when compared to contralateral transplanted wild-type tumors. After 24 h and doses of 550, 1200 or 2400 MBq/m(2) hNIS-expressing tumors lost 89, 89 and 91% of the initial activity, respectively. Dosimetric calculations showed that 1200 MBq/m(2) resulted in 3+/-0.5 Gy (wild-type tumor 0.15+/-0.1 Gy) and 2400 MBq/m(2) resulted in 3.1+/-0.9 Gy (wild-type tumor 0.26+/-0.02 Gy). Although transduction of the hNIS gene induces iodide transport in rat prostate adenocarcinoma a rapid efflux occurs, which leads to a low absorbed dose in genetically modified tumors. With regard to a therapeutic application additional conditions need to be defined leading to iodide trapping.

Absorption↗

PubMed2001

Apoptosis and changes in glucose transport early after treatment of Morris hepatoma with gemcitabine.

Apoptosis has been described as an energy-consuming process. This combined in vivo/in vitro study investigated the effects of the antineoplastic agent gemcitabine on tumour metabolism and on the induction of apoptosis. Dynamic positron emission tomography (PET) measurements of fluorine-18 fluorodeoxyglucose (FDG) uptake were done in rats bearing Morris hepatoma prior to and after therapy with 90 mg gemcitabine/kg b.w. Furthermore, thymidine (TdR) incorporation into the DNA of these tumours was determined. In vitro measurements of FDG and TdR uptake were performed immediately and 24 h after the end of gemcitabine treatment, and the amount of apoptotic cells was determined using the TUNEL reaction. In vivo an increase in FDG transport and phosphorylation occurred early after gemcitabine treatment, although TdR incorporation into the DNA of the tumours declined. In vitro, an enhanced glucose transport, an increase in TdR uptake in the cytoplasm and a decrease in TdR incorporation in the nucleic acid fraction early after treatment occurred. Inhibition of glucose transport caused an increase in the amount of apoptotic cells. The increase in glucose uptake and TdR metabolism early after therapy is interpreted as a stress reaction of the tumour cells, protecting the cells from apoptosis during this early period after exposure to cytotoxic drugs like gemcitabine.

3-O-Methylglucose↗

PubMed2001

Iodide uptake in human anaplastic thyroid carcinoma cells after transfer of the human thyroid peroxidase gene.

Human thyroperoxidase (hTPO) is critical for the accumulation of iodide in thyroid tissues. Poorly differentiated and anaplastic thyroid tumours which lack thyroid-specific gene expression fail to accumulate iodide and, therefore, do not respond to iodine-131 therapy. We consequently investigated whether transfer of the hTPO gene is sufficient to restore the iodide-trapping capacity in undifferentiated thyroid and non-thyroid tumour cells. The human anaplastic thyroid carcinoma cell lines C643 and SW1736, the rat Morris hepatoma cell line MH3924A and the rat papillary thyroid carcinoma cell line L2 were used as in vitro model systems. Employing a bicistronic retroviral vector based on the myeloproliferative sarcoma virus for the transfer of the hTPO and the neomycin resistance gene, the C643 cells and SW1736 cells were transfected while the L2 cells and MH3924A cells were infected with retroviral particles. Seven recombinant C643 and seven SW1736 cell lines as well as four recombinant L2 and four MH3924A cell lines were established by neomycin selection. They were studied for hTPO expression using an antibody-based luminescence kit, followed by determination of the enzyme activity in the guaiacol assay and of the iodide uptake capacity in the presence of Na125I. Genetically modified cell lines expressed up to 1,800 times more hTPO as compared to wild type tumour cells. The level of hTPO expression varied significantly between individual neomycin-resistant cell lines, suggesting that the recombinant retroviral DNA was integrated at different sites of the cellular genome. The accumulation of iodide, however, was not significantly enhanced in individual recombinant cell lines, irrespective of low or high hTPO expression. Moreover, there was no correlation between hTPO expression and enzyme activity in individual cell lines. The transduction of the hTPO gene per se is not sufficient to restore iodide trapping in non-iodide-concentrating tumour cells. Future studies will have to concentrate on the possible expression of enzymatically active proteins or the transfer of multiple genes involved in iodide trapping.

Animals↗

PubMed2001

Glucose transport and apoptosis after gene therapy with HSV thymidine kinase.

The relation between tumour metabolism and induction of apoptosis by gene therapy was investigated in a rat Morris hepatoma (MH3924A) model expressing the HSV thymidine kinase (HSVtk) gene. In vivo the amount of glucose transporter (GLUT1 and GLUT3 isoforms) expressing cells was determined in tumours of untreated and treated animals using immunohistochemistry. In vitro uptake studies with 2-fluoro-2-deoxy-D-glucose (FDG), 3-O-methylglucose and thymidine (TdR) and a TUNEL (TdT-mediated dUTP nick end labelling) assay for the assessment of apoptosis were done immediately and 24 h after treatment of the recombinant cells with different doses of ganciclovir (GCV). Immunohistochemistry revealed a significant increase in GLUT1 in treated tumours which showed enhanced transport activity for FDG. In vitro the FDG and 3-O-methylglucose uptake increased to 186% when compared with that of the non-treated cells immediately after incubation with GCV. However, 24 h later the FDG uptake had declined to its normal level, whereas the accumulation of 3-O-methylglucose remained elevated. The uptake of TdR, which was determined simultaneously, decreased in the acid-insoluble fraction of the cells to 27% and 11%, respectively, immediately and 24 h after therapy, while in the acid-soluble fraction it increased to 229% and to 167%, respectively. Employing the TUNEL technique, 25% of cells were found to be apoptotic 24 h after the termination of GCV treatment. Inhibition of glucose transport by cytochalasin B or competition with deoxyglucose resulted in a 78% (cytochalasin B) and 88% (deoxyglucose) decrease in FDG uptake in the recombinant hepatoma cells and in an increase in the apoptotic cell fraction. It is concluded that inhibition of enhanced glucose transport in GCV-treated cells increased apoptosis. Therefore, enhanced glucose transport seems to represent a stress reaction of tumour cells dedicated for the prevention of cell death.

3-O-Methylglucose↗

PubMed2001

Tissue-specific gene expression in medullary thyroid carcinoma cells employing calcitonin regulatory elements and AAV vectors.

Calcitonin (CT), the major secretory product of the C cell, is also expressed in C-cell-derived neoplasia. To investigate the role of the CT gene regulatory sequence in tissue-specific gene expression, the genes coding for the herpes simplex virus thymidine kinase (HSVtk) and for the enhanced green fluorescent protein (EGFP) regulated by the CT promoter (rAAV/CT266tkneo), the CT promoter/enhancer element (rAAV/CTenhtkneo), or the cytomegalovirus (CMV) promoter (rAAV/CMVtkneo) were transduced by recombinant adenoassociated viral (AAV) vectors into the medullary thyroid carcinoma (MTC) cell lines TT and hMTC and into HeLa cells as controls. In TT cell lines and hMTC cell lines transiently infected by the rAAV/CT266tkneo viruses, a significant increase in (3)H ganciclovir uptake was observed. Upon ganciclovir treatment, TT cells infected by rAAV/CT266tkneo revealed a significant growth inhibition, which was less tissue-specific because HeLa cells were also affected by these particles (74.5%). In contrast, a minor but more tissue-specific growth inhibition (33.6%) was observed for TT cells after transient infection with the rAAV/CTenhtkneo particles. Employing EGFP controlled by CMV promoter and the individual CT regulatory sequences for transduction by rAAV particles, similar results were obtained indicating that both the CT promoter and enhancer element are required for tissue-specific gene expression in MTC.

Calcitonin↗

PubMed2001

Imaging methods in gene therapy of cancer.

Clinical gene therapy needs non invasive tools to evaluate the efficiency of gene transfer. This includes the evaluation of infection efficiency as well as the verification of successful gene transfer in terms of gene transcription. These informations can be used for therapy planning, follow up studies in treated tumors and as an indicator of prognosis. Therapy planning is performed by the assessment of gene expression for example using radiolabeled specific substrates to determine the activity of suicide enzymes as the Herpes Simplex Virus thymidine kinase or cytosine deaminase. Furthermore, other in vivo reporter genes as receptors, antigens or transport proteins may be used in bicistronic vector systems for the evaluation of gene transduction and expression. This is done using radiolabeled ligands, antigens or substrates. Follow up studies with magnetic resonance imaging, single photon emission tomography or positron emission tomography may be done to evaluate early or late effects of gene therapy on tumor volume, metabolism or proliferation. Finally, enhancement of radioactive isotope accumulation in tumors by transfer of the appropriate genes may be used for the treatment of malignant tumors.

Animals↗

PubMed2001

Transfer of the human NaI symporter gene enhances iodide uptake in hepatoma cells.

UNLABELLED: The characteristic feature of thyroid cells of taking up iodide enables benign thyroid diseases and differentiated thyroid carcinoma to be successfully treated with radioiodide therapy. The transport of iodide across the cell membrane is mediated by the human NaI symporter (hNIS). We therefore investigated whether the accumulation of iodide may be induced by the retroviral transfer of the hNIS gene in nonthyroid tumor cells. METHODS: With use of a bicistronic retroviral vector for the transfer of the hNIS coding sequence and the hygromycin resistance gene, rat Morris hepatoma (MH3924A) cells were infected with retroviral particles and 32 hNIS-expressing cell lines were generated by hygromycin selection. After incubation of the genetically modified and wild-type hepatoma cells and the rat thyroid cell line FRTL5 with Na125I, the uptake and efflux of iodide were determined. In addition, the iodide distribution in rats bearing wild-type and genetically modified hepatomas was monitored. RESULTS: Genetically modified MH3924A cell lines accumulated up to 235 times more iodide than did noninfected hepatoma cells. The maximal iodide uptake in the cells was observed after 60 min incubation time. Competition experiments in the presence of sodium perchlorate revealed a dose-dependent decrease of iodide uptake (87%-92%). Moreover, carbonyl cyanide p-trifluoromethoxyphenylhydrazone led to a loss of accumulated I- (32%), whereas 4,4'-diisothiocyano-2,2'-disulfonic acid stilbene increased the I- uptake into the cells (22%). However, a rapid efflux of the radioactivity (80%) was observed during the first 10 min after 125I(-)-containing medium had been replaced by nonradioactive medium. In rats, the hNIS-expressing tumors accumulated six times more iodide than did the contralateral wild-type tumor as monitored by scintigraphy. The ex vivo quantitation of the iodide content performed 1 h after tracer administration in 1 g of tumor tissue revealed a 17-fold higher iodide accumulation in the genetically modified tumors. In accordance with the in vitro data, we also observed a rapid efflux of radioactivity from the tumor in vivo. CONCLUSION: The transduction of the hNIS gene per se is sufficient to induce 125I transport in Morris hepatoma cells in vitro and in vivo. With regard to a therapeutic application, however, additional conditions need to be defined that inhibit the iodide efflux from the tumor cells.

Animals↗

PubMed2000

Hippocampal malrotation with normal corpus callosum: a new entity?

Among 527 MRI examinations of patients with a suspicion of epilepsy in 5 years, we found 32 cases of hippocampal malrotation (HIMAL). The characteristic features are: incomplete inversion of the hippocampus with and abnormally round shape; unilateral involvement of the whole hippocampus; normal signal intensity and size; blurred internal structure; an abnormal angle of collateral sulcus; abnormal position and size of the fornix; normal size of the temporal lobe; enlargement and particular configuration of the temporal horn, typical of corpus callosum agenesis; and a normal corpus callosum. In 7 cases (22%) HIMAL occurred together with developmental disorders. It was predominantly seen in men. The clinical features were varied. Based on some MRI features, the presence of developmental disorders, the male predominance, the frequently positive family history, and a review of the literature, we think HIMAL may be the consequence of a mild hemisphere developmental disorder. It is probably not the basic cause of epilepsy in such varied clinical setting, but may be a sign of a developmental disorder and can help in selecting patients for more meticulous investigation. It also may give some new understanding of brain development.

Adult↗

PubMed1999

Is sudden infant death syndrome still more common in very low birthweight infants in the 1990s?

OBJECTIVE: To determine the rate of sudden infant death syndrome (SIDS) in very low birthweight children (VLBW) relative to children with low (LBW) and normal birthweights. DESIGN, SETTING AND SUBJECTS: Cohort study of consecutive live births in Victoria, 1993-1997 inclusive. MAIN OUTCOME MEASURES: All sudden unexpected deaths in early childhood over this five-year period; all deaths from SIDS (defined as a sudden unexpected death without a definite pathological explanation); and the proportion of SIDS in live births in three birthweight subgroups (VLBW, 500-1499 g; LBW, 1500-2499 g; and normal birthweight, > 2499 g). RESULTS: There were 316,028 live births (with known birthweight) in Victoria over the five-year period; 224 (0.71 per 1000 live births) died unexpectedly. In 10 of these deaths there was a definite pathological explanation, giving a rate of SIDS of 0.68 per 1000 live births. The rate of SIDS in VLBW children was 2.52 per 1000 live births, lower than the rate reported before the 1990s. The rate of SIDS in VLBW children was not significantly different from the rate in LBW children of 1.98 per 1000 live births (difference per 1000 live births, 0.53; 95% CI, -1.45 to 2.52), but was significantly higher than the rate in normal birthweight children of 0.59 per 1000 live births (difference per 1000 live births, 1.93; 95% CI, 0.06-3.79). CONCLUSIONS: The rate of SIDS in VLBW children has fallen in the 1990s, along with the overall fall in the rate of SIDS, but remains higher than that in normal birthweight children.

Humans↗

PubMed1998

Ganciclovir uptake in human mammary carcinoma cells expressing herpes simplex virus thymidine kinase.

Assessment of suicide enzyme activity would have considerable impact on the planning and the individualization of suicide gene therapy of malignant tumors. This may be done by determining the pharmacokinetics of specific substrates. We generated ganciclovir (GCV)-sensitive human mammary carcinoma cell lines after transfection with a retroviral vector bearing the herpes simplex virus thymidine kinase (HSV-tk) gene. Thereafter, uptake measurements and HPLC analyses were performed up to 48 h in an HSV-tk-expressing cell line and in a wild-type cell line using tritiated GCV. HSV-tk-expressing cells showed higher GCV uptake and phosphorylation than control cells, whereas in wild-type MCF7 cells no phosphorylated GCV was detected. In bystander experiments the total GCV uptake was related to the amount of HSV-tk-expressing cells. Furthermore, the uptake of GCV correlated closely with the growth inhibition (r = 0.92). Therefore, the accumulation of specific substrates may serve as an indicator of the HSV-tk activity and of therapy outcome. Inhibition and competition experiments demonstrated slow transport of GCV by the nucleoside carriers. The slow uptake and low affinity to HSV-tk indicate that GCV is not an ideal substrate for the nucleoside transport systems or for HSV-tk. This may be the limiting factor for therapy success, necessitating the search for better substrates of HSV-tk.

Antiviral Agents↗

PubMed1998

Uncoupling of 2-fluoro-2-deoxyglucose transport and phosphorylation in rat hepatoma during gene therapy with HSV thymidine kinase.

This animal study investigates the application of positron emission tomography (PET) with tracers of tumour metabolism for monitoring suicide gene therapy with herpes simplex virus thymidine kinase (HSVtk). After transplantation of HSVtk-expressing Morris hepatoma cells into ACI rats, dynamic PET measurements of 18F-labeled 2-fluoro-2-deoxyglucose (FDG) uptake were performed in animals 2 days (n = 7) and 4 days (n = 5) after the onset of therapy with 100 mg ganciclovir (GCV)/kg body weight as well as after administration of sodium chloride (n = 8). The arterial FDG plasma concentration was measured dynamically in an extracorporeal loop and the rate constants for FDG transport (K1, k2) and FDG phosphorylation (k3) were calculated using a three-compartment model modified for heterogeneous tissues. Also, quantification using the metabolic rate of FDG turnover and the standardized uptake value (SUV) was done. Furthermore, the thymidine incorporation into the tumour DNA was determined after i.v. administration of 3H-thymidine. An uncoupling of FDG transport and phosphorylation was found with enhanced K1 and k2 values and a normal k3 after 2 days of GCV treatment. The increase in FDG transport normalized after 4 days whereas the phosphorylation rate k3 increased. Quantification using the metabolic rate or the SUV showed congruent but less sensitive results compared with the modeling approach. The thymidine incorporation into the DNA of the tumours declined to 10.5% of the controls after 4 days of GCV treatment. The data indicate that PET with 18FDG and 11C-thymidine may be applied for monitoring of gene therapy with the HSVtk/GCV suicide system. Increased transport rates are evidence of stress reactions early after therapy. The measurement of thymidine incorporation into the tumour DNA can be used as an indicator of therapy efficacy.

Animals↗

PubMed1998

Functions encoded by pyrrolnitrin biosynthetic genes from Pseudomonas fluorescens.

Pyrrolnitrin is a secondary metabolite derived from tryptophan and has strong antifungal activity. Recently we described four genes, prnABCD, from Pseudomonas fluorescens that encode the biosynthesis of pyrrolnitrin. In the work presented here, we describe the function of each prn gene product. The four genes encode proteins identical in size and serology to proteins present in wild-type Pseudomonas fluorescens, but absent from a mutant from which the entire prn gene region had been deleted. The prnA gene product catalyzes the chlorination of L-tryptophan to form 7-chloro-L-tryptophan. The prnB gene product catalyzes a ring rearrangement and decarboxylation to convert 7-chloro-L-tryptophan to monodechloroaminopyrrolnitrin. The prnC gene product chlorinates monodechloroaminopyrrolnitrin at the 3 position to form aminopyrrolnitrin. The prnD gene product catalyzes the oxidation of the amino group of aminopyrrolnitrin to a nitro group to form pyrrolnitrin. The organization of the prn genes in the operon is identical to the order of the reactions in the biosynthetic pathway.

Blotting, Western↗

PubMed1997

Specificity of human cytotoxic T lymphocytes induced by a human papillomavirus type 16 E7-derived peptide.

In order to establish tumour-specific cytotoxic T lymphocyte (CTL) cell lines, T cells from a human papillomavirus (HPV) type 16-positive patient with a cervical carcinoma in situ and from a healthy volunteer were stimulated in vitro with autologous dendritic cells loaded with peptides derived from the viral transforming proteins E6 and E7 and corresponding to potential HLA-A*0201-restricted T cell epitopes. From each donor a small number of low-affinity CTL lines against the peptide E7/86-93 was obtained, which specifically lysed HLA-A*0201-expressing B-lymphocytes (cell line 721) loaded with this peptide. Cytotoxicity was also observed against two HLA-A*0201-E7-positive epithelial cell lines, the cervical carcinoma cell line CaSki and the HPV-16-immortalized foreskin-keratinocyte cell line HPK IA. However, since none of the CTL recognized both cell lines, and E7-expressing 721 transfectants were never lysed, it was concluded that the reactivity against CaSki and HPK IA cells was due to cross-reactivity on allogeneic HLA molecules rather than to E7 recognition, which emphasizes that the specificity of tumour cell lysis by peptide-induced CTL has to be interpreted with caution.

Carcinoma in Situ↗

PubMed1997

Monitoring gene therapy with herpes simplex virus thymidine kinase in hepatoma cells: uptake of specific substrates.

UNLABELLED: This study investigates the application of PET with specific substrates for the assessment of enzyme activity after transfer of the herpes simplex virus thymidine kinase (HSV-tk) gene. METHODS: After transfection of a rat hepatoma cell line with a retroviral vector containing the HSV-tk gene, different clones were established by G418 selection. Uptake measurements were performed up to 48 hr in a TK-expressing cell line and in a control cell line using thymidine (TdR; measured under therapy conditions), fluorodeoxycytidine (FdCyt) and ganciclovir (GCV). Additionally, bystander experiments and inhibition/competition studies were done. RESULTS: In TK-expressing cells GCV treatment caused an increased (up to 250%) TdR uptake in the acid-soluble fraction and a decrease to 5.5% in the acid-insoluble fraction. The FdCyt uptake was higher in the TK-expressing cells than in controls with a maximum after 4 hr (12-fold and 3-fold higher in the acid-insoluble and acid-soluble fraction). GCV accumulated up to 180-fold more in the acid-insoluble and 26-fold more in the acid-soluble fraction. GCV uptake occurred mainly by the nucleoside transport systems. Bystander experiments revealed a relation between growth inhibition or GCV uptake and the amount of TK-expressing cells. GCV uptake and growth inhibition were correlated with r = 0.96. CONCLUSION: Assessment of GCV accumulation may serve as an indicator of the enzyme activity and of therapy outcome. TdR may be useful to measure therapy effects on DNA synthesis, whereas the potential of FdCyt has to be investigated in further studies.

Animals↗

PubMed1997

Multitracer studies during gene therapy of hepatoma cells with herpes simplex virus thymidine kinase and ganciclovir.

UNLABELLED: Using different tracers of tumor metabolism, the application of PET for monitoring gene therapy with the suicide gene herpes simplex virus thymidine kinase (HSVtk) is investigated in this in vitro study. METHODS: Morris hepatoma cells were transfected with a retroviral vector bearing the HSVtk gene, and different clones were established by selection with the neomycin analog G418. Thereafter, uptake measurements using fluorodeoxyglucose (FDG), 3-O-methylglucose, aminoisobutyric acid and methionine were performed in a thymidine kinase (TK)-expressing cell line and in control cells bearing the empty vector in the presence of different concentrations of ganciclovir (GCV). These experiments were done up to 48 hr after the onset of therapy. The values were expressed as Bq/well or as Bq/10(5) cells. RESULTS: During GCV treatment therapy, a decrease of the uptake/well was measured for all tracers in the TK-expressing cell line. After normalization to the viable cell number, the uptake for FDG and 3-O-methylglucose increases up to 195% after 24 hr incubation with GCV. A high-pressure liquid chromatography analysis revealed a decline of the FDG-6-phosphate fraction after 48 hr incubation with GCV. Consequently, a normalization of FDG uptake was observed after this incubation period, whereas the 3-O-methylglucose uptake was still increased. Experiments performed with different amounts of TK-expressing cells and control cells showed that these effects are dependent on the percentage of TK-expressing cells. The aminoisobutyric acid uptake decreases to 47%, while the methionine uptake decreases in the acid-insoluble fraction (to 17%) and increases in the acid-soluble fraction (to 150%). CONCLUSION: These data indicate that combinations of the PET tracers used in these experiments may be applied for monitoring gene therapy with HSVtk. The increase in FDG and 3-O-methylglucose uptake in vitro is interpreted as stress reaction of the tumor cells. However, an uncoupling of transport and phosphorylation was observed after 48 hr incubation. The amino acid uptake experiments point to an inhibition of protein synthesis as well as of the neutral amino acid transport.

3-O-Methylglucose↗

PubMed1997

PET 2-fluoro-2-deoxyglucose uptake in rat prostate adenocarcinoma during chemotherapy with gemcitabine.

UNLABELLED: This study was performed to investigate the effect of the new chemotherapeutic agent gemcitabine on glucose transport and metabolism in prostate carcinoma in vitro and in vivo. METHODS: After transplantation of rat prostate adenocarcinoma cells, dynamic PET measurements with fluorine-18-labeled 2-fluoro-2-deoxy-D-glucose (18FDG) were performed in 15 animals before and 1 day after therapy with 90 mg/kg of body weight (n = 8) and 180 mg/kg of body weight (n = 7) gemcitabine. In the second examination, the animals received a simultaneous injection of 18FDG and [3H]thymidine. Quantitative evaluation of the PET data was done using the standardized uptake value (SUV) as well as a three-compartment pharmacokinetic model. Furthermore, the incorporation of [3H]thymidine into the DNA was determined. In vitro measurements of the FDG, 3-O-methylglucose and thymidine uptake were performed immediately and 4 hr after a 24-hr incubation period with different doses of gemcitabine. RESULTS: FDG-SUV and the metabolic rate of FD 3 utilization did not change significantly after therapy. However, the values for the transport rate constants K1 and K2 increased significantly. The incorporation of thymidine into the DNA of treated tumors showed an 80% decline as compared with a control group. In the cell culture experiments, a dose-dependent increase of FDG (up to 178%) and 3-O-methylglucose uptake (up to 305%) was demonstrated. The thymidine uptake showed a 96% decline in the nucleic acid fraction and an increase of up to 337% in the cytoplasmic fraction. CONCLUSION: The more global measures of FDG metabolism as SUV and metabolic rate of FDG utilization were unchanged after therapy, while DNA synthesis and cell viability declined. However, in vitro and in vivo evidence of an enhancement of glucose transport is presented, indicating that quantification by modelling may be superior for the evaluation of metabolic effects during chemotherapy.

Adenocarcinoma↗

PubMed1996

Human papillomavirus type 16 (HPV 16) E7 and major histocompatibility complex (MHC) class I and II expression in human keratinocytes in culture.

The low expression of major histocompatibility complex (MHC) class I antigens on human papillomavirus (HPV)-infected cervical carcinoma cells may be responsible for an insufficient cytotoxic T cell response against these cells. To investigate in vitro whether the HPV type 16 early gene product E7 influences cell surface expression of MHC class I and II molecules the HPV negative keratinocyte cell line HaCaT was either stably transfected with the E7 gene or infected with E7-recombinant vaccinia viruses. No difference in MHC class I transcription was detected between E7-transfected and untransfected HaCaT cells. MHC class I cell surface expression as determined by FACS analysis was stronger in some of the transfectants and less intensive in others when compared to untransfected HaCaT cells. In wildtype as well as in E7-recombinant vaccinia virus infected HaCaT cells downregulation of MHC class I molecules on protein and transcriptional level was observed. The alterations in MHC class I expression were independent of the presence and amount of E7-specific transcripts. None of the transfectants or infected HaCaT cells had MHC class II molecules on their cell surface. Hence, our data did not show a correlation between HPV 16 E7 and MHC expression in vitro.

Cell Line↗