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-11267848-B2
Publication Date
2022-03-08
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 relates to soluble, mature trimeric RSV F molecules comprising specific point mutations that introduce tyrosine substitutions at amino acid residues 428, 226, and 185, and additional point mutations to phenylalanine at residue 190 and to leucine at residue 207, or corresponding residues as determined by alignment. The described molecular forms include F0 precursor, F1 and F2 polypeptides, protomers, and mature soluble or membrane-bound trimers of RSV F, with the overall focus on pre-fusion molecular stabilization.
The invention also relates to RSV F immunogens stabilized in the pre-fusion conformation using di-tyrosine cross-links introduced by the above mutations. The described cross-link designs include an intramolecular di-tyrosine cross-link and an intermolecular di-tyrosine cross-link, and the disclosure includes configurations with multiple di-tyrosine links.
The disclosed pre-fusion stabilized mutant RSV F immunogens are intended to improve binding to pre-fusion/site Ø antibodies and to elicit neutralizing antibodies for vaccination, therapy, and antibody production. The document further discusses functional and immunological goals associated with the stabilized pre-fusion antigens, including improved serum neutralization titers and reduced tendency to form aggregates.
Claims Coverage
The disclosure includes an independent claim directed to a soluble, mature trimeric RSV F molecule defined by a specific set of point mutations that underpin pre-fusion stabilization via introduced tyrosines. Dependent claim refinements further constrain included sequence regions and add di-tyrosine crosslinks, nucleic acids, pharmaceutical compositions with adjuvant, and vaccination methods.
Soluble, mature trimeric RSV F with defined point mutations
A soluble, mature trimeric RSV F molecule comprising point mutation to tyrosine at amino acid residue 428, at residue 226, and at residue 185, point mutation to phenylalanine at amino acid residue 190, and point mutation to leucine at amino acid residue 207, or corresponding residues as determined by alignment to SEQ ID NO.1.
Sequence-region constraints for the RSV F molecule
The RSV F molecule further comprises amino acid residues 26-109 and 137-513 of SEQ ID No. 81.
Di-tyrosine crosslinks and stabilization in the pre-F conformation
The RSV F molecule comprises one or more di-tyrosine cross links and is stabilized in the pre-F conformation.
Encoding nucleic acid for the defined RSV F molecule
A nucleic acid molecule encodes an RSV F molecule as defined in claim 2.
Pharmaceutical composition with an adjuvant
A pharmaceutical composition according to claim 16 further comprises an adjuvant.
Vaccination method by administering an effective amount
A method of vaccinating a subject against RSV by administering an effective amount of a pharmaceutical composition according to claim 22 to a subject in need thereof.
The claim coverage centers on a soluble, mature trimeric RSV F antigen defined by a specific set of point mutations at residues 428, 226, 185, 190, and 207, with additional features including sequence-region constraints, di-tyrosine crosslinks and pre-F conformational stabilization, nucleic acids, adjuvanted pharmaceutical compositions, and vaccination methods.
Stated Advantages
Reduces RSV/A/Tracy viral titers in lung and nasal wash after prime-boost vaccination.
Generates RSV/A/Tracy neutralizing antibodies.
Generates cross-reactive RSV/B/18537 neutralizing antibodies.
Improved serum neutralization titers.
Stabilization of the pre-fusion conformation.
Reduced aggregate formation.
Documented Applications
Vaccination of animals, including mice and cotton rats, using DT-AVR02/DT-preF formulated with an alum adjuvant (Alhydrogel), with prime-boost administration and RSV/A/Tracy challenge measurements.
Vaccination methods involving administering a pharmaceutical composition comprising the RSV F molecule to a subject in need thereof.
Therapeutic methods for RSV in connection with eliciting neutralizing antibodies for therapy.
Antibody production methods in connection with eliciting neutralizing antibodies for antibody production.
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