Partial purification of a protein growth inhibitor from multicellular spheroids

Freyer, James P.; Schor, Patricia L.; Saponara, A.G.

doi:10.1016/s0006-291x(88)80736-8

Cited by 22 publications

(21 citation statements)

References 11 publications

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“…The high rates of OxPhos determined in young spheroids and monolayer culture cells (see Table 1) were similar to the total oxygen consumption rates documented in melanoma WIX‐c 600‐µm spheroids and monolayer cells (Hystad and Rofstad, 1994), and in Rat1‐T1 500‐µm spheroids (Wu et al, 1999). Furthermore, the marked diminution in OxPhos capacity in HeLa and Hek293 old‐spheroids in comparison to their respective young counterparts (see Table 1) agreed with the diminution (1.2–4.5 times) in cellular oxygen consumption described for large spheroids (diameter >800 µm) from tumor human glioma U‐118MG, human colon carcinoma HT29 (Carlsson and Acker, 1988), rat glioma C6 (Acker et al, 1992), rat transformed fibroblast Rat1‐T1 and MR1 (Kunz‐Schughart et al, 2000), mouse mammary tumor EMT6/Ro (Freyer, 1988b), and normal hamster fibroblast V‐79 (Freyer et al, 1984).…”

Section: Discussionsupporting

confidence: 76%

Novel paclitaxel‐eluting, biodegradable polymer coated stent in the treatment of de novo coronary lesions: A prospective multicenter registry

Buszman

Trznadel

Milewski

et al. 2007

Cathet Cardio Intervent

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Objectives: The purpose of the present study was to evaluate the efficacy and safety of a biodegradable polymer coated, paclitaxel eluting stent (Luc-Chopin 2 ) based on 9-months angiographic and 12-months clinical follow-up results. Background: First-generation drug-eluting stents utilize nonbioabsorbable polymeric coatings, whose persistent presence in the arterial wall may negatively affect long-term outcomes. Bioabsorbable coatings with a degradation period matched to that of the drug elution may be a better alternative, clinically and economically. Methods: We conducted a prospective, multicenter first-in-man registry of a novel, locally developed, bioabsorbable-coated, paclitaxel-eluting coronary stent in 116 patients with single-lesion de novo coronary disease. Results: Major adverse cardiac events occurred in 7.8% patients within 12 months. There were no late thrombotic events, death, stroke, or surgical revascularization in that period. There were two myocardial infarctions, one related to recent subacute stent thrombosis and another associated with restenosis. By 12 months, target vessel revascularization was performed in 7.8%; 2.9% were ischemia-driven and the rest were mandated at 9 months in accordance with a control angiography protocol. Core-lab assessed binary in-stent restenosis (≥50% DS) was noted in 11.9% patients and mean late loss was 0.46 6 0.47 mm. Conclusions: This first-in-man experience obtained in a multicenter registry of real-world de novo lesions (almost half of lesions were class B2 or C by AHA classification) showed a favorable safety profile and acceptable efficacy through 12 months. Randomized comparison with a benchmark nonbioabsorbable polymer coated paclitaxel eluting stent should be undertaken to validate this initial positive experience. '

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Section: Discussionsupporting

confidence: 76%

Novel paclitaxel‐eluting, biodegradable polymer coated stent in the treatment of de novo coronary lesions: A prospective multicenter registry

Buszman

Trznadel

Milewski

et al. 2007

Cathet Cardio Intervent

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“…Furthermore, the marked diminution in OxPhos capacity in HeLa and Hek293 old-spheroids in comparison to their respective young counterparts (see Table 1) agreed with the diminution (1.2-4.5 times) in cellular oxygen consumption described for large spheroids (diameter >800 mm) from tumor human glioma U-118MG, human colon carcinoma HT29 (Carlsson and Acker, 1988), rat glioma C6 (Acker et al, 1992), rat transformed fibroblast Rat1-T1 and MR1 , mouse mammary tumor EMT6/Ro (Freyer, 1988b), and normal hamster fibroblast V-79 (Freyer et al, 1984).…”

Section: Is There a Diminished Mitochondrial Function During Mcts Grosupporting

confidence: 73%

“…However in these last studies and others, where oxygen consumption has been analyzed (Freyer et al, 1984;Carlsson and Acker, 1988;Freyer, 1988b;Acker et al, 1992;Hystad and Rofstad, 1994;Kunz-Schughart et al, 2000), the oligomycinsensitive respiration was not assessed. Therefore, due to the high non-mitochondrial oxidase activities, especially heme oxygenase (Murphy et al, 1993), and cytochrome P 450 activity Enzyme activities were determined as described in the Materials and Methods Section.…”

Section: Is There a Diminished Mitochondrial Function During Mcts Gromentioning

confidence: 99%

Energy metabolism transition in multi‐cellular human tumor spheroids

Rodrı́guez-Enrı́quez

Gallardo‐Pérez

Avilés‐Salas

et al. 2008

Journal Cellular Physiology

124

101

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It is thought that glycolysis is the predominant energy pathway in cancer, particularly in solid and poorly vascularized tumors where hypoxic regions develop. To evaluate whether glycolysis does effectively predominate for ATP supply and to identify the underlying biochemical mechanisms, the glycolytic and oxidative phosphorylation (OxPhos) fluxes, ATP/ADP ratio, phosphorylation potential, and expression and activity of relevant energy metabolism enzymes were determined in multi-cellular tumor spheroids, as a model of human solid tumors. In HeLa and Hek293 young-spheroids, the OxPhos flux and cytochrome c oxidase protein content and activity were similar to those observed in monolayer cultured cells, whereas the glycolytic flux increased two- to fourfold; the contribution of OxPhos to ATP supply was 60%. In contrast, in old-spheroids, OxPhos, ATP content, ATP/ADP ratio, and phosphorylation potential diminished 50-70%, as well as the activity (88%) and content (3 times) of cytochrome c oxidase. Glycolysis and hexokinase increased significantly (both, 4 times); consequently glycolysis was the predominant pathway for ATP supply (80%). These changes were associated with an increase (3.3 times) in the HIF-1alpha content. After chronic exposure, both oxidative and glycolytic inhibitors blocked spheroid growth, although the glycolytic inhibitors, 2-deoxyglucose and gossypol (IC(50) of 15-17 nM), were more potent than the mitochondrial inhibitors, casiopeina II-gly, laherradurin, and rhodamine 123 (IC(50) > 100 nM). These results suggest that glycolysis and OxPhos might be considered as metabolic targets to diminish cellular proliferation in poorly vascularized, hypoxic solid tumors.

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“…There, because of unknown factors, the cell will be unable to replicate its DNA and arrest, or will pass into G 2 phase and arrest. In both cell types, the S-and G 2 -phase arrest may be due to specific alterations in S-and G 2 -phase regulatory proteins, or may reflect the effect of a substance released from the spheroid necrotic center that nonspecifically induces arrest in all phases of the cell cycle (6,41). S-phase arrest has been observed in cells from tumors, and is indicative of extreme physiological conditions (37,42).…”

Section: Discussionmentioning

confidence: 99%

Microenvironmental Regulation of Proliferation in Multicellular Spheroids Is Mediated through Differential Expression of Cyclin-Dependent Kinase Inhibitors

2004

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Multicellular spheroids composed of transformed cells are known to mimic the growth characteristics of tumors and to develop gradients in proliferation with increasing size. This progressive accumulation of quiescent cells is presumably an active process that occurs in response to the microenvironmental stresses that develop within the three-dimensional structure, and, yet, little is known regarding either the signals that induce the cell cycle arrest or the molecular basis for the halt in proliferation. We have previously reported that regulation of cyclin-dependent kinase (CDK) inhibitors (CKIs) differs in monolayer versus spheroid cell culture. In this study, we have examined the expression of three CKIs in EMT6 mouse mammary carcinoma and MEL28 human melanoma spheroids, as a function both of spheroid size and of location within the spheroid. We report that expression of the CKIs p18INK4c , p21 waf1/cip1 , and p27 Kip1 all increase as the spheroid grows and develops a quiescent cell fraction. However, by examining protein expression in discrete regions of the spheroid, we have found that only p18INK4c and p27 Kip1 expression positively correlate with growth arrest, whereas p21 waf1/cip1 is expressed predominantly in proliferating cells. Further analysis indicated that, in the quiescent cells, p18INK4c is found in increasing association with CDK6, whereas p27Kip1 associates predominantly with CDK2. In MEL28 cells, CDK2 activity is completely abrogated in the inner regions of the spheroid, whereas in EMT6 cells, CDK2 activity decreases in accordance with a decrease in expression. We also observed a decrease in all cell cycle regulatory proteins in the innermost spheroid fraction, including CDKs, CKIs, and cyclins. Induction of CKIs from separate families, as well as their association with distinct target CDKs, suggests that there may be multiple checkpoints activated to ensure cell cycle arrest in non-growthconducive environments. Furthermore, because very similar observations were made in both a human melanoma cell line and a mouse mammary carcinoma cell line, our results indicate that these checkpoints, as well as the signal transduction pathways that activate them, are highly conserved.

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Partial purification of a protein growth inhibitor from multicellular spheroids

Cited by 22 publications

References 11 publications

Novel paclitaxel‐eluting, biodegradable polymer coated stent in the treatment of de novo coronary lesions: A prospective multicenter registry

Novel paclitaxel‐eluting, biodegradable polymer coated stent in the treatment of de novo coronary lesions: A prospective multicenter registry

Energy metabolism transition in multi‐cellular human tumor spheroids

Microenvironmental Regulation of Proliferation in Multicellular Spheroids Is Mediated through Differential Expression of Cyclin-Dependent Kinase Inhibitors

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