Looking for breakthrough ideas for innovation challenges? Try Patsnap Eureka!

Production of polyketides

a polyketide and recombinant technology, applied in the field of recombinant methods and materials for producing polyketides, can solve the problems of affecting the production of polyketides,

Inactive Publication Date: 2006-10-05
KOSAN BIOSCI
View PDF0 Cites 0 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Benefits of technology

"The present invention provides a method for producing polyketides, specifically epothilones, in a host cell of the suborder Cystobacterineae. This is achieved through the use of recombinant host cells containing recombinant expression vectors that encode heterologous PKS genes. The invention also provides recombinant DNA vectors capable of chromosomal integration or extrachromosomal replication in the host cells. The recombinant host cells produce epothilones at higher levels than natural occurring organisms, and can produce mixtures of epothilones that are less complex than natural occurring organisms. The invention also provides novel epothilone derivative compounds that can be used in agriculture, veterinary practice, medicine, and the manufacture of another compound. The patent text also describes a method of treating cancer by administering a therapeutically effective amount of a novel epothilone compound of the invention."

Problems solved by technology

While such methods are valuable and highly useful, certain polyketides are expressed only at very low levels or are toxic to the heterologous host cell employed.
However, the production of epothilone was only about 50 to 100 μg / L and appeared to have a deleterious effect on the producer cells.
Despite the success of these efforts, the chemical synthesis of the epothilones is tedious, time-consuming, and expensive.
Indeed, the methods have been characterized as impractical for the full-scale pharmaceutical development of an epothilone.

Method used

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
View more

Image

Smart Image Click on the blue labels to locate them in the text.
Viewing Examples
Smart Image
  • Production of polyketides
  • Production of polyketides
  • Production of polyketides

Examples

Experimental program
Comparison scheme
Effect test

example 1

Construction of a Myxococcus xanthus Expression Vector

[0176] The DNA providing the integration and attachment function of phage Mx8 was inserted into commercially available pACYC184 (New England Biolabs). An ˜2360 bp MfeI-SmaI from plasmid pPLH343, described in Salmi et al., February 1998, J. Bact. 180(3): 614-621, was isolated and ligated to the large EcoRI-XmnI restriction fragment of plasmid pACYC184. The circular DNA thus formed was ˜6 kb in size and called plasmid pKOS35-77.

[0177] Plasmid pKOS35-77 serves as a convenient plasmid for expressing recombinant PKS genes of the invention under the control of the epothilone PKS gene promoter. In one illustrative embodiment, the entire epothilone PKS gene with its homologous promoter is inserted in one or more fragments into the plasmid to yield an expression vector of the invention.

[0178] The present invention also provides expression vectors in which the recombinant PKS genes of the invention are under the control of a Myxococcus ...

example 2

Construction of a Bacterial Artificial Chromosome (BAC) for Expression of Epothilone in Myxococcus xanthus

[0185] To express the epothilone PKS and modification enzyme genes in a heterologous host to produce epothilones by fermentation, Myxococcus xanthus, which is closely related to Sorangium cellulosum and for which a number of cloning vectors are available, is employed in accordance with the methods of the invention. M. xanthus and S. cellulosum are myxobacteria and so may share common elements of gene expression, translational control, and post translational modification (if any). M. xanthus has been developed for gene cloning and expression: DNA can be introduced by electroporation, and a number of vectors and genetic markers are available for the introduction of foreign DNA, including those that permit its stable insertion into the chromosome. M. xanthus can be grown with relative ease in complex media in fermentors and can be subjected to manipulations to increase gene expres...

example 3

Process for the Production of Epothilones B and D

A. Production of Epothilone B

[0197] I. Flasks

[0198] A 1 mL vial of the K111-32-25 strain is thawed and the contents transferred into 3 mL of CYE seed media in a glass tube. This culture is incubated for 72±12 hours at 30° C., followed by the subculturing of 3 mL of this tube culture into 50 mL of CYE media within a 250 mL baffled Erlenmeyer flask. This CYE flask is incubated for 24±8 hours at 30° C., and 2.5 mL of this seed (5% v / v) used to inoculate the epothilone production flasks (50 mL of CTS-TA media in a 250 mL baffled Erlenmeyer flask). These flasks are then incubated at 30° C. for 48±12 hours, with a media pH at the beginning of 7.4. The peak epothilone A titer is 0.5 mg / L, and the peak epothilone B titer is 2.5 mg / L.

[0199] II. Fermentors

[0200] A similar inoculum expansion of K111-32-25 as described above is used, with the additional step that 25 mL of the 50 mL CYE seed is subcultured into 500 mL of CYE. This secondary ...

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to View More

PUM

PropertyMeasurementUnit
total volumeaaaaaaaaaa
total volumeaaaaaaaaaa
total volumeaaaaaaaaaa
Login to View More

Abstract

Recombinant host cells of the suborder Cystobacterineae containing recombinant expression vectors that encode heterologous PKS genes can produce polyketides synthesized by the PKS enzymes encoded on those vectors at high levels.

Description

CROSS-REFERENCE TO RELATED APPLICATIONS [0001] This application claims priority to U.S. patent application Ser. No. 09 / 443,501, filed 19 Nov. 1999; PCT patent application US99 / 27438, filed 19 Nov. 1999; and U.S. provisional application Ser. Nos. 60 / 130,560, filed 22 Apr. 1999; 60 / 122,620, filed 3 Mar. 1999; 60 / 119,386, filed 10 Feb. 1999; and 60 / 109,401, filed 20 Nov. 1998, each of which is incorporated herein by reference.REFERENCE TO GOVERNMENT FUNDING [0002] This invention was supported in part by SBIR grant 1R-43-CA79228-01. The U.S. government has certain rights in this invention.FIELD OF THE INVENTION [0003] The present invention provides recombinant methods and materials for producing polyketides in recombinant host cells. The recombinant host cells are from the suborder Cystobacterineae, preferably from the genera Myxococcus and Stigmatella that have been transformed with recombinant DNA expression vectors of the invention that encode modular or iterative polyketide synthase...

Claims

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to View More

Application Information

Patent Timeline
no application Login to View More
Patent Type & Authority Applications(United States)
IPC IPC(8): C12P17/00C07H21/04C12N1/21C07D417/02C07D277/22C07D277/28C07D313/00C07D405/06C07D413/06C07D417/06C07D491/04C07D493/04C07D493/08C07D498/08C07D513/08C12N1/20C12N9/00C12N15/52C12N15/74C12P17/16C12P17/18
CPCC07D313/00C12R1/01C07D409/06C07D413/06C07D417/06C07D491/04C07D493/04C07D493/08C07D513/08C12N1/20C12N9/00C12N15/1058C12N15/52C12N15/74C12P17/16C12P17/181C07D405/06C12N1/205C12R2001/01
Inventor JULIEN, BRYANKATZ, LEONARDKHOSLA, CHAITAN
Owner KOSAN BIOSCI
Who we serve
  • R&D Engineer
  • R&D Manager
  • IP Professional
Why Patsnap Eureka
  • Industry Leading Data Capabilities
  • Powerful AI technology
  • Patent DNA Extraction
Social media
Patsnap Eureka Blog
Learn More
PatSnap group products

Browse by: Latest US Patents, China's latest patents, Technical Efficacy Thesaurus, Application Domain, Technology Topic, Popular Technical Reports.

© 2024 PatSnap. All rights reserved.Legal|Privacy policy|Modern Slavery Act Transparency Statement|Sitemap|About US| Contact US: help@patsnap.com