Pharmaceutical compositions and methods useful for modulating angiogenesis and inhibiting metastasis and tumor fibrosis
a technology of angiogenesis and compositions, applied in the direction of drug compositions, immunoglobulins against animals/humans, cardiovascular disorders, etc., can solve the problems of inability to use unmodified oligonucleotides, interference with correct splicing, and limited solid tumor growth
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The role of LOR1 in Angiogenesis
[0173] A study was conducted in efforts to further substantiate and characterize the role of LOR-1 in angiogenes is.
[0174] Materials and Methods
[0175] Human recombinant platelet factor-4 (PF4, GenBank Accession number M20901) which was produced in bacteria and subsequently refolded was supplied by Dr. Maione of Repligen Corp. (Boston, USA). Estrogen slow release pellets were obtained from Innovative Research of America, Sarasota, Fla., USA.
[0176] Construction of LOR-1 expression vector, transfection into MCF-7 cells, and expression: The LOR-1 cDNA (SEQ ID NO: 1) was cloned into a pCDNA3.1-hygro expression vector (Invitrogen Inc., USA) under the control of a CMV promoter. Clones of cells expressing LOR-1 were selected using hygromycine and assayed for LOR-1 expression using the polyclonal antisera described below.
[0177] Construction of platelet factor-4 affinity columns and purification of LOR-1 on such columns: PF4 was coupled to sepharose using a mod...
example 2
MCF-7 Breast Cancer Cells Expressing Recombinant Lysyl Oxidase Related Protein-1 (LOR-1) form Invasive Tumors Characterized by Extensive Fibrosis
[0200] A study was conducted in efforts to further substantiate and characterize the role of LOR-1 in inhibiting metastasis and tumor fibrosis.
[0201] Materials and Methods
[0202] Estrogen pellets (17.beta.-estradiol, 0.72 mg / pellet, 60-days release) were from Innovative Research of America, FL, USA. Masson Trichrome stain kit was purchased from Bio-Optica (Milano, Italy), Reverse Transcriptase, G418 were from GIBCO BRL (U.K.), Hygromycin B, Tetracycline hydrochloride, Reticulum stain kit fast green and Sirius red (direct red 80) were from Sigma (USA) , Restriction enzymes, T4 ligase were from New England Biolabs (USA). The bacterial expression vector pQE-30 and the nickel affinity column were obtained from Qiagen (Germany) , .sup.32p-dATP was purchased from NEN (USA). Monoclonal anti cytokeratin-7 (CAM 5.2) coupled to FITC was acquired from ...
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