Simultaneous, integrated selection and evolution of antibody/protein performance and expression in production hosts
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Publication Number
US-11976381-B2
Publication Date
2024-05-07
Expiration Date
Abstract
The present disclosure provides methods of integrating therapeutic protein and antibody generation and/or selection, evolution and expression in a eukaryotic host for manufacturing in a single system. Therapeutic proteins, including antibodies, are generated, optimized and manufactured in the same eukaryotic host system. The disclosed system of Comprehensive Integrated Antibody Optimization (CIAO!™) allows for simultaneous evolution of protein performance and expression optimization.
Core Innovation
The disclosed invention provides a method of evolution of a template human protein in a manufacturing host that includes mutating the template human protein to produce a set of mutant human proteins in the manufacturing host, while the manufacturing host incorporates an unnatural amino acid into the mutant human proteins. The method screens the set of mutant human proteins for at least one predetermined property, characteristic or activity, and selects an up-mutant human protein from the screened set based on the predetermined property, characteristic or activity.
After selecting an up-mutant, the method manufactures the selected up-mutant human protein by expressing the up-mutant human protein in the same manufacturing host used in the mutating step. The disclosed platform is described as an integrated approach for evolving and manufacturing therapeutic proteins and antibodies in a single eukaryotic production host using cell-surface display for high-throughput selection.
The disclosed approach further includes functional mapping via comprehensive positional evolution and related non-stochastic confirmed evolution methods, including comprehensive positional insertion, comprehensive positional deletion, and combinatorial protein synthesis. Mutants are screened under manufacturing-mimicking conditions and undesirable immune responses are screened for and eliminated, enabling selection of up-mutants based on optimized properties and enhanced or simultaneous expression.
Claims Coverage
The partial content identifies one independent claim. Across dependent claims, the inventive features are refined to specify the manufacturing host context, including eukaryotic cell production host and cell-surface display, define screening targets and selection criteria, exclude undesirable immune responses, and combine property optimization with modified expression, including enhanced expression.
Unnatural amino acid mutagenesis in a manufacturing host
Mutating the template human protein to produce a set of mutant human proteins in the manufacturing host, wherein the manufacturing host incorporates an unnatural amino acid into the mutant human proteins.
Screening for predetermined property, characteristic, or activity
Screening the set of mutant human proteins produced to identify at least one predetermined property, characteristic or activity.
Selecting an up-mutant based on screened properties
Selecting an up-mutant human protein from the set of mutant human proteins screened based upon the at least one predetermined property, characteristic or activity.
Manufacturing by expressing the selected up-mutant in the same host
Manufacturing the up-mutant human protein by expressing the up-mutant human protein in the same manufacturing host as was used in the mutating step.
Evolving in a eukaryotic cell production host with cell-surface display
Evolving the template human protein in a eukaryotic cell production host to generate mutant human proteins that are displayed on the cell surface.
Two-part selection using property optimization and modified expression
Selecting the up-mutant human protein based on optimizing at least one predetermined property versus a template human protein and selecting based on modified expression versus the template human protein.
Enhanced expression selection
Selecting the up-mutant based on modified expression that is an enhanced expression.
Silent and missense screening compared to the template antibody
Screening includes screening for a silent mutation and screening for a missense mutation, each compared to the template antibody.
Screening for and excluding an undesirable immune response
Screening identifies mutant human proteins that produce an undesirable immune response, and selecting is performed to obtain mutant proteins that do not exhibit that undesirable immune response.
The independent claim centers on evolving a template human protein in a manufacturing host by incorporating an unnatural amino acid during mutagenesis, screening mutants for predetermined properties, selecting an up-mutant based on those properties, and manufacturing by expressing the up-mutant in the same host. Dependent claims further specify eukaryotic cell surface display, silent and missense screening, exclusion of undesirable immune responses, and selection that combines property optimization with modified expression, including enhanced expression.
Stated Advantages
Enables evolving and manufacturing in a single eukaryotic production host by expressing the selected up-mutant in the same manufacturing host used for mutagenesis.
Supports high-throughput selection using cell-surface display.
Allows functional mapping and selection based on optimized predetermined properties and enhanced or simultaneous expression.
Screens for and eliminates undesirable immune responses.
Documented Applications
Integrated evolution and manufacturing of therapeutic proteins/antibodies in a single eukaryotic production host, using cell-surface display for high-throughput selection.
Biosimilar-oriented antibody/protein development is referenced via mention of biosimilar.
Humanized anti-antigen antibody libraries are referenced as a context for the approach.
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