Multi-epitope fusion protein of an HCV antigen and uses thereof

Patent metadata

Field	Value
Publication number	US-11078241-B2
Application number	US-202016751609-A
Country	US
Kind code	B2
Filing date	Jan 24, 2020
Priority date	Jul 27, 2017
Publication date	Aug 3, 2021
Grant date	Aug 3, 2021

Abstract

The disclosure relates to a multi-epitope fusion protein as well as to its use as calibrator and/or control in an in vitro diagnostics immunoassay for detecting HCV core antigen. The multi-epitope fusion protein has two to six different non-overlapping linear peptides present in the amino acid sequence of hepatitis C virus (HCV) core protein, wherein each of the peptides is separated from the other peptides by a spacer consisting of a non-HCV amino acid sequence and having a chaperone amino acid sequence. No further HCV specific amino acid sequences are present in the polypeptide. A further aspect relates to a reagent kit for detecting HCV core antigen containing said multi-epitope fusion protein as calibrator or control or both.

First claim

The invention claimed is:

1. A multi-epitope fusion protein comprising: two to six different non-overlapping linear peptides of hepatitis C virus (HCV) core protein, wherein each of said two to six different non-overlapping linear peptides is at least 5 amino acids in length and present one to five-fold, and wherein each of said two to six different non-overlapping linear peptides is separated from the other said peptides by a spacer, wherein the spacer comprises a chaperone peptide specific of a chaperone protein, and wherein no further HCV specific amino acid sequences are present in said multi-epitope fusion protein.

2. A multi-epitope fusion protein according to claim 1 , wherein said HCV core protein corresponds to an amino acid sequence of SEQ ID NO: 1.

3. A multi-epitope fusion protein according to claim 1 , wherein each of said two to six linear peptides has a length of 5 to 50 amino acids.

4. A multi-epitope fusion protein according to claim 1 , wherein three different linear peptides present in the amino acid sequence of said HCV core protein are part of said multi-epitope fusion protein.

5. A multi-epitope fusion protein according to claim 1 , wherein said three different linear peptides comprises a linear amino acid sequence present within amino acid positions 55 to 80 of SEQ ID NO:1, amino acid positions 90 to 120 of SEQ ID NO:1 and amino acid positions 145 to 175 of SEQ ID NO:1.

6. A multi-epitope fusion protein according to claim 1 , having the formula
X-spacer-Y-spacer-Z
wherein X, Y and Z designate different non-overlapping linear peptides of HCV core protein;
wherein Z is present one to five-fold and
wherein spacer designates a chaperone peptide.

7. A multi-epitope fusion protein according to claim 6 , wherein X comprises a linear amino acid sequence present within amino acid positions 55 to 80 of SEQ ID NO:1, Y comprises linear amino acid sequence present within amino acid positions 90 to 120 of SEQ ID NO:1, Z comprises linear amino acid sequences present within amino acid positions 145 to 175 of SEQ ID NO:1.

8. A multi-epitope fusion protein according to claim 1 , wherein said spacer comprises a sequence of at least 25 non-HCV amino acids.

9. A multi-epitope fusion protein according to claim 1 , consisting of any of SEQ ID NOs:4 to 6.

10. A method of producing a soluble multi-epitope fusion protein, said method comprising the steps of
a) culturing host cells transformed with an expression vector comprising operably linked a recombinant DNA molecule encoding a multi-epitope fusion protein according to claim 1 ,
b) expression of said polypeptide and
c) purification of said polypeptide.

11. A reagent kit for the detection of HCV core antigen, comprising in separate containers or in separated compartments of a single container at least one HCV core antigen-specific binding agent and at least one multi-epitope fusion protein according to claim 1 as calibrator material.

12. A reagent kit according to claim 11 , comprising at least two HCV core antigen-specific binding agents, wherein one of said at least two HCV core antigen-specific binding agents is detectably labeled.

13. A reagent kit according to claim 11 , additionally comprising microparticles.

Assignees

• Roche Diagnostics Operations Inc

Inventors

• Upmeier Barbara
• Bollhagen Ralf
• Zarnt Toralf

Classifications

• G01N33/56983

  Viruses · CPC title

• G01N2469/10

  Detection of antigens from microorganism in sample from host · CPC title

• C07K14/005 Primary

  from viruses · CPC title

• C07K2319/35

  containing a fusion for enhanced stability/folding during expression, e.g. fusions with chaperones or thioredoxin · CPC title

• G01N2496/00

  Reference solutions for assays of biological material · CPC title