Recombinant HIV-1 envelope proteins and their use

Patent metadata

Field	Value
Publication number	US-10400015-B2
Application number	US-201515508885-A
Country	US
Kind code	B2
Filing date	Sep 4, 2015
Priority date	Sep 4, 2014
Publication date	Sep 3, 2019
Grant date	Sep 3, 2019

Abstract

HIV-1 Env ectodomain trimers stabilized in a prefusion mature closed conformation and methods of their use and production are disclosed. In several embodiments, the HIV-1 Env ectodomain trimers and/or nucleic acid molecules can be used to generate an immune response to HIV-1 in a subject. In additional embodiments, the therapeutically effective amount of the HIV-1 Env ectodomain trimers can be administered to a subject in a method of treating or preventing HIV-1 infection.

First claim

We claim:

1. An isolated immunogen, comprising:
a recombinant HIV-1 Env ectodomain trimer stabilized in a prefusion mature closed conformation by one or more amino acid substitutions compared to a native HIV-1 Env sequence, wherein the one or more amino acid substitutions comprise cysteine substitutions to introduce a non-natural disulfide bond between at HIV-1 Env positions 201 and 433 that stabilizes the HIV-1 Env ectodomain trimer in the prefusion mature closed conformation, wherein the HIV-1 Env positions correspond to a HXB2 reference sequence set forth as SEQ ID NO: 1; wherein
the recombinant HIV-1 Env ectodomain trimer comprises three gp120-gp41 protomers comprising a gp120 polypeptide and a gp41 ectodomain; and
the recombinant HIV-1 Env ectodomain trimer does not specifically bind to a 17b antibody when incubated with a molar excess of sCD4.

2. The immunogen of claim 1 , wherein:
the N-terminal residue of the gp120 polypeptide is one of HIV-1 Env positions 1-35;
the C-terminal residue of the gp120 polypeptide is one of HIV-1 Env positions 503-512;
the N-terminal residue of the gp41 ectodomain is one of HIV-1 Env positions 512-522; and/or
the C-terminal residue of the gp41 ectodomain is one of HIV-1 Env positions 624-705, and
wherein the HIV-Env positions correspond to a HXB2 reference sequence set forth as SEQ ID NO: 1.

3. The immunogen of claim 2 , wherein the gp120 polypeptide comprises or consists of HIV-1 Env positions 31-507, and the gp41 ectodomain comprises or consists of HIV-1 Env positions 512-664, wherein the HIV-Env positions correspond to a HXB2 reference sequence set forth as SEQ ID NO: 1.

4. The immunogen of claim 1 , wherein the gp120 polypeptide and the gp41 ectodomain comprise amino acid sequences of HIV-1 Env positions 31-511 and 512-664 (HXB2 numbering), respectively, of the HIV-1 Env amino acid sequence set forth as SEQ ID NO: 2 (BG505), SEQ ID NO: 51 (CAP256.SU), SEQ ID NO: 81 (BB201.B42), SEQ ID NO: 107 (KER2018.11), SEQ ID NO: 174 (CH070.1), SEQ ID NO: 745 (ZM233.6), SEQ ID NO: 746 (Q23.17), SEQ ID NO: 747 (A244), SEQ ID NO: 2114 (T250-4), or SEQ ID NO: 748 (WITO.33), further comprising the one or more amino acid substitutions that stabilize the HIV-1 Env ectodomain trimer in the prefusion mature closed conformation, or sequences at least 80% identical thereto that comprise the one or more one amino acid substitutions that stabilize the recombinant HIV-1 Env ectodomain in the prefusion mature closed conformation.

5. The immunogen of claim 1 , wherein the recombinant HIV-1 Env ectodomain trimer is a chimera comprising amino acid sequences from at least two HIV-1 strains, wherein
the gp41 ectodomain, an N-terminal region of the gp120 polypeptide comprising a β-4 strand, and a C-terminal region of the gp120 polypeptide comprising a β26 strand, are from a first strain of HIV-1; and
the remaining sequence of the gp120 polypeptide are from a heterologous strain of HIV-1; and
the sequences of the first and heterologous strains have been modified to comprise one or more amino acid substitutions that stabilize the recombinant HIV-1 Env ectodomain trimer in the prefusion mature closed conformation.

6. The immunogen of claim 1 , wherein the recombinant HIV-1 Env ectodomain trimer is a chimera comprising amino acid sequences from at least two HIV-1 strains, wherein the recombinant HIV-1 Env ectodomain trimer comprises a V1V2 domain of an HIV-1 Env protein from a heterologous HIV-1 strain, and the remainder of the recombinant HIV-1 Env ectodomain is from a first strain of HIV-1.

7. The immunogen of claim 1 , wherein the recombinant HIV-1 Env ectodomain trimer is a chimera comprising amino acid sequences from at least three HIV-1 strains, wherein
the gp41 ectodomain, an N-terminal region of the gp120 polypeptide comprising a β-4 strand, and a C-terminal region of the gp120 polypeptide comprising a β26 strand, are from a first strain of HIV-1;
a V1V2 domain of the gp120 polypeptide is from a second strain of HIV-1; and
the remaining sequence of the gp120 polypeptide is from a heterologous strain of HIV-1; and
the sequences of the first, second, and heterologous strains have been modified to comprise the one or more amino acid substitutions that stabilize the recombinant HIV-1 Env ectodomain trimer in the prefusion mature closed conformation.

8. The immunogen of claim 1 , wherein the one or more amino acid substitutions that stabilize the HIV-1 Env ectodomain trimer in the prefusion mature closed conformation further comprise:
one of the following substitutions: 66P, 68P, 70F, 70Y, 71P, 75F, 75W, 75M, 111F, 111Y, 112I, 120T, 154W, 154F, 159Y, 164F, 172F, 202F, 204W, 204F, 208F, 208W, 223W, 302F, 307F, 307W, 309W, 315F, 316W, 323Y, 423F, 427M, 429N, 430W, 430F, 432F, 432W, 436W, 544Y, 580L, 583L, and 594N; or
one of the following sets of substitutions: 154F, 300L, 302M, and 320L substitutions,
159Y and 323Y substitutions, 544Y and 537Y substitutions, 120F and 315M substitutions, 177W and 420M substitutions, 580L and 583L substitutions, 573T and 594N substitutions, and 304C and 440C substitutions: and
wherein the HIV-Env positions correspond to a HXB2 reference sequence set forth as SEQ ID NO: 1.

9. The immunogen of claim 1 , wherein the one or more amino acid substitutions that stabilize the HIV-1 Env ectodomain trimer in the prefusion mature closed conformation further comprise:
(a) a non-natural disulfide bond between cysteine substitutions at HIV-1 Env positions 501 and 605;
(b) a proline substitution at HIV-1 Env position 559; or
(c) a combination of (a) and (b); and
wherein the HIV-Env positions correspond to a HXB2 reference sequence set forth as SEQ ID NO: 1.

10. The immunogen of claim 1 , wherein:
the recombinant HIV-1 Env ectodomain further comprises one or more amino acid substitutions that introduce an N-linked glycosylation site at one of HIV-1 Env position 33, 35, 502, 504, 658, or 661, or a combination of two or more thereof; and wherein the HIV-Env positions correspond to a HXB2 reference sequence set forth as SEQ ID NO: 1.

11. The immunogen of claim 1 , wherein the gp120-gp41 protomers are single chain HIV-1 Env ectodomains, and wherein the N-terminal residue of the gp120 polypeptide is linked to the C-terminal residue of the gp41 ectodomain by a heterologous peptide linker.

12. The immunogen of claim 11 , wherein
the heterologous peptide linker joins gp120 and gp41 at HIV-1 -Env positions 507 and 512, 503 and 519, 504 and 519, 503 and 522, or 504 and 522, respectfully, wherein the HIV-Env positions correspond to a HXB2 reference sequence set forth as SEQ ID NO: 1.

13. The immunogen of claim 1 , wherein the gp120-gp41 protomers in the recombinant HIV-1 Env ectodomain trimer are linked to a transmembrane domain.

14. The immunogen of claim 9 , comprising the combination of
cysteine substitutions at positions 501 and 605 to introduce a disulfide bond in the HIV-1 Env ectodomain trimer; and
a proline substitution at position 559.

15. The immunogen of claim 1 , wherein the gp120-gp41 protomers in the recombinant HIV-1 Env ectodomain trimer comprise the amino acid sequence set forth as SEQ ID NO: 26.

16. The immunogen of claim 1 , further comprising a D368R substitution, an N-linked glycosylation site at HIV-1 Env position 332, an R6 substitution, or a combination thereof, wherein the HIV-Env positions correspond to a HXB2 reference sequence set forth as SEQ ID NO: 1.

17. The immunogen of claim 1 , wherein the gp120-gp41 protomers in the recombinant HIV-1 Env ectodomain trimer comprise:
the amino acid sequence set fort

Assignees

• Us Health

Inventors

• Kwong Peter
• Pancera Marie
• Zhou Tongqing
• Georgiev Ivelin
• Joyce Michael Gordon
• Acharya Priyamvada
• Gorman Jason
• Yang Yongping
• Druz Aliaksandr
• Stewart-Jones Guillaume
• Chen Rita
• Chuang Gwo-Yu
• Baxa Ulrich
• Mascola John
• Lynch Rebecca
• Zhang Baoshan
• Cheng Cheng

Classifications

• C07K16/1145

  Env proteins, e.g. gp41, gp110/120, gp160, V3, principal neutralising domain [PND] or CD4-binding site · CPC title

• C12N2740/16034

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

• C12N2740/16022

  New viral proteins or individual genes, new structural or functional aspects of known viral proteins or genes · CPC title

• C07K14/005

  from viruses · CPC title

• A61K2039/55577

  Saponins; Quil A; QS21; ISCOMS · CPC title