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Conditional immortalisation of cells

A conditional, cell-based technology, applied in the field of immortalization, can solve the problem that cells are not suitable for transplantation therapy

Inactive Publication Date: 2002-10-09
RENEURON LTD +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

This means that such cells are not suitable for transplant therapy

Method used

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  • Conditional immortalisation of cells
  • Conditional immortalisation of cells

Examples

Experimental program
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Embodiment 1

[0036] 1. Isolation of mammary interlobular fibroblasts (HMF) and mammary microvascular endothelial (MMVE) cells:

[0037]Cultures of mammary interlobular fibroblasts and mammary microvascular endothelial cells were prepared from normal breast tissue of patients undergoing plastic surgery (mammoplasty) with consent obtained. The culture of interlobular fibroblasts was prepared according to the method of Atherton et al., J. Cell Sci., 107: 2931-2939, and Dulbecco's modified Eagles medium supplemented with 10% fetal bovine serum and antibiotics (penicillin / streptomycin) ( DMEM) to maintain the culture. Endothelial cells from microvessels were isolated from primary cultures by immunomagnetic sorting using the anti-clotting regulator QBEND-40 mouse monoclonal antibody essentially according to Drake and Lock, Human Reprod. , 1992; 6: 1156-1159 as described in the method operation. Endothelial cultures were established and maintained in EGM-2 medium (Biowhittaker).

[0038] 2. Cu...

Embodiment 2

[0054] This example demonstrates the preparation of a suitable expression vector for the co-expression of three genes; the three genes are: heat-labile T antigen (U19tsA58) from a non-DNA-binding mutant in the early region of SV40; telomere end transfer The catalytic subunit hTERT (cDNA) of the enzyme complex (Counter et al., PNAS, 1998; 95(25): 14723-14728); and a dominant selective neomycin phosphotransferase resistance marker (Neo), which Encodes resistance to G418 (Clontech). The final construct was assembled in the Moloney murine leukemia virus (Mo MuLV)-based high-titer reverse transcription expression vector pBabe (Morgenstern and Land, supra). Transfer of the SV40 early region through the retroviral life cycle into viruses producing only the large T antigen. All constructs were assembled in rec-A host (JS4-rec-A derivative of MC1061) by ampicillin selection.

[0055] A total of two versions of the vector were constructed;

[0056] Structure 1

[0057] This structur...

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Abstract

A recombinant, or genetically engineered, mammalian cell, comprises a conditionally-inducible oncogene and an exogenous polynucleotide encoding the catalytic sub-unit of the telomerase complex. The recombinant cells are useful in therapy, to replace lost or damaged cells.

Description

field of invention [0001] The present invention relates to the immortalization of mammalian cells for therapeutic applications. Background of the invention [0002] Awareness and understanding of the importance of transplantation therapy in the treatment of tissue and organ damage is increasing. And while organ transplants are being widely practiced, transplant therapies based on single-cell transplantation are still relatively early in clinical development. [0003] For example, there is growing awareness that transplanting suitable cells into the injured brain may improve or correct any impairments in perception, movement, behavior, or psychology that result from the injury. [0004] For cell-based therapies to be useful, it must be possible to obtain sufficient cells for transplantation. One way to ensure this is to culture undifferentiated cells under conditions that allow repeated cell division and growth. One difficulty with using undifferentiated cells is that unre...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/09A61K35/12A61K35/30A61K38/00A61K48/00A61P25/28C07K14/82C12NC12N5/10C12N9/12C12N15/12C12N15/54
CPCA61K35/12A61K38/00C07K14/82C12N9/1241C12N2799/027A61P25/28
Inventor P·杰特
Owner RENEURON LTD
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