Reversible depression of transcription during hibernation.

Mammalian hibernators downregulate processes of energy production and consumption while maintaining cellular homeostasis. Energetic costs of transcription must be balanced with demands for gene products. Data from nuclear run-on assays indicate transcriptional initiation is reduced two fold in torpid golden-mantled ground squirrels ( Spermophilus lateralis) as compared to euthermic animals between bouts of torpor. In addition, elongation rates across the temperature range experienced by hibernators indicate a virtual arrest of transcription at the low body temperatures of torpor. Finally, there is no seasonal compensation or species-specific adaptation for increased elongational capacity in the cold. Thus, it appears that hibernators are not specifically adapted to continue transcription during torpor. Taken together, these data indicate that transcription arrests during torpor because of a moderate depression of initiation and a more severe inhibition of elongation, largely due to temperature effects. Restoration of euthermic body temperatures during the interbout arousals reverses this transcriptional depression and permits gene expression.

Ubiquitin conjugate dynamics in the gut and liver of hibernating ground squirrels.

Protein synthesis is severely depressed in hibernating mammals. In the absence of significant protein synthesis, the continued turnover of proteins as a function of normal cellular activity would result in the net depletion of protein pools. We measured levels of ubiquitylated proteins in the gut of thirteen-lined ground squirrels ( Spermophilus tridecemlineatus) and liver of golden-mantled ground squirrels ( Spermophilus lateralis). In both tissues, ubiquitin conjugate concentrations increased during entrance into torpor and were elevated 2-3 fold by late torpor compared with levels in active animals. The data are consistent with a depression of proteolysis with a resultant high level of ubiquitylated proteins during the natural hypothermia of torpor. The periodic returns to euthermy during the hibernation season allow for degradation of these conjugated proteins and may serve to restore protein pools.

Translational initiation is uncoupled from elongation at 18 degrees C during mammalian hibernation.

Cellular and organismal homeostasis must be maintained across a body temperature (Tb) range of 0 to 37 degrees C during mammalian hibernation. Hibernators depress biosynthetic activities including protein synthesis, concordant with limited energy availability and temperature effects on reaction rates. We used polysome analysis to show that initiation of protein synthesis ceases during entrance into torpor in golden-mantled ground squirrels (Spermophilus lateralis) when Tb reaches 18 degrees C. Elongation of preinitiated polypeptides continues slowly throughout the torpor bout. As Tb begins to rise, initiation resumes even at temperatures below 18 degrees C, although the euthermic polysome pattern is not reestablished. At precisely 18 degrees C, there is a large increase in initiation events and a complete restoration of euthermic polysome distribution patterns. These data indicate a role for both passive and active depression of translation during torpor and are consistent with a requirement for new protein biosynthesis during each interbout arousal.

Characterization of ATP-dependent proteolysis in embryos of the brine shrimp, Artemia franciscana.

Under anoxia, embryos of Artemia franciscana enter a state of quiescence. During this time protein synthesis is depressed, and continued degradation of proteins could jeopardize the ability to recover from quiescence upon return to favorable conditions. In this study, we developed an assay for monitoring ATP/ ubiquitin-dependent proteolysis in order to establish the presence of this degradation mechanism in A. franciscana embryos, and to describe some characteristics that may regulate its function during anoxia-induced quiescence. For lysates experimentally depleted of adenylates, supplementation with ATP and ubiquitin stimulated protein degradation rates by 92 +/- 17% (mean +/- SE) compared to control rates. The stimulation by ATP was maximal at concentrations > or =11 micromol x l(-1). In the presence of ATP and ubiquitin, ubiquitin-conjugated proteins were produced by lysates during the course of the 4-h assays, as detected by Western blotting. Acute acidification of lysates to values approximating the intracellular pH observed under anoxia completely inhibited ATP/ubiquitin-dependent proteolysis. Depressed degradation was also observed under conditions where net ATP hydrolysis occurred. These results suggest that ATP/ubiquitin-dependent proteolysis is markedly inhibited under cellular conditions promoted by anoxia. Inhibition of proteolysis during quiescence may be one critical factor that increases macromolecular stability, which may ultimately govern the duration of embryo survival under anoxia.

Depression of nuclear transcription and extension of mRNA half-life under anoxia in Artemia franciscana embryos.

Transcriptional activity, as assessed by nuclear run-on assays, was constant during 10 h of normoxic development for embryos of the brine shrimp Artemia franciscana. Exposure of embryos to only 4 h of anoxia resulted in a 79.3+/-1 % decrease in levels of in-vivo-initiated transcripts, and transcription was depressed by 88. 2+/-0.7 % compared with normoxic controls after 24 h of anoxia (means +/- s.e.m., N=3). Initiation of transcription was fully restored after 1 h of normoxic recovery. Artificially lowering the intracellular pH of aerobic embryos to the value reflective of anoxia (pH 6.7) showed that acidification alone explained over half the transcriptional arrest. Initiation of transcription was not rescued by application of 80 % carbon monoxide under anoxia, which suggests that heme-based oxygen sensing is not involved in this global arrest. When these transcriptional data are combined with the finding that mRNA levels are unchanged for at least 6 h of anoxia, it is clear that the half-life of mRNA is extended at least 8.5-fold compared with that in aerobic embryos. In contrast to the activation of compensatory mechanisms to cope with anoxia that occurs in mammalian cells, A. franciscana embryos enter a metabolically depressed state in which gene expression and mRNA turnover are cellular costs apparently not compatible with survival and in which extended tolerance supercedes the requirement for continued metabolic function.

Paclitaxel for malignant pleural mesothelioma: a phase II study of the EORTC Lung Cancer Cooperative Group.

The EORTC Lung Cancer Cooperative Group undertook a phase II study of paclitaxel in 25 chemotherapy-naive patients with malignant pleural mesothelioma. Paclitaxel was given intravenously at a dose of 200 mg m-2 as a 3 h infusion every 3 weeks, after standard premedication with corticosteroids and antihistamines. This regimen was well tolerated, with < 4% of cycles resulting in severe toxicity. No major objective responses were observed and ten patients had stable disease. Median survival time was 39 weeks and the 1 year survival rate was 30%. In conclusion, paclitaxel at the dose and schedule investigated in this trial had no major activity in the treatment of malignant pleural mesothelioma.

Epirubicin in malignant mesothelioma: a phase II study of the European Organization for Research and Treatment of Cancer Lung Cancer Cooperative Group.

PURPOSE: The doxorubicin analog, epirubicin (EPI), was tested in patients with malignant mesothelioma. PATIENTS AND METHODS: Sixty-three patients with malignant mesothelioma were given EPI 110 mg/m2 every 3 weeks. Histology was reviewed and confirmed by a pathology panel. On the basis of unconvincing or wrong histology, insufficient material or cytology only, nine cases were considered ineligible for the study. None of the patients had received prior chemotherapy. RESULTS: The main side effects were myelosuppression, alopecia, and gastrointestinal toxicity. Tumor response, assessed by computed tomographic (CT) scans, was assessable in 48 patients. Seven patients (15%) achieved a partial response that lasted a median of 37 weeks; 19 patients had stable disease, and 22 patients progressed on treatment. Median survival time was 40 weeks from the start of chemotherapy, and the median survival of responding patients was 87 weeks. One responding patient is still alive and free of relapse 4 years from the start of chemotherapy. CONCLUSION: We conclude that further testing of EPI in malignant mesothelioma is warranted.

Disuse atrophy in the hibernating golden-mantled ground squirrel, Spermophilus lateralis.

Disuse (inactivity, bed rest, and spaceflight) may lead to a loss of muscle mass and a decrease in oxidative capacity in skeletal muscle. If such changes were to occur in hibernating animals, both locomotor and thermogenic function would be compromised. Muscle masses and oxidative capacities (as assessed by citrate synthase activity) were measured in the gastrocnemius and semitendinosus muscles, cardiac muscle (ventricle), and brown fat (axillary pad) in a group (n = 7) of prehibernating ground squirrels (Spermophilus lateralis) and after 6 mo of hibernation (n = 8). Hibernation produced significant atrophy in the gastrocnemius (14%) and semitendinosus (42%) muscles. Cardiac tissue increased (21%) in mass, as did brown adipose tissue (150%). That such changes were not due simply to fluid shifts was evidenced by similar protein concentrations between groups. In contrast to many other disuse studies, oxidative capacity was increased significantly in the gastrocnemius (65%) and semitendinosus (37%). Citrate synthase was also higher in cardiac tissue of hibernators (20%) but was not significantly different in brown fat.

Mitoxantrone in malignant pleural mesothelioma: a study by the EORTC Lung Cancer Cooperative Group.

46 patients with malignant pleural mesothelioma were entered in a phase II study of mitoxantrone 14 mg/m2 every 3 weeks. Histology was confirmed by a pathology panel. None of the patients had received previous chemotherapy. Toxicity was mainly mild gastrointestinal and haematological side-effects. Out of 34 patients evaluated for response, only 1 partial response was recorded. Mitoxantrone at this dose and schedule has marginal activity in malignant mesothelioma.

Vindesine, cisplatin, and bleomycin combination chemotherapy in non-small cell lung cancer: survival and quality of life.

Although combination chemotherapy is applied on a large scale in advanced non-small cell lung cancer (NSCLC), we still lack evidence indicating in which subsets of patients survival or quality of life might be improved. We studied these issues among a sample of 28 NSCLC patients with a high performance status, who received a tri-weekly vindesine, cisplatin, and bleomycin combination. Treatment was extended for an additional two courses only if a response was observed after the initial three courses. An overall response rate of 13/27 evaluable patients (48%) was found (complete response 1/27 and partial response 12/27) with a median response duration of 24 weeks. Median survival was 33 weeks (47 for responders and 26 for non-responders). Toxicity was primarily related to cisplatin, including severe nausea and vomiting and nephrotoxicity in 68% and 21% of the patients, respectively. Performance status and body weight dropped significantly during chemotherapy both among responders and non-responders. Performance status after discontinuation of chemotherapy approached pre-treatment scores in responders only. While the antitumor effect of this drug combination was confirmed, we conclude that treatment-associated toxicity and deterioration of the patients' well-being offset any potential survival advantage for the majority of patients.

Efficacy of amino-hydroxypropylidene bisphosphonate in hypercalcemia: observations on regulation of serum calcium.

For 2 weeks 27 patients with hypercalcemia received a standard oral treatment with (3-amino-1-hydroxypropylidene)-1,1-bisphosphonate (APD) as the sole agent. Results were grouped according to causes of hypercalcemia and compared with effects of APD in 13 normocalcemic patients with Paget's disease of bone and 7 with osteoporosis. In 12 hypercalcemic patients with osteolytic bone lesions and in the 20 normocalcemic patients, the mean serum calcium decreased to final levels that were subnormal and significantly lower than those obtained after treatment of 8 patients with primary hyperparathyroidism. In 3 patients with myeloma and in 4 tumor patients without bone lesions, serum calcium did not always decrease to the normal range. Implications of these observations for the mechanism of hypercalcemia are discussed.

Inhibition of osteolytic bone lesions by (3-amino-1-hydroxypropylidene)-1, 1-bisphosphonate (A.P.D.).

14 patients with osteolytic bone disease due to breast cancer or myeloma, 7 of whom had hypercalcaemia, received oral treatment with (3-amino-1-hydroxypropylidene)-1, 1-bisphosphonate (A.P.D.). Serum-calcium dropped to low normal values in all 14 patients, accompanied by a decrease in urine calcium and hydroxyproline excretion-rate. The results show that A.P.D. may inhibit tumour-induced osteolysis.