Use of marrow-derived glial progenitor cells as gene delivery vehicles into the central nervous system

a technology of marrow-derived glial progenitor cells and central nervous system, which is applied in the direction of cell culture active agents, blood/immune system cells, viruses, etc., can solve the problems of glia in adult brains and the debatable origin of microglia, and achieve the effect of greater therapeutic precision

Inactive Publication Date: 2006-06-22
UNITED STATES OF AMERICA
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Benefits of technology

[0016] Although many neurotrophic factors show promise in the treatment of CNS disorders, their use has been hindered by their inability to cross the blood-brain barrier and by their limited diffusion into CNS tissues (34). In addition, adverse effects have been reported after systemic administration of some neurotrophins (35). Using marrow-derived cells to deliver therapeutic proteins directly to the site of CNS pathology likely would be more benign than systemic administration of toxic molecules. In addition, using vectors with cell type-specific promoters could restrict gene expression specifically to reactive astroglia or microglia, thereby providing greater therapeutic precision for gene therapy of CNS disease.

Problems solved by technology

However, it is unclear whether the glia in adult brains free of disease or injury originate solely from cells present in the brain since the fetal stage of development, or if there is further input into such adult brains from cells originating outside the central nervous system (CNS).
However, the developmental origin of microglia remains debatable (2,3).

Method used

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  • Use of marrow-derived glial progenitor cells as gene delivery vehicles into the central nervous system
  • Use of marrow-derived glial progenitor cells as gene delivery vehicles into the central nervous system
  • Use of marrow-derived glial progenitor cells as gene delivery vehicles into the central nervous system

Examples

Experimental program
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Effect test

experiment 1

Gene Transfer and Bone Marrow Transplantation

[0020] Gene transfer into hematopoietic precursors was done as previously described (11, 12), with the addition of stem cell factor to optimize transduction of reconstituting hematopoietic stem cells (13). C57BL / 6J mice (Jackson Laboratories, Bar Harbor, Me.), 6-8 weeks old, were used as donors. Forty-eight hours before marrow harvest, the mice were injected with 5-fluorouracil at a dose of 150 mg / kg to ablate mature blood cells and thereby induce progenitor cells into cycle. Upon harvest, marrow was placed into liquid culture in suspension dishes and grown in Dulbecco's modified Eagle's medium containing 15% fetal bovine serum (Whittaker Bioproducts, Walkersville, Md.) and supplemented with IL-3 (50 ng / ml), IL-6 (100 ng / ml) and stem cell factor (100 ng / ml). Growth factors were used to maintain early hematopoietic cells in cycle (13). All were obtained from R & D Systems (Minneapolis, Minn.). After 48 hr in culture with growth factors, m...

experiment 2

In situ Hybridization Histochemistry

[0023] Tissues were evaluated with both oligonucleotide and RNA probes. To detect neo R transcripts, two oligonucleotide probes were prepared, complementary to the sequence of the neoR gene either from nucleotides 222-269 or from nucleotides 447-494 (numbering with the A of the initiation codon as 1). The oligonucleotides were labeled using terminal transferase (Boehringer-Mannheim, Indianapolis, Ind.) and 35S-dATP (New England Nuclear, Boston, Mass.) as described previously (16). An RNA probe, complementary to the entire neoR coding region, was labeled with 35S-UTP using SP6 polymerase (17). Labeling with radioactive probes was detected by dipping hybridized sections in photographic emulsion. Emulsion was exposed for 14 days, then developed and sections were stained, air dried, and coverslipped for microscopic examination. To detect male bone marrow cells transplanted into female recipients, sequences specific to the donor mouse Y chromosome wer...

experiment 3

Nuclear Staining

[0025] To confirm that Y chromosome ISHH coincided with cell nuclei, sections were counterstained with ethidium bromide or 4′,6-diamidino-2-phenylindole (DAPI). Staining was detected by illumination with a mercury lamp using a microscope equipped for fluorescence micrography.

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Abstract

The present disclosure relates to a method for introducing a hematopoietic cell into the brain of a mammal, by administering bone marrow-derived progenitor cells into the body of the mammal by intravenous injection. The bone marrow-derived cell is preferably a cell that differentiates into a glial cell. The disclosure also relates to a method for delivery of therapeutic protein molecules into the brain of a mammal, by administering to a mammal an effective amount of bone marrow-derived progenitor cells which contain a gene having a nucleic acid sequence that encodes a functional therapeutic protein. Isolated recombinant cells and a pharmaceutical composition are also provided.

Description

REFERENCE TO RELATED APPLICATIONS [0001] This is a divisional of U.S. patent application Ser. No. 10 / 122,703, filed Apr. 11, 2002, now allowed, which is a continuation-in-part of U.S. patent application Ser. No. 09 / 819,096, filed Feb. 16, 2001, now abandoned, which is a continuation of U.S. patent application Ser. No. 09 / 058,160, filed Apr. 10, 1998, now abandoned, which claims the benefit of U.S. Provisional Application No. 60 / 036,592, filed Apr. 10, 1997.FIELD [0002] The present disclosure relates to methods for introducing hematopoietic cells into the brain of a mammal, the differentiation of adult bone marrow cells into glial cells, and the use of marrow-derived glial progenitor cells as gene delivery vehicles into the brain. BACKGROUND [0003] Glial cells are thought to derive embryologically from either myeloid cells of the hematopoietic system (microglia) or neuroepithelial progenitor cells (astroglia and oligodendrocytes). However, it is unclear whether the glia in adult brai...

Claims

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Application Information

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Patent Type & Authority Applications(United States)
IPC IPC(8): A61K48/00C12N5/08C12N15/867A61K38/18C07K14/475C12N5/0789
CPCA61K48/00C07K14/475C12N5/0647C12N15/87C12N2501/125C12N2501/23C12N2510/00C12N2510/02C12N2799/027A61K38/185A61K35/28A61K2300/00
Inventor EGLITIS, MARTIN A.MEZEY, EVAMOURADIAN, MARY MARAL
Owner UNITED STATES OF AMERICA
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