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Sarcoma Molecular Biology Laboratory
Recent Publications
Ecteinascidin-743
drug resistance in sarcoma cells: transcriptional and cellular alterations
Shao L, Kasanov J, Hornicek FJ, Morii T, Fondren G, Weissbach L.
Biochem Pharmacol. 2003 Dec 15;66(12):2381-95
Orthopaedic Research Laboratories, Massachusetts General Hospital and Harvard Medical School, GRJ 1124, 55 Fruit Street, Boston, MA 02114, USA.
A human chondrosarcoma cell line, CS-1, was treated successively with increasing concentrations of the marine chemotherapeutic Ecteinascidin-743 (ET-743), yielding a variant cell line displaying a significant degree of resistance to the cytotoxic action of this drug. Various experiments were performed to discern molecular aberrations between the parent and resistant cell line, and also identify potential molecular markers indicative of drug resistance. Although no significant differences in the levels of membrane transporters such as P-glycoprotein or multidrug resistance protein 1 (MRP1) were detected, the cell migratory ability of the ET-743-resistant cell variant was reduced, as was its attachment capability to gelatin-coated cell culture dishes. Staining of the actin-containing cytoskeleton with fluorescent-labeled phalloidin revealed marked differences in the cytoskeleton architecture between the parent and ET-743-resistant CS-1 cell lines. Comparison of serum-free conditioned medium from both cell lines showed conspicuous differences in the levels of several proteins, including a quartet of high molecular weight proteins (> or =140 kDa). The protein sequences of two of these high molecular weight proteins, present at significantly higher concentrations in conditioned medium obtained from the parent cell line, corresponded to subunits of types I and IV collagen. Analysis of type I collagen alpha1 chain mRNA revealed a significantly lower level in the ET-743-resistant CS-1 cell line. Thus, prolonged exposure to ET-743 may cause changes in cell function through cytoskeleton rearrangement and/or modulation of collagen levels.
Sphingosine
1-phosphate and cell migration: resistance to angiogenesis inhibitors
Morii T, Weissbach L.
Exp Cell Res. 2003 Jul 1;287(1):166-77.
Orthopaedic Research Laboratories, Massachusetts General Hospital and Harvard Medical School, Boston, Massachusetts 02114, USA.
Naturally occurring angiogenesis inhibitors can inhibit different steps of the angiogenic process, such as endothelial cell migration. However, the mechanisms underlying this inhibition have not been elucidated. We demonstrate that migration of human umbilical vein endothelial cells induced by the potent endothelial cell chemoattractant sphingosine 1-phosphate is refractory to inhibition by well-characterized angiogenesis inhibitors such as endostatin and plasminogen-related protein-B. Our data support the contention that for effective blockage of tumor-induced angiogenesis, antagonists of both G protein-coupled receptor signaling and receptor tyrosine kinase signaling must be combined.
Interaction
of plasminogen-related protein B with endothelial and smooth muscle cells
in vitro
Morioka H, Morii T, Vogel T, Hornicek FJ, Weissbach L.
Orthopaedic Research Laboratories, Massachusetts General Hospital and Harvard Medical School, GRJ 1124, 55 Fruit Street, Boston, MA 02114, USA.
Plasminogen-related protein B (PRP-B) closely resembles the N-terminal plasminogen activation peptide, which is released from plasminogen during conversion to plasmin. We have previously demonstrated that the steady-state level of mRNA encoding PRP-B is increased within tumor tissues, and that recombinant PRP-B antagonizes neoplastic growth when administered systemically to mice harboring tumors, but no insights into the cell targets of PRP-B have been presented. Employing serum-free medium optimized for culturing human endothelial or smooth muscle cells, we show that recombinant PRP-B inhibits basic fibroblast growth factor-dependent cell migration for both cell types, as well as tube formation of endothelial cells. Comparison with the angiogenesis inhibitors angiostatin and endostatin revealed similar results. Recombinant PRP-B is effective in promoting cell attachment of endothelial and smooth muscle cells, and antibody interference experiments reveal that the interaction of recombinant PRP-B with endothelial cells is mediated at least in part by alpha(v)-containing integrins. Inhibition of angiogenesis in vivo by PRP-B was demonstrated in the chicken chorioallantoic membrane assay. PRP-B and other antiangiogenic molecules may elicit metabolic perturbations in endothelial cells as well as perivascular mesenchymal cells such as smooth muscle cells and pericytes.
Antiangiogenesis
treatment combined with chemotherapy produces chondrosarcoma necrosis
Morioka H, Weissbach L, Vogel T, Nielsen GP, Faircloth GT, Shao L, Hornicek
FJ.
Orthopedic Research Laboratories, Massachusetts General Hospital and Harvard Medical School, 55 Fruit Street, Boston, MA 02114, USA.
A combination therapy protocol using a marine chemotherapeutic and an antiangiogenic molecule was tested in a mouse tumor xenograft model for the ability to curtail the growth of a human chondrosarcoma (CHSA). Ecteinascidin-743 (ET-743), a marine-derived chemotherapeutic, was effective at slowing the growth of a primary CHSA. Plasminogen-related protein B, which antagonizes various endothelial cell activities, also elicited a significant inhibition of neoplastic growth, albeit with reduced effectiveness. The combination of the two agents resulted in only a modest further repression of tumor growth over that associated with ET-743 treatment alone, as measured by tumor volume (82% versus 76% inhibition, respectively). However, analysis of the extent of tumor necrosis and vascularization of the tumor revealed that the coadministration of the two compounds was clearly more effective, eliciting a 2.5-fold increase in tumor necrosis relative to single-agent treatment. The combination therapy also was most effective at antagonizing tumor-associated microvessel formation, as assessed by CD31 immunostaining, suggesting that combination therapy may hold promise for treating CHSA. Tumor necrosis produced by combination therapy of ET-743 and recombinant plasminogen-related protein B was also significantly greater than that produced by conventional doxorubicin treatment, further corroborating the efficacy of combination therapy.
