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Nucleic acids, polypeptides, and methods for modulating apoptosis

A technology for apoptosis and cancer cells, applied in the fields of nucleic acid, polypeptide, and regulation of cell apoptosis, which can solve the problems of normal cells having little effect and failing to occur in time.

Inactive Publication Date: 2011-01-26
SENESCO TECHNOLOGIES INC
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Thus, depletion of activated apoptosis-specific eIF-5A due to inhibition of deoxy-8-hydroxy-2,7,10-triaminodecanoic acid synthase activity may not occur in time to block toxicity induced by spermidine apoptosis
Second, polyamines are competitive inhibitors of the deoxy-8-hydroxy-2,7,10-triaminodecanoic acid reaction and thus cannot completely block the reaction even at toxic concentrations
Because these mRNAs are transcribed in cancer cells, but not in adjacent normal cells, apoptotic eIF-5A is expected to promote apoptosis in cancer cells but has minimal, if any, effect on normal cells

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
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Effect test

Embodiment 1

[0134] This example demonstrates the isolation and characterization of a full-length cDNA encoding a rat eIF-5A nucleic acid with apoptosis-specific expression.

[0135] Superovulation and induction of apoptosis in rat corpus luteum

[0136] Subcutaneously inject 50 IU of PMSG (pregnant mare serum gonadotropin) to immature (21-30 days old) female rats, and 60-65 hours later, inject 50 IU of HCG (human chorionic gonadotropin) to make It superovulates. After being treated with HCG for 7 days, 500 mg of PGF-2α was subcutaneously injected to induce apoptosis of corpus luteum cells. After treatment with PGF-2[alpha], rats are sacrificed at various times (eg, 1, 8, and 24 hours), and the corpus luteum is excised and placed in liquid nitrogen. The corpus luteum tissues of the control group were obtained from rats sacrificed immediately before PGF-2α treatment.

[0137] Dispersion of luteal cells in rat ovary

[0138] Six to nine days after superovulation, rats were subcutaneously...

Embodiment 2

[0174] This example demonstrates the regulation of apoptosis using apoptosis-specific eIF-5A and DHS.

[0175] Culture of COS-7 cells and isolation of RNA

[0176] COS-7, an African green monkey kidney fibroblast-like cell line transformed with an SV40 mutant encoding wild-type T antigen, can be used for all transfection-based experiments. COS-7 cells were cultured in Dulbecco's modified Eagle's medium (DMEM) containing 0.584 g L-glutamine, 4.5 g glucose, and 0.37% sodium bicarbonate per liter. The medium was also supplemented with 10% fetal bovine serum (FBS) and 100 units of penicillin / streptomycin. cells at 37°C, 5% CO 2 and 95% air humid environment to grow. Every 3-4 days, the cells were cultured again by detaching adherent cells with a solution of 0.25% trypsin and 1 mM EDTA. Disperse the isolated cells at a ratio of 1:10 in a new Petri dish containing fresh medium.

[0177] COS-7 cells used for RNA isolation were grown in 150-mm tissue culture dishes (Corning). Ce...

Embodiment 3

[0195] This example demonstrates the regulation of apoptosis using apoptosis-specific eIF-5A and DHS.

[0196] Using the method and general procedure described in the previous examples, Figure 23 is a flowchart illustrating the transient transfection of COS-7 cells, wherein the cells in serum-free medium were incubated for 4 hours in the plasmid DNA of lipofectamine, added Serum, cells were incubated for an additional 40 hours. Either culture the cells in regular medium containing serum for an additional 48 hours (i.e., without further treatment) prior to analysis, induce apoptosis by depriving serum for 48 hours prior to analysis, or Previously, cells were treated with actinomycin D for 48 hours to induce apoptosis.

[0197] Figure 22 illustrates the transient expression of exogenous proteins in COS-7 cells after transfection with pHM6. Proteins were mock-transfected, or with pHM6-LacZ, pHM6-antisense 3'rF5A (pHM6-antisense 3'UTR rat apoptosis eIF-5A), or pHM6-sense rF5A (p...

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Abstract

The invention discloses a nucleic acids, polypeptides, and methods for modulating apoptosis. The present invention relates to isolated and / or purified rat apoptosis-specific eucaryotic initiation Factor-5A (eIF-5A) and deoxyhypusine synthase (DHS) nucleic acids and polypeptides. The present invention also relates to methods of modulating apoptosis using apoptosis-specific eIF-5A and DHS, and antisense oligonucleotides and expression vectors of apoptosis-specific eIF-5A and DHS useful in such methods.

Description

[0001] This application is a divisional application of Chinese patent application 02818632.X "Nucleic Acid, Polypeptide, and Method for Regulating Cell Apoptosis" filed on July 23, 2002. technical field [0002] The present invention relates to the nucleic acid and polypeptide of apoptosis-specific eukaryotic initiation factor-5A (eIF-5A) and deoxy 8-hydroxy-2,7,10-triaminodecanoic acid synthase (DHS), and the use thereof Apoptosis-specific methods of eIF-5A and DHS to regulate apoptosis. Background technique [0003] 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. (1972) Br. J. Cancer, 26, 239-257; Wyllie et al. (1980) Int. Rev. Cytol., 68, 251-306. It plays an important role in normal tissue development and homeostasis, and defects in the apoptotic program are thought to contribute to a variety of human disease...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): A61K48/00A61P35/00C12N15/12H01S5/022H01S5/024H01S5/0683
CPCA61K31/132A61P35/00
Inventor C·泰勒T·-W·王L·佩特罗夫J·C·卡尔森R·纳拉延辛J·E·汤普森D·克利歇M·考普
Owner SENESCO TECHNOLOGIES INC
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