Conformationally stabilized RSV pre-fusion F proteins
Inventors
Marshall, Christopher • Yondola, Mark • Mariani, Roberto • Zomback, Aaron • Gidwani, Sonal
Assignees
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Publication Number
US-11926649-B2
Publication Date
2024-03-12
Expiration Date
Abstract
The present invention provides mutant RSV F molecules, such as those that can be, or are stabilized, in a pre-fusion conformation by the introduction of one or more DT cross-links. The present invention also provides methods of making such mutant RSV F molecules, compositions comprising such mutant RSV F molecules, and methods of use of such mutant RSV F molecules, for example in vaccination methods, therapeutic methods, and antibody production methods.
Core Innovation
The invention provides a soluble, mature, trimeric respiratory virus fusion (F) protein complex for RSV in which each protein in the trimeric complex comprises point mutations to tyrosine at amino acid residues 428, 226, and 185, with the amino acid numbering determined by alignment to SEQ ID NO.1. The invention further provides structural stabilization of the F protein in the pre-F conformation by incorporating one or more di-tyrosine (DT) cross-links, including an intramolecular 198Y-226Y cross-link and an intermolecular 185Y-428Y cross-link.
The stabilized soluble mature trimeric RSV F immunogens are described as useful for generating and producing anti-RSV neutralizing antibodies. The pre-F conformation better displays potent neutralizing antigenic site Ø, and the disclosed constructs are described as providing increased neutralization titers, an improved antigenic profile, and increased trimer production.
The partial content also indicates sequence examples for wild type and multiple mutant constructs, including variants with tyrosine substitutions at key residues and additional engineered mutations, along with optional C-terminal tags/linkers and artificial sequence elements. It further describes characterization and comparative evaluation of related variants, including designs identified as AVR02/DT-AVR02 and related mutants, to support maintaining the desired pre-F conformation.
The partial content further describes use in non-human mammalian settings, including cotton rats, to evaluate vaccination outcomes. It reports reduced RSV titers after vaccination and enhanced neutralizing antibody responses versus controls, and references formulation-related discussion, including sucrose to reduce aggregation, and analytical characterization context such as size-exclusion chromatography (SEC).
Claims Coverage
The document provides at least two independent claims, covering a soluble, mature, trimeric respiratory virus fusion (F) protein complex and a method for eliciting anti-RSV antibodies by administering that complex. The main inventive features include the specific residue-to-tyrosine point mutations at residues 428, 226, and 185 and, in refinements, conformational stabilization in the pre-F state via one or more di-tyrosine cross-links.
Soluble, mature, trimeric respiratory virus fusion (F) protein complex with tyrosine point mutations at residues 428, 226, and 185
A soluble, mature, trimeric respiratory virus fusion (F) protein complex in which each protein in the trimeric complex comprises point mutations to tyrosine at amino acid residue 428, amino acid residue 226, and amino acid residue 185, where the amino acid numbering is determined by alignment to SEQ ID NO.1.
Eliciting production of anti-RSV antibodies by administering a stabilized soluble, mature trimeric RSV F protein complex
A method of eliciting production of anti-RSV antibodies in a non-human mammalian subject by administering an effective amount of a soluble, mature trimeric RSV F protein complex, wherein each protein in the trimeric complex comprises point mutations to tyrosine at amino acid residue 428, amino acid residue 226, and amino acid residue 185, with amino acid numbering determined by alignment to SEQ ID NO.1.
Di-tyrosine cross-links and stabilization in the pre-F conformation
The respiratory virus F protein complex includes one or more di-tyrosine cross-links and is stabilized in the pre-F conformation.
Overall, the claim coverage centers on soluble, mature, trimeric RSV F immunogens defined by specific tyrosine point mutations at residues 428, 226, and 185. In refinements tied to the claim set, the complexes are further characterized by one or more di-tyrosine cross-links and stabilization in the pre-F conformation, and these immunogens are used to elicit anti-RSV antibody production in a non-human mammalian subject.
Stated Advantages
Increased neutralization titers.
Improved antigenic profile.
Increased trimer production.
Better display of potent neutralizing antigenic site Ø.
Reduced RSV titers after vaccination in cotton rats.
Enhanced neutralizing antibody responses versus controls.
Documented Applications
Vaccination and therapeutic intervention using soluble mature trimeric RSV F immunogens.
Antibody production, including producing anti-RSV neutralizing antibodies.
Producing anti-RSV antibodies in a non-human mammalian subject by administering an effective amount of the soluble, mature trimeric RSV F protein complex.
Vaccination of non-human mammalian subjects using the soluble, mature trimeric RSV F complex to elicit anti-RSV antibodies, with reporting of reduced RSV titers and enhanced neutralizing antibody responses versus controls.
Progression to higher-order testing and human clinical trial descriptions is mentioned in the partial content, including a phase I human clinical trial context.
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