Methods and compositions for dengue virus vaccines and diagnostics

Patent metadata

Field	Value
Publication number	US-10772948-B2
Application number	US-201615766304-A
Country	US
Kind code	B2
Filing date	Sep 20, 2016
Priority date	Oct 7, 2015
Publication date	Sep 15, 2020
Grant date	Sep 15, 2020

Abstract

The present invention provides compositions directed to recombinant flavivirus E glycoprotein ectodomain dimers for use in diagnostic and immunotherapeutic methods.

First claim

What is claimed is:

1. A method of producing recombinant soluble dengue virus E ectodomain dimers, comprising:
a) preparing a first recombinant soluble monomeric dengue virus E ectodomain comprising a functional first linking moiety at one terminus;
b) contacting the first ectodomain with a second linking moiety that associates with the first linking moiety, wherein the second linking moiety is attached to a solid substrate, thereby attaching the ectodomain to the solid substrate in a specific orientation;
c) contacting the first ectodomain attached to the solid substrate with a second recombinant monomeric dengue virus E ectodomain lacking a functional first linking moiety under conditions whereby dimerization of the first ectodomain and second ectodomain can occur;
d) detaching the recombinant soluble dengue virus E ectodomain dimers from the solid substrate; and
e) collecting the recombinant soluble dengue virus E ectodomain dimers.

2. The method of claim 1 , wherein the first linking moiety and second linking moiety are selected from the group consisting of: 1) a histidine tag (HIS) and Ni 2+ , respectively; 2) biotin and avidin, respectively; and 3) a primary α-helix and a secondary α-helix, respectively.

3. The method of claim 1 , wherein the dengue virus is selected from the group consisting of dengue virus serotype 1 (DENV-1), dengue virus serotype 2 (DENV-2), dengue virus serotype 3 (DENV-3), and dengue virus serotype 4 (DENV-4).

4. A method of producing a recombinant dengue virus E ectodomain dimer attached to a carrier, comprising:
a) preparing a first recombinant monomeric dengue virus E ectodomain comprising a functional first linking moiety at one terminus;
b) contacting the first ectodomain with a second linking moiety that associates with the first linking moiety, wherein the second linking moiety is attached to a carrier, thereby attaching the ectodomain to a solid substrate in a specific orientation; and
c) contacting the first ectodomain attached to the carrier with a second recombinant monomeric dengue virus E ectodomain lacking a functional first linking moiety under conditions whereby dimerization of the first ectodomain and second ectodomain can occur.

5. The method of claim 4 , wherein the first linking moiety and second linking moiety are selected from the group consisting of: 1) a histidine tag (HIS) and Ni 2+ , respectively; 2) biotin and avidin, respectively; and 3) a primary α-helix and a secondary α-helix, respectively.

6. The method of claim 4 , wherein the dengue virus is selected from the group consisting of dengue virus serotype 1 (DENV-1), dengue virus serotype 2 (DENV-2), dengue virus serotype 3 (DENV-3), and dengue virus serotype 4 (DENV-4).

Assignees

• Univ North Carolina Chapel Hill

Inventors

• Metz Stefan
• Desilva Aravinda
• Miley Michael

Classifications

• A61K39/12 Primary

  Viral antigens · CPC title

• A61P31/14

  for RNA viruses · CPC title

• G01N33/56983

  Viruses · CPC title

• C12N2770/24134

  Use of virus or viral component as vaccine, e.g. live-attenuated or inactivated virus, VLP, viral protein · CPC title

• C12N2770/24151

  Methods of production or purification of viral material · CPC title