# US 5,707,864 A · Nucleic acids encoding mutated human immunodeficiency virus matrix proteins

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- Number: US 5,707,864 A
- Document ID: US5707864A
- Status: Granted
- Kind: A
- Application number: 07/979,966
- Filed: 1992-11-23
- Granted: 1998-01-13
- Earliest claimed: 1992-11-23
- Assignees: President and Fellows of Harvard College
- Inventors: Myron E. Essex; Xiaofang Yu; Tun-Hou Lee
- Web page: https://patentgrep.com/patent/US5707864A

## CPC

- C07K 14/005: from viruses
- A61P 31/12: Antivirals
- A61P 31/18: for HIV
- A61K 38/00: Medicinal preparations containing peptides (peptides containing beta-lactam rings A61K31/00; cyclic dipeptides not having in their molecule any other peptide link than those which form their ring, e.g. piperazine-2,5-diones, A61K31/00; ergot alkaloids of the cyclic peptide type A61K31/48; containing macromolecular compounds having statistically distributed amino acid units A61K31/74; medicinal preparations containing antigens or antibodies A61K39/00; medicinal preparations characterised by the non-active ingredients, e.g. peptides as drug carriers, A61K47/00)
- C12N 2740/16222: New viral proteins or individual genes, new structural or functional aspects of known viral proteins or genes
- Y10S 930/221: Retrovirus related, or human immunodeficiency virus related, or simian immunodeficiency virus related
- C07K: PEPTIDES (peptides in foodstuffs A23; obtaining protein compositions for foodstuffs, working-up proteins for foodstuffs A23J; preparations for medicinal purposes A61K; peptides containing beta-lactam rings C07D; cyclic dipeptides not having in their molecule any other peptide link than those which form their ring, e.g. piperazine-2,5-diones, C07D; ergot alkaloids of the cyclic peptide type C07D519/02; macromolecular compounds having statistically distributed amino acid units in their molecules, i.e. when the preparation does not provide for a specific; but for a random sequence of the amino acid units, homopolyamides and block copolyamides derived from amino acids C08G69/00; macromolecular products derived from proteins C08H1/00; preparation of glue or gelatine C09H; single cell proteins, enzymes C12N; genetic engineering processes for obtaining peptides C12N15/00; compositions for measuring or testing processes involving enzymes C12Q; investigation or analysis of biological material G01N33/00)
- A61P: SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61K: PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES (devices or methods specially adapted for bringing pharmaceutical products into particular physical or administering forms A61J3/00; chemical aspects of, or use of materials for deodorisation of air, for disinfection or sterilisation, or for bandages, dressings, absorbent pads or surgical articles A61L; soap compositions C11D)
- C12N: MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA (microbiological testing media C12Q1/00)
- Y10S: TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS

## Abstract

Nucleic acid constructs encoding mutated human immunodeficiency virus matrix proteins are described. The mutated proteins lower the incorporation of envelope polypeptides in viral particles, disrupt viral assembly or disrupt viral entry into uninfected cells.

## Claims

### Claim 1

A DNA construct comprising nucleic acid encoding a mutated human immunodeficiency virus (HIV) matrix (MA) polypeptide capable of undergoing wild-type gag proteolytic processing, said polypeptide being effective in lowering the incorporation of env polypeptides in the viral particle; disrupting viral assembly; or disrupting viral entry into uninfected cells.

### Claim 2 (depends on 1)

The construct of claim 1 comprising nucleic acid encoding said mutated MA polypeptide, said nucleic acid being in an expressible genetic construction capable of transforming cells.

### Claim 3 (depends on 2)

The construct of claim 2 in which said nucleic acid is part of a viral vector capable of infecting isolated cells.

### Claim 4 (depends on 1)

The construct of claim 1 in which said nucleic acid further comprises a sequence encoding a CD4-binding polypeptide.

### Claim 5 (depends on 1)

The construct of claim 1 in which said nucleic acid further comprises a sequence encoding a gp120-binding polypeptide.

### Claim 6 (depends on 3)

The construct of claim 3, wherein said viral vector is human immunodeficiency virus type I.

### Claim 7

Nucleic acid encoding a mutated MA polypeptide, said mutated MA polypeptide containing a deletion of a region comprising at least one or more of amino acids 11 to 20 and said mutated MA polypeptide being effective to lower the incorporation of env polypeptides in the viral particle.

### Claim 8

Nucleic acid encoding a mutated MA polypeptide, said mutated MA polypeptide containing a deletion of amino acids 21 to 31 of wild-type MA polypeptide, and said mutated MA polypeptide being effective to lower the incorporation of env polypeptides in the viral particle.

### Claim 9

Nucleic acid encoding a mutated MA polypeptide, said mutated MA polypeptide containing a deletion of a region comprising at least one or more of amino acids 32 to 49, and said mutated MA polypeptide being effective to lower the incorporation of env polypeptides in the viral particle.

### Claim 10

Nucleic acid encoding a mutated MA polypeptide, said mutated MA polypeptide containing a deletion of a region comprising at least one or more of amino acids 79 to 90, and said mutated MA polypeptide being effective to lower the incorporation of env polypeptides in the viral particle.

### Claim 11

Nucleic acid encoding a mutated MA polypeptide, said mutated MA polypeptide containing a deletion of a region comprising at least one or more of amino acids 91 to 103, and said mutated MA polypeptide being effective to lower the incorporation of env polypeptides in the viral particle.

### Claim 12

Nucleic acid encoding a mutated MA polypeptide, said mutated MA polypeptide containing a deletion of a region comprising at least one or more of amino acids 105 to 114, and said mutated MA polypeptide being effective to disrupt entry of viral particles into uninfected cells.

### Claim 13

Nucleic acid encoding a mutated MA polypeptide, said mutated MA polypeptide containing a deletion of a region comprising at least one or more of amino acids 116 to 128, and said mutated MA polypeptide being effective to disrupt entry of viral particles into uninfected cells.

### Claim 14

Nucleic acid encoding a mutated MA polypeptide, said mutated MA polypeptide containing a change of amino acids trp36 and ala37 to arg and pro, respectively, and said mutated MA polypeptide being effective to lower the incorporation of env polypeptides in the viral particle.

### Claim 15

Nucleic acid encoding a mutated MA polypeptide, said mutated MA polypeptide containing a change of amino acid ala37 to pro, and said mutated MA polypeptide being effective to lower the incorporation of env polypeptides in the viral particle.

### Claim 16

Nucleic acid encoding a mutated MA polypeptide, said mutated MA polypeptide containing a substitution of thr20 for arg20, and substitution of asn18, 26, 27, 28, 30 and 32 for lys18, 26, 27, 28, 30 and 32, and said mutated MA polypeptide being effective to disrupt viral assembly.

### Claim 17

A nucleic acid construct encoding a mutated HIV matrix (MA) polypeptide, wherein said mutated MA polypeptide contains a deletion of at least one of the following regions of a wild type MA polypeptide:
a) amino acids 5-16 of said wild-type;
b) amino acids 11-20 of said wild-type;
c) amino acids 21-31 of said wild-type;
d) amino acids 32-49 of said wild-type;
e) amino acids 79-90 of said wild-type;
f) amino acids 91-103 of said wild-type;
g) amino acids 105-114 of said wild-type;
h) amino acids 116-128 of said wild-type.

### Claim 18

A nucleic acid construct encoding a mutated HIV matrix (MA) polypeptide, wherein said mutated MA polypeptide contains at least one of the following substitutions of a wild type MA polypeptide:
a) substitution of arg for trp36;
b) substitution of pro for ala37;
c) substitution of thr for arg20;
d) substitution of glu for one or more of lys26, 27, 28, and 32;
e) substitution of asn for one or more of lys18, 26, 27, 28, 30 and 32.

### Claim 19 (depends on 18)

A nucleic acid construct according to claim 18 in which the substitution mutation is selected from the group consisting of:
a) substitution of Arg for Trp36 and substitution of Pro for Ala37 (R3WA);
b) substitution of Pro for Ala37 (R3AP);
d) substitution of Thr for Arg20 and substitution of Glu for each of Lys26, 27, 28, and 32 (B5); and
e) substitution of Thr for Arg20 and substitution of Asn for each of Lys18, 26, 27, 28, 30 and 32 (B8).

### Claim 20 (depends on 17, 18, 19)

A nucleic acid construct according to claim 17, 18 or 19, said substitution being characterized in that said mutated MA polypeptide is effective for at least one of the following:
(a) lowering the incorporation of Env polypeptides in the viral particle;
(b) disrupting viral assembly; or
(c) disrupting viral entry into uninfected cells.

### Claim 21

A composition comprising cells containing nucleic acid encoding a mutated MA polypeptide, said nucleic acid being in an expressible genetic construction, wherein said mutated MA polypeptide is effective in lowering the incorporation of env polypeptides in the viral particle, disrupting viral assembly, or disrupting viral entry into uninfected cells.

### Claim 22 (depends on 21)

The composition of claim 21 wherein said nucleic acid comprises a viral vector capable of infecting said cells.

### Claim 23 (depends on 21)

The composition of claim 21 wherein said nucleic acid further comprises a sequence capable of encoding a CD4-binding polypeptide.

### Claim 24 (depends on 21)

The composition of claim 21 wherein said nucleic acid further comprises a sequence encoding a gp120-binding polypeptide.