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Nucleic acids, polypeptides, and methods for modulating apoptosis

A technology of apoptosis and cells, applied in the direction of chemical instruments and methods, biochemical equipment and methods, apoptosis-related proteins, etc., can solve the problem of not completely blocking apoptosis

Inactive Publication Date: 2006-02-08
SENESCO TECHNOLOGIES INC
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, almost all chemotherapeutic agents induce p53-independent apoptosis if dosed sufficiently, suggesting that even in drug-resistant tumors, there is no complete blockade of the apoptotic pathway

Method used

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  • Nucleic acids, polypeptides, and methods for modulating apoptosis
  • Nucleic acids, polypeptides, and methods for modulating apoptosis
  • Nucleic acids, polypeptides, and methods for modulating apoptosis

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0148] This example demonstrates the isolation and characterization of a full-length cDNA encoding a rat eIF-5A nucleic acid exhibiting apoptosis-specific expression.

[0149] Induction of superovulation and apoptosis in rat corpus luteum

[0150] Immature (21-30-day-old ) female rats with superovulation. Seven days after treatment with HCG, luteal cell apoptosis was induced by subcutaneous injection of 500 mg of PGF-2α. Rats are sacrificed at various times (eg, 1, 8, and 24 hours) after PGF-2α treatment, and the corpus luteum is removed and placed in liquid nitrogen. Rats were sacrificed immediately before PGF-2α treatment to obtain control corpus luteum tissue.

[0151] Dispersion of luteal cells in rat ovary

[0152] 6-9 days after superovulation, rats were treated by subcutaneous injection of 500 mg PGF-2α at multiple sites. After 15-30 minutes, ovaries were removed from superovulated rats, placed in EBSS (Gibco) on ice, blotted dry, and weighed. Cut off the connecti...

Embodiment 2

[0187] This example demonstrates modulation of apoptosis with apoptosis-specific eIF-5A and DHS.

[0188] Culture of COS-7 cells and isolation of RNA

[0189] COS-7 (a Vero monkey kidney fibroblast-like cell line transformed with an SV40 mutant encoding wild-type T antigen) was used for all transfection-based experiments. COS-7 cells were cultured in Dulbecco's modified Eagle's medium (DMEM) containing 0.584 g / LL-glutamine, 4.5 g / L glucose and 0.37% sodium bicarbonate. The medium was supplemented with 10% fetal bovine serum (FBS) and 100 units penicillin / streptomycin. at 5% CO 2 Cells were cultured at 37°C in a humidified environment with 95% air. Cells were subcultured every 3 to 4 days by detaching attached cells with a solution of 0.25% trypsin and 1 mM EDTA. Disperse the detached cells in a new Petri dish containing fresh medium at a dissociation ratio of 1:10.

[0190] In 150-mm tissue culture-treated dishes (Corning), COS-7 cells to be used for RNA isolation were cu...

Embodiment 3

[0208] This example demonstrates modulation of apoptosis with apoptosis-specific eIF-5A and DHS.

[0209] Using the general procedure and method described in the previous examples, Figure 23 is a flow chart illustrating the process of transient transfection of COS-7 cells, wherein the cells in serum-free medium were incubated with plasmid DNA (in lipofectAMINE) for 4 hours, serum was added, and the cells were incubated for an additional 40 hours. Cells were then incubated in normal medium containing serum for an additional 48 h (i.e., no further treatment) prior to analysis, serum was removed for 48 h prior to analysis to induce apoptosis, or treated with actinomycin D prior to analysis Cells were incubated for 48 hours to induce apoptosis.

[0210] Figure 22 is a Western blot illustrating the transient expression of foreign proteins in COS-7 cells following transfection with pHM6. After sham transfection, pHM6-LacZ, pHM6-antisense 3′rF5A (pHM6-antisense 3′UTR rat apoptosis ...

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Abstract

The present invention provides methods of identifying an incidence of ischemia in mammalian tissue, particularly mammalian heart tissue. Further, a method of reducing apoptosis in mammalian tissue, preferably heart tissue, is provided. These methods involve measuring and comparing the gene expression levels of both apoptosis-specific eIF-5A and proliferating eIF-5A and correlating an incidence of ischemia when the expression level of apoptosis-specific eIF-5a is higher than proliferating eIF-5A. In the method of reducing apoptosis in mammalian tissue, there is provided an agent that inhibits expression of apoptosis-specific eIF-5A. Preferred agents are antisense oligonucleotides to human apoptosis-specific eIF-5A.

Description

[0001] related application [0002] This application is a continuation-in-part of U.S. Application Serial No. 10 / 200,148, filed July 23, 2002, which is a continuation-in-part of U.S. Application Serial No. 10 / 141,647, filed May 7, 2002, which is a continuation-in-part of U.S. Application Serial No. 10 / 141,647, filed May 7, 2002, which is a 2001 Continuation-in-Part of US Application Serial No. 9 / 909,796 filed July 23, 2009. field of invention [0003] The present invention relates to cell apoptosis (apoptosis)-specific eukaryotic initiation factor-5A (eIF-5A) and deoxy 8-hydroxyl-2,7,10-triaminodecanoic acid synthase (DHS) nucleic acid and polypeptide and Methods of modulating apoptosis using apoptosis-specific eIF-5A and DHS. Background of the invention [0004] Apoptosis is a genetically programmed cellular event characterized by well-defined morphological features such as cell shrinkage, chromatin condensation, nuclear fragmentation, and membrane blebbing. Kerr et al. (...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61K48/00C12Q1/68C07H21/04C07K14/47H01S5/022H01S5/024H01S5/0683
CPCC12Q1/6883C07K14/4747C12Q2600/158A61P9/10
Inventor J·E·汤普逊C·泰勒D·克利彻C·迪纳雷洛L·雷兹尼科夫B·波梅兰茨
Owner SENESCO TECHNOLOGIES INC
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