Polypeptide serving as angiogenic marker and dna thereof

a polypeptide and marker technology, applied in the field of polypeptides and polynucleotides, can solve the problems that the pathogenesis mechanism has not been perfectly elucidated, and achieve the effect of increasing the expression of kiaa1036 polypeptides

Inactive Publication Date: 2005-07-21
SATO YASUFUMI +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Benefits of technology

[0005] The present inventors found that expression of KIAA1036 polypeptide was increased in ovarian cancer and colon cancer and confirmed that the peptide was useful as a marker for cancer. As results of further researches, they found that KIAA1036 polypeptide was a new marker for neo...

Problems solved by technology

Though the research of genes relating to the above-mentioned various diseases have been conducted, the mechanism ...

Method used

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  • Polypeptide serving as angiogenic marker and dna thereof
  • Polypeptide serving as angiogenic marker and dna thereof
  • Polypeptide serving as angiogenic marker and dna thereof

Examples

Experimental program
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Effect test

example 1

An Identification of Gene Whose Expression in Human Umbilical Vein Endothelial Cells (HUVEC) is Enhanced by the Stimulation of Vascular Endothelial Cell Propagation Factors (VEGF)

[0228] Three Endocell HUVEC (KURABO) which is a human umbilical vein vascular endothelial cell were thawed and then inoculated on 16 culture plates coating with collagen (IWAKI) poured EBM medium including 10% fetal calf serum (FCS) (Sanko Junyaku). After culturing at 37° C. for 3 days, the medium was changed with the same medium and cultured for 2 days. After the cells reached to confluence, the medium was changed with M199 medium including 1% FCS (Nissui) and cells were cultured for 24 hours. For a culture with VEGF treatment, the medium was changed to M199 medium including 1% FCS and 1 nM human VEGF165 (R&D Systems). For a culture without VEGF (a control), medium was changed with the same medium not including human VEGF165. Cells were cultured at 37° C. and collected after 0 hour, 0.5 hours, 2 hours, 6...

example 2

Expression of KIAA1036 in Human Normal Tissues

[0230] Cloning of KIAA1036 gene from human umbilical vein vascular endothelial cell was tried. Two kinds of oligonucleotides were synthesized as primers for PCR on the basis of a nucleic acid sequence of 3′ untranslated region of KIAA1036 gene from NCBI database.

Primer1:5′-GTTCAGGACTGTCTTTCAGC-3′(SEQ ID NO: 4)Primer2:5′-GTCAATACTGATGGACTTGC-3′(SEQ ID NO: 5)

[0231] Total RNA was prepared according to an attached manual with RNeasy Midi (QIAGEN) from HUVEC which are human umbilical vein vascular endothelial cells. Single strand cDNA fragments were prepared from the obtained total RNA (10 μg) with SuperscriptII (Invitrogen). The obtained single strand cDNAs were used as templates and gene fragments were amplified with Primer1 (SEQ ID NO: 4) and Primer2 (SEQ ID NO: 5) by PCR.

[0232] To one fiftieth of single strand cDNAs, was added TaKaRa Ex Taq (TaKaRa) (1.25 unit), Primer1 and Primer2 (1 μM each), a buffer (Mg2+ plus) for PCR reaction a...

example 3

Expression of KIAA1036 in Human Umbilical Vein Endothelial Cells

[0235] As the VEGF-induced expression of KIAA1036 was originally found by Gene Expression Microarray Analysis using DNA chip as described in Example 1, the result was confirmed by Northern blot with KIAA1036 probe and total RNA prepared from VEGF-treated HUVEC. Endocell HUVEC (KURABO) which are human umbilical vein endothelial cells were cultured in M199 medium containing 1 nM human VEGF165 (R&D Systems) and 1% FCS as described in Example 1. After 0, 0.5, 1, 2, 6, 12 and 24 hours of culture, total RNA was prepared from these cells according to an attached manual with RNeasy Midi (QIAGEN).

[0236] After electrophoresis of each RNA sample (5 μg), the samples were transferred to a nylon membrane, Hybond-XL (Amersham Pharmacia Bioteque). That nylon membrane was immersed with GMC buffer and prehybridization was held at 65° C. for 1 hour. After prehybridization, the nylon membrane was immersed with GMC buffer containing prob...

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Abstract

A marker for neovascularization, vascular disease, inflammatory disease, entoptic neovascular disease, reproductive system disease, central nervous system disease and cancer, the method of detection of the marker and a diagnosis kit of the diseases are provided. Additionally, therapeutic agents of the diseases are provided. The expression of KIAA1036 is enhanced in ovarian cancer and large bowel cancer and KIAA1036 expresses in umbilical vein endothelial cell and inhibits DNA synthesis in the cells, cell migrating and lumen formation. Therefore KIAA1036 is useful as a marker for neovascularization, vascular disease, inflammatory disease, entoptic neovascular disease, reproductive system disease, central nervous system disease or cancer. Additionally, KIAA1036 is useful for screening of agonists, antagonists, DNA synthesis inhibitors, cell migrating inhibitors and neovascular inhibitors. The substances obtained by the screening, KIAA1036 and the antibodies are useful as therapeutic agents the above disease.

Description

TECHNICAL FIELD [0001] The present invention relates to a polypeptide and a polynucleotide useful as a marker for neovascularization, vascular disease, inflammatory disease, entoptic neovascular disease, reproductive system disease, central nervous system disease or cancer. And the present invention relates to the treatment of those diseases. BACKGROUND ART [0002] By a double adapter method, shotgun library of human genome was constructed and a nucleotide sequence of AF055021 (SEQ ID NO: 3), EST clone, was determined with the DNA library (ANLITICAL BIOCHEMISTRY, 236, 107-113 (1996).). Subsequently, a nucleotide sequence of full-length cDNA of AF055021 (SEQ ID NO: 1) was determined and named KIAA1036 (DNA Research, 6 (3) 197-205 (1999).). KIAA1036 polypeptide (SEQ ID NO: 2) is a polypeptide consisting of 365 amino acids and the function has not been elucidated yet. [0003] Since cells need oxygen and nutriments provided from blood to survive in vivo, neovascularization is vigorous in ...

Claims

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Application Information

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IPC IPC(8): A61K38/00A61P9/00A61P15/00A61P27/02A61P29/00A61P35/00A61P43/00C07K14/47C12N15/12C12Q1/68
CPCA61K38/00C07K14/47C12Q2600/158C12Q1/6883C12N2799/026A61P9/00A61P15/00A61P25/00A61P27/02A61P29/00A61P35/00A61P43/00G01N33/53
Inventor SATO, YASUFUMISONODA, HIKARUOHTA, HIDEKI
Owner SATO YASUFUMI
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