Manufacturing optimization of GL-2045, a multimerizing stradomer
Inventors
Block, David S. • Merigeon, Emmanuel Y. • Olsen, Henrik
Assignees
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Publication Number
US-12195499-B2
Publication Date
2025-01-14
Expiration Date
Abstract
The present disclosure involves optimized methods for production of biologically active proteins termed optimally manufactured stradomers. The present disclosure further provides compositions and methods useful in the treatment of diseases and conditions including autoimmune diseases, inflammatory diseases, or infectious diseases.
Core Innovation
The invention is directed to a composition comprising a recombinantly produced homodimer and a multimer of the homodimer, where the homodimer comprises two monomers each comprising amino acids 21-264 of SEQ ID NO:4, and where the highest order multimer fractions comprise at least about 30% of the total composition. The multimer profile is described as dominated by higher-order multimers and is linked to the desired drug substance multimer profile for GL-2045.
The patent addresses the manufacturing challenge of retaining a defined multimer pattern during production and purification. It describes that repeated protein A affinity purification can lead to loss of low molecular weight bands after successive runs because high-order multimers occupy protein A binding sites, requiring more frequent and stringent CIP to regenerate the column.
To preserve the multimer profile across purification cycles, the document describes production in Chinese Hamster Ovary (CHO) cells stably transfected with an expression vector comprising a nucleic acid sequence encoding SEQ ID NO:4, with a growth-temperature shift from 37°C ±1°C to 32.5°C ±1°C. It further describes chromatography and polishing steps, including protein A affinity chromatography, ion exchange, hydrophobic interaction chromatography, and analytical SEC-HPLC or HPLC, to separate multimer species and maintain the defined final multimer composition range.
Claims Coverage
The partial claim set includes two independent claims. The inventive features center on a defined recombinant homodimer composition with a highest-order multimer fraction threshold, and on a CHO-cell production method with a defined temperature shift and cell viability constraint.
Composition with high-order multimer fraction
A composition comprising a recombinantly produced homodimer and a multimer of the homodimer, wherein the homodimer comprises two monomers each comprising amino acids 21-264 of SEQ ID NO:4, and wherein the highest order multimer fractions of the composition comprise at least about 30% of the total composition.
Producing the homodimer and multimer using stably transfected CHO cells with temperature shift
A method for producing a composition comprising a homodimer and a multimer of the homodimer, wherein the homodimer comprises two monomers each comprising amino acids 21-264 of SEQ ID NO:4, comprising culturing Chinese Hamster Ovary (CHO) cells that have been stably transfected with an expression vector comprising a nucleic acid sequence encoding SEQ ID NO:4 at a growth temperature of 37°C ±1°C for 5 ±1 days, shifting the growth temperature to 32.5°C ±1°C, and harvesting the homodimer and the multimer from the culture media, wherein cell viability exceeds 85% at day 10, day 11, day 12, day 13, day 14, or longer of culture.
The claims cover a specific recombinantly produced homodimer defined by amino acids 21-264 of SEQ ID NO:4 with multimers such that the highest order multimer fractions are at least about 30% of the total composition, together with a production method using stably transfected CHO cells, a defined temperature shift, and a cell viability threshold exceeding 85% for the specified culture period.
Stated Advantages
Retains a defined multimer pattern dominated by higher-order multimers.
Requires fewer and less stringent CIP to regenerate the column by preserving the multimer profile across purification cycles.
Achieves cell viability exceeding 85% at day 10 through day 14 or longer of culture.
Provides higher-order multimer fractions associated with enhanced binding/functional efficacy, including FcγRIIIa avidity, CDC inhibition, and chemotaxis inhibition.
Documented Applications
Treatment or prevention of an inflammatory, autoimmune, or infectious disease or disorder in a subject using the claimed composition.
Treatment or prevention of idiopathic thrombocytopenia purpura, chronic inflammatory demyelinating polyradiculoneuropathy, multifocal motor neuropathy, myasthenia gravis, organ transplantation, and rheumatoid arthritis.
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