Granted Patent B2

US 12,460,193 · App. 18/914,016 · Granted Nov 4, 2025

Inventors: Cheng Liu (Orinda, CA); Hongbing Zhang (Moraga, CA); Ziyou Cui (Seattle, WA); Zhiyuan Yang (Richmond, CA); Jinyun Chen (Emeryville, CA); Jingbao Liu (Castro Valley, CA); Guangyan Xiong (Emeryville, CA); Warner Greene (Hillsborough, CA)

Assignee: INVISISHIELD TECHNOLOGIES LTD.

C12N9/485A61P31/14C12Y304/17023A61K38/00C07K2319/00

Abstract

Claims (49)

1 . A chimeric protein comprising:

(a) a target-binding moiety comprising an extracellular binding domain (EBD) of an angiotensin-converting enzyme 2 (ACE2) protein or a fragment or a variant thereof that specifically binds to a spike(S) protein; and

(b) a mucoadhesive peptide fragment comprising at least 5 positively charged amino acid residues, wherein the mucoadhesive peptide fragment does not comprise six consecutive histidines; and

wherein the mucoadhesive peptide fragment facilitates attachment of the chimeric protein to a mucosa.

2 . The chimeric protein of claim 1 , wherein the chimeric protein comprises:

(i) a single polypeptide chain; or

(ii) two or more polypeptide chains, and wherein the chimeric protein comprises two or more mucoadhesive peptide fragments.

3 . The chimeric protein of claim 1 , wherein the mucoadhesive peptide fragment comprises at least 6 positively charged amino acid residues.

4 . The chimeric protein of claim 1 , wherein the positively charged amino acid residues are selected from the group consisting of lysine, arginine, histidine, ornithine, and combinations thereof.

5 . The chimeric protein of claim 1 , wherein:

(i) the mucoadhesive peptide fragment comprises at least 5 contiguous positively charged amino acid residues;

(ii) the positively charged amino acid residues are interspersed with one or more non-positively charged amino acid residues;

(iii) the mucoadhesive peptide fragment is no more than about 15 kD; and/or

(iv) the mucoadhesive peptide fragment has an isoelectric point (pI) higher than the pH of the mucosa.

6 . The chimeric protein of claim 1 , wherein the mucoadhesive peptide fragment comprises an amino acid sequence of any one of SEQ ID NOs: 31-62 and 128-134, or a variant thereof having at least about 90% sequence identity to the amino acid sequence of any one of SEQ ID NOs: 31-62 and 128-134.

7 . The chimeric protein of claim 1 , wherein the mucoadhesive peptide fragment is fused to the target-binding moiety via a peptide linker.

8 . The chimeric protein of claim 1 , wherein the mucoadhesive peptide fragment is fused to a C-terminus of the target-binding moiety.

9 . The chimeric protein of claim 1 , wherein the target-binding moiety comprises the EBD of a human ACE2 (hACE2) protein or a fragment or a variant thereof that specifically binds to a spike(S) protein.

10 . The chimeric protein of claim 9 , wherein the target-binding moiety comprises:

(a) (i) amino acids 30-41 of a full-length hACE2 protein, or a variant thereof having at least about 90% sequence identity to amino acids 30-41 of a full-length hACE2 protein; and/or

(ii) the amino acid sequence of SEQ ID NO: 102, or a variant thereof having at least about 90% sequence identity to the amino acid sequence of SEQ ID NO: 102;

(b) (i) amino acids 24-42 of a full-length hACE2 protein, or a variant thereof having at least about 90% sequence identity to amino acids 24-42 of a full-length hACE2 protein; and/or

(ii) The amino acid sequence of SEQ ID NO: 8, or a variant thereof having at least about 90% sequence identity to the amino acid sequence of SEQ ID NO: 8; or

(c) the amino acid sequence of any one of SEQ ID NOs: 1-7, 9-14, and 135, or a variant thereof having at least about 90% sequence identity to the amino acid sequence of any one of SEQ ID NOs: 1-7, 9-14, and 135.

11 . The chimeric protein of claim 10 , wherein the chimeric protein comprises the amino acid sequence of any one of SEQ ID NOs: 84, 86-88, 136, and 138-140, or a variant thereof having at least about 90% sequence identity to the amino acid sequence of any one of SEQ ID NOs: 84, 86-88, 136, and 138-140.

12 . The chimeric protein of claim 1 , wherein the target-binding moiety comprises the EBD of an animal ACE2 protein or a fragment or a variant thereof that specifically binds to a spike (S) protein.

13 . The chimeric protein of claim 12 , wherein the target-binding moiety comprises:

(a) (i) amino acids 30-41 of a full-length animal ACE2 protein, or a variant thereof having at least about 90% sequence identity to amino acids 30-41 of a full-length animal ACE2 protein, wherein the full-length animal ACE2 protein is not a chicken or canine ACE2 protein; or

(ii) amino acids 29-40 of a full-length animal ACE2 protein, or a variant thereof having at least about 90% sequence identity to amino acids 29-40 of a full-length animal ACE2 protein, wherein the full-length animal ACE2 protein is a chicken or canine ACE2 protein; or

(b) the amino acid sequence of any one of SEQ ID NOs: 15-27 and 110-122, or a variant thereof having at least about 90% sequence identity to any one of SEQ ID NOs: 15-27 and 110-122.

14 . The chimeric protein of claim 13 , wherein the chimeric protein comprises the amino acid sequence of any one of SEQ ID NOs: 90-93, or a variant thereof having at least about 90% sequence identity to the amino acid sequence of any one of SEQ ID NOs: 90-93.

15 . A pharmaceutical composition comprising the chimeric protein of claim 1 , and a pharmaceutically acceptable carrier.

16 . The chimeric protein of claim 12 , wherein the animal ACE2 protein is a murine, guinea pig, equine, ferret, macaque, chimpanzee, swine, canine, feline, bovine, rabbit, mink, or chicken ACE2 protein or a fragment or a variant thereof.

17 . The chimeric protein of claim 1 , wherein the S protein is a coronavirus S protein, and wherein the coronavirus is selected from the group consisting of SARS-COV, SARS-COV-2, and HCoV-NL63.

18 . The chimeric protein of claim 17 , wherein the S protein comprises the amino acid sequence of any one of SEQ ID NOs: 96-100, 103, 123-127, and 141-143, or a variant thereof having at least about 90% sequence identity to the amino acid sequence of any one of SEQ ID NOs: 96-100, 103, 123-127, and 141-143.

19 . The chimeric protein of claim 7 , wherein the peptide linker comprises an oligomerization or multimerization domain.

20 . The chimeric protein of claim 19 , wherein the oligomerization or multimerization domain is an Fc region or a fragment thereof.

21 . An isolated nucleic acid or a vector encoding the chimeric protein of claim 1 .

22 . A host cell expressing the chimeric protein of claim 1 .

23 . A method of preparing a chimeric protein, comprising:

(a) culturing the host cell of claim 22 under a condition effective to express the chimeric protein; and

(b) obtaining the expressed chimeric protein from the host cell.

24 . A method of preventing or treating an infection caused by a virus in an individual, comprising administering to the individual an effective amount of the pharmaceutical composition of claim 15 , wherein the chimeric protein specifically binds to a spike(S) protein of the virus.

25 . An in vitro method of killing or neutralizing a virus, comprising contacting the virus with the chimeric protein of claim 1 in the presence of at least one component of the complement system, wherein the chimeric protein specifically binds to a spike(S) protein of the virus.

26 . A method of killing or neutralizing a virus in an individual, comprising administering to the individual an effective amount of the pharmaceutical composition of claim 15 , wherein the chimeric protein specifically binds to a spike(S) protein of the virus.

27 . A method of activating the complement pathway in an individual, comprising administering to the individual an effective amount of the pharmaceutical composition of claim 15 .

28 . The method of claim 24 , wherein the virus is a coronavirus.

29 . The method of claim 28 , wherein the S protein comprises the amino acid sequence of any one of SEQ ID NOs: 96-100, 103, 123-127, and 141-143, or a variant thereof having at least about 90% sequence identity to the amino acid sequence of any one of SEQ ID NOs: 96-100, 103, 123-127, and 141-143.

30 . The method of claim 28 , wherein the coronavirus is selected from the group consisting of SARS-COV, SARS-COV-2, and HCoV-NL63.

Assignments (4)

Continuity (4)

References Cited (176)