Cell surface display of polypeptide isoforms by stop codon readthrough

US10724033B2 · kind B2 · utility

Assignee

• Novartis AG · CH

Inventors

• Thomas Jostock · Neuenburg am Rhein, DE
• Hans-Peter Knopf · Schallstadt, DE
• Burkhard Wilms · Weil am Rhein, DE
• Audrey Nommay · Blotzheim, FR

Key dates

• Filing date: Aug 8, 2017
• Grant date: Jul 28, 2020
• Expiry date: Aug 8, 2037

Classification

• Technology area (CPC C): Chemistry; Metallurgy
• CPC primary: C07K2319/035
• WIPO field: Biotechnology
• WIPO sector: Chemistry

Abstract

A method for selecting mammalian host cells that express a polypeptide of interest with high yield is described. The host cells comprise an expression cassette comprising at least a first polynucleotide encoding the polypeptide of interest, at least one leaky stop codon downstream of the first polynucleotide, and a second polynucleotide downstream of the leaky stop codon encoding an immunoglobulin transmembrane anchor. The host cells are cultivated to allow expression of the polypeptide of interest such that at least a portion of the polypeptide of interest is expressed as a fusion polypeptide that is displayed on the cell surface. The cells are selected based upon the presence or amount of the displayed fusion polypeptide. Also provided are methods for producing a polypeptide that employ a respective selection to identify high expressing host cells.