Soluble and immunoreactive variants of HTLV capsid antigen P24
Inventors
Faatz, Elke • Scholz, Christian • Muench, Peter
Assignees
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Publication Number
US-11567079-B2
Publication Date
2023-01-31
Expiration Date
Abstract
The invention concerns soluble and antigenic HTLV p24 variants that can be fused to chaperones and their use in diagnostic applications such as immunoassays for detecting antibodies against HTLV-I or HTLV-II in an isolated biological sample. In particular, the invention relates to a soluble HTLV-I or HTLV-II p24 antigen comprising either the N- or the C-terminal domain of p24 and lacking the other domain. Moreover, the invention covers recombinant DNA molecules encoding these HTLV-I and -II fusion antigens as well as their recombinant production using expression vectors and host cells transformed with such expression vectors. In addition, the invention focuses on compositions of these HTLV p24 antigens with HTLV gp21 antigen and on an immunoassay method for detection of HTLV antibodies using the antigens of the invention. Also the use of HTLV p24 antigens in an in vitro diagnostic assay as well as a reagent kit for detection of anti-HTLV-antibodies comprising said HTLV antigens is encompassed.
Core Innovation
The invention relates to an HTLV-I/HTLV-II diagnostic platform based on soluble, immunoreactive HTLV p24 capsid antigen domain fragments. The platform uses p24 N-terminal domain and p24 C-terminal domain variants in which the presented antigen lacks the corresponding other p24 domain, fused to an oligomeric chaperone and expressed as an individual distinct molecule in the composition.
Soluble p24 antigen domain fragments are provided as fusion polypeptides with an oligomeric chaperone selected from SlyD, FkpA, and Skp. The diagnostic compositions include antigen components comprising HTLV gp21 antigen together with the HTLV p24-chaperone fusion polypeptide for use in immunoassay formats including a double antigen sandwich and double antigen bridge format.
Technical rationale and results are documented showing that full-length p24 aggregates, while isolated CTD fused to FkpA or Skp shows improved early IgM detection. Spectroscopic evidence indicates reversible refolding and maintenance of native-like ordered conformation for Skp-p24/CTD and FkpA-p24/CTD.
Claims Coverage
The independent claim covers a composition-defined HTLV p24 N-terminal domain-only fusion polypeptide architecture fused to an oligomeric chaperone and expressed as an individual distinct molecule. The claim set also includes separate HTLV gp21 antigen and sequence-defined fusion polypeptide variants with at least 80% or at least 95% identity.
HTLV p24 N-terminal domain lacks CTD fused to an oligomeric chaperone and expressed as an individual distinct molecule
A composition comprising an HTLV p24-chaperone fusion polypeptide comprising an N-terminal domain of HTLV p24, wherein the HTLV p24 antigen lacks the C-terminal domain, fused to an oligomeric chaperone, and expressed as an individual distinct molecule.
Oligomeric chaperone selected from Skp or FkpA
The soluble HTLV p24 antigen is prepared using an oligomeric chaperone selected from Skp or FkpA.
Separate polypeptides comprising HTLV p24-chaperone fusion polypeptide and HTLV gp21 antigen
A composition is provided that includes separate polypeptides: an HTLV p24-chaperone fusion polypeptide and an HTLV gp21 antigen with an amino acid sequence of SEQ ID NO:25.
Fusion polypeptide defined by selected SEQ ID sequences with at least 80% identity variants
The fusion polypeptide composition includes an amino acid sequence corresponding to SEQ ID NO:14, 15, 16, 23, or 24, or a variant that is at least 80% identical.
Fusion polypeptide defined by selected SEQ ID sequences with at least 95% identity variants
The fusion polypeptide has an amino acid sequence matching SEQ ID NO:14, 15, 16, 23, or 24, or a variant that is at least 95% identical.
The claim set centers on compositions containing HTLV p24 N-terminal domain-only antigen fused to an oligomeric chaperone and expressed as a distinct molecule, optionally together with HTLV gp21 antigen SEQ ID NO:25 as a separate polypeptide. Sequence scope is tightened through SEQ ID-referenced fusion-polypeptide options and variant identity thresholds.
Stated Advantages
Improved early IgM detection when isolated CTD is fused to FkpA or Skp, especially with Skp-p24/CTD.
Spectroscopic evidence indicates Skp-p24/CTD and FkpA-p24/CTD refold reversibly and maintain native-like ordered conformation.
Documented Applications
Immunoassay detection of anti-HTLV IgG/IgM in samples using antigen compositions comprising HTLV p24-chaperone fusion polypeptides and HTLV gp21 antigen SEQ ID NO:25, including double antigen sandwich and double antigen bridge formats.
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