Modified EGF protein, production method therefor, and use thereof
Inventors
Assignees
Interested in licensing this patent?
MTEC can help explore whether this patent might be available for licensing for your application.
Publication Number
US-11339197-B2
Publication Date
2022-05-24
Expiration Date
Abstract
A modified EGF protein, a production method therefor, and a use thereof are disclosed. The modified EGF protein allows an effective delivery of EGF into the cell and exhibits an enhanced half-life span. A method for producing the modified EGF protein includes culturing conditions where host cell growth phase and protein expression phase employ different temperature conditions. A use of the modified EGF protein is also disclosed.
Core Innovation
The invention relates to a modified EGF (EGF-deri) comprising a defined amino-acid sequence, including the amino-acid sequence of SEQ ID NO: 4. In optional forms, the invention also includes a signal-sequence form corresponding to SEQ ID NO: 2. The modified EGF-deri is intended to improve cell membrane binding for effective intracellular delivery.
The modified EGF-deri is further intended to extend in vivo half-life/persistence and to provide stability under severe conditions. The disclosure describes that the modified EGF-deri maintains biological activity under severe storage conditions, including preservation of activity after severe storage and stability through multiple freeze/thaw cycles.
The disclosure provides production and characterization supporting the intended properties. It describes production using an expression vector and host cells, and reports characterization showing active monomer predominance, long-term cell line stability over many passages, and biological activity comparable to standard EGF, while also describing enhanced skin permeability in mouse models.
Claims Coverage
The independent claim provides a structural definition of the modified EGF protein by a specific amino-acid sequence (SEQ ID NO: 4). Dependent claims refine the invention by adding corresponding nucleotide information (SEQ ID NO: 5), production-related culturing conditions, purification-related affinity chromatography using a Protein A-coupled resin, and formulation/use for skin-condition and skin-disease treatment.
Modified EGF protein defined by SEQ ID NO: 4
A modified EGF protein comprising the amino acid sequence of SEQ ID NO: 4.
Nucleotide encoding the modified EGF protein
The modified EGF protein comprising the amino acid sequence of SEQ ID NO: 4 is encoded by the nucleotide sequence of SEQ ID NO: 5.
Two-phase host cell culturing temperatures
Produce the modified EGF protein by culturing a host cell line with the encoding nucleic acid at 35°C to 38°C, then culturing under expression conditions at 32°C to 34°C.
Expression-phase culture at defined pH
The culture in step 2 is carried out at a pH of 6.8 to 7.1.
Affinity chromatography with Protein A-coupled resin
Affinity chromatography uses a Protein A-coupled resin.
Skin-condition improvement and skin disease treatment by administration
Improve skin condition and/or treat skin disease by administering an effective amount of the composition of claim 19 to a subject in need thereof.
Claim coverage centers on a modified EGF protein defined by SEQ ID NO: 4, with corresponding nucleotide encoding, host-cell production involving specified temperature ranges and a defined expression-phase pH, and purification by Protein A-coupled resin affinity chromatography. The dependent use embodiments focus on administering an effective amount of a composition to improve skin condition and/or treat skin disease.
Stated Advantages
Improves cell membrane binding for effective intracellular delivery.
Extends in vivo half-life/persistence.
Provides stability under severe conditions.
Preserves biological activity after severe storage.
Maintains stability through multiple freeze/thaw cycles.
Shows enhanced skin permeability in mouse models.
Provides no relevant blood EGF increase.
Exhibits comparable in vitro biological activity to standard EGF.
Documented Applications
Cosmetic compositions for skin-condition improvement.
Pharmaceutical compositions for preventing/treating skin diseases.
Administration of a composition to a subject in need thereof to improve skin condition and/or treat skin disease.
Interested in licensing this patent?