# US 11,364,306 B2 · Detargeted adenovirus variants and related methods

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- Number: US 11,364,306 B2
- Document ID: US11364306B2
- Status: Granted
- Kind: B2
- Application number: 16/460,160
- Filed: 2019-07-02
- Published: 2019-12-26
- Granted: 2022-06-21
- Earliest claimed: 2015-01-20
- Assignees: Adcure Biotechnologies, LLC.
- Applicants: Adcure Biotechnologies, LLC.
- Inventors: Dmitry M. Shayakhmetov; Nelson C. Di Paolo
- Web page: https://patentgrep.com/patent/US11364306B2

## CPC

- A61K 48/00: Medicinal preparations containing genetic material which is inserted into cells of the living body to treat genetic diseases; Gene therapy
- A61K 35/76: Viruses; Subviral particles; Bacteriophages
- A61K 35/761: Adenovirus
- A61P 43/00: Drugs for specific purposes, not provided for in groups A61P1/00-A61P41/00
- C07K 14/005: from viruses
- C07K 14/075: Adenoviridae
- C12N 7/00: Viruses; Bacteriophages; Compositions thereof; Preparation or purification thereof (preparing medicinal viral antigen or antibody compositions, e.g. virus vaccines, A61K39/00)
- C12N 7/06: {Inactivation or attenuation} by chemical treatment
- C12N 15/86: Viral vectors
- C12N 15/8616: {Special methods for targeting systems}
- C12N 2710/00043: viral genome or elements thereof as genetic vector
- C12N 2710/00061: Methods of inactivation or attenuation
- C12N 2710/10021: Viruses as such, e.g. new isolates, mutants or their genomic sequences
- C12N 2710/10322: New viral proteins or individual genes, new structural or functional aspects of known viral proteins or genes
- C12N 2710/10332: Use of virus as therapeutic agent, other than vaccine, e.g. as cytolytic agent
- C12N 2710/10333: Use of viral protein as therapeutic agent other than vaccine, e.g. apoptosis inducing or anti-inflammatory
- C12N 2710/10334: Use of virus or viral component as vaccine, e.g. live-attenuated or inactivated virus, VLP, viral protein
- C12N 2710/10345: Special targeting system for viral vectors
- C12N 2810/405: Vectors comprising RGD peptide
- 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)
- A61P: SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- 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)
- C12N: MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA (microbiological testing media C12Q1/00)

## Abstract

The present disclosure describes the generation and the use of Ad variants (Ad) possessing any combination of mutations in genes that code for the hexon, penton, fiber, and non-structural proteins, where simultaneous modification of hexon and penton are made to avoid the trapping of Ad in the liver and to reduce toxicity after intravascular virus administration. Such liver de-targeted Ad can be useful tool for selective and specific gene delivery to extra-hepatic tissues and cells, including disseminated metastatic cancer cells.

## Claims

### Claim 1

A method of delivering a gene to a non-hepatic mammalian cell in a subject, comprising the steps of:
administering to the subject an effective amount of an adenovirus comprising:
a mutation in the RGD motif of the penton protein, wherein the mutation causes reduced binding of β3 integrins of a host cell;
a mutation in the HVR1 region of the hexon protein, wherein the mutation causes reduced virus trapping in Kupffer cells; and
a mutation in the HVR7 region of the hexon protein,
wherein the combination of the mutation in the RGD motif, the mutation in the HVR1 region and the mutation in the HVR7 region causes reduced liver sequestration of the adenovirus.

### Claim 2 (depends on 1)

The method of claim 1, wherein the RGD motif of the penton protein is substituted with a non-RGD motif-containing peptide, capable of binding to non-β3 cellular integrins.

### Claim 3 (depends on 1)

The method of claim 1, wherein the adenovirus is a species B, C, or D adenovirus.

### Claim 4 (depends on 1)

The method of claim 1, wherein the adenovirus is serotype 5 or 2.

### Claim 5 (depends on 1)

The method of claim 1, wherein the adenovirus further comprises a mutation in the HVR3 or HVR5 region of the hexon protein.

### Claim 6 (depends on 5)

The method of claim 5, wherein the mutation in the HVR3 or HVR5 region of the hexon protein is a substitution.

### Claim 7 (depends on 1)

The method of claim 1, wherein the adenovirus comprises a single virus genomic DNA molecule comprising a nucleotide sequence that encodes SEQ ID NO: 2.

### Claim 8 (depends on 1)

The method of claim 1, wherein the mutation in the HVR1 region of the hexon protein is a substitution.

### Claim 9 (depends on 1)

The method of claim 1, wherein the adenovirus comprises a gene of interest that encodes a polypeptide selected from the group consisting of cytokines, cellular receptors, nuclear receptors, ligands, coagulation factors, CFTR proteins, insulins, dystrophins, growth hormones, immune-stimulatory or immune-suppressing proteins of eukaryotic or prokaryotic origin, enzymes, enzyme inhibitors, polypeptides with antitumor effect, polypeptides capable of inhibiting a bacterial, parasitic or viral infection, antibodies, toxins, immunotoxins, ribozymes, and markers.

### Claim 10 (depends on 1)

The method of claim 1, wherein the adenovirus comprises a single virus genomic DNA molecule comprising a nucleotide sequence that encodes SEQ ID NO:3 or SEQ ID NO:4.

### Claim 11 (depends on 1)

The method of claim 1, wherein the adenovirus is administered intravenously.

### Claim 12 (depends on 1)

The method of claim 1, wherein the adenovirus is a replication-competent adenovirus.

### Claim 13 (depends on 1)

The method of claim 1, wherein the adenovirus comprises a gene encoding a shRNA or a LncRNA.