# US 10,765,711 B2 · Replication competent attenuated vaccinia viruses with deletion of thymidine kinase with and without the expression of human FLT3L or GM-CSF for cancer immunotherapy

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- Number: US 10,765,711 B2
- Document ID: US10765711B2
- Status: Granted
- Kind: B2
- Application number: 16/673,475
- Filed: 2019-11-04
- Published: 2020-07-30
- Granted: 2020-09-08
- Earliest claimed: 2016-02-25
- Assignees: Memorial Sloan Kettering Cancer Center
- Applicants: Memorial Sloan Kettering Cancer Center
- Inventors: Liang Deng; Stewart Shuman; Jedd Wolchok; Taha Merghoub; Weiyi Wang; Peihong Dai; Ning Yang
- Web page: https://patentgrep.com/patent/US10765711B2

## CPC

- A61K 35/768: Oncolytic viruses not provided for in groups A61K35/761 - A61K35/766
- A61K 35/76: Viruses; Subviral particles; Bacteriophages
- A61K 39/3955: {against proteinaceous materials, e.g. enzymes, hormones, lymphokines}
- A61K 48/00: Medicinal preparations containing genetic material which is inserted into cells of the living body to treat genetic diseases; Gene therapy
- A61P 35/00: Antineoplastic agents
- C07K 14/521: {Chemokines}
- C07K 14/535: Granulocyte CSF; Granulocyte-macrophage CSF
- C07K 16/2818: {against CD28 or CD152}
- C07K 16/2827: {against B7 molecules, e.g. CD80, CD86}
- C12N 15/86: Viral vectors
- A61K 2039/505: {comprising antibodies}
- C07K 2317/76: Antagonist effect on antigen, e.g. neutralization or inhibition of binding
- C12N 2710/24121: Viruses as such, e.g. new isolates, mutants or their genomic sequences
- C12N 2710/24132: Use of virus as therapeutic agent, other than vaccine, e.g. as cytolytic agent
- C12N 2710/24133: Use of viral protein as therapeutic agent other than vaccine, e.g. apoptosis inducing or anti-inflammatory
- C12N 2710/24143: viral genome or elements thereof as genetic vector
- C12N 2710/24161: Methods of inactivation or attenuation
- 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 invention relates generally to the fields of oncology, virology and immunotherapy. More particularly, it concerns the use of poxviruses, specifically the replication competent attenuated vaccinia virus with deletion of thymidine kinase (VC-TK−) with and without the expression of human Flt3L or GM-CSF as oncolytic and immunotherapy. The foregoing poxviruses can also be used in combination with immune checkpoint blocking agents. The foregoing poxviruses can also be inactivated via Heat or UV-treatment and the inactivated virus can be used as immunotherapy either alone or in combination with immune checkpoint blocking agents.

## Claims

### Claim 1

A method for eliciting an immune response in a subject in need thereof, the method comprising delivering to the subject a therapeutically effective amount of a composition comprising a recombinant vaccinia virus selected from the group consisting of: (i) E3LΔ83N-TK−-hFlt3L; (ii) E3LΔ83N-TK−; (iii) E3LΔ83N-TK−-GM-CSF; and (iv) combinations thereof, wherein the virus is in replicative or inactivated form.

### Claim 2 (depends on 1)

The method of claim 1, wherein the recombinant vaccinia virus comprises E3LΔ83N-TK−-hFlt3L in replicative or inactivated form.

### Claim 3 (depends on 1)

The method of claim 1, wherein eliciting the immune response in the subject comprises one or more of the following immunological effects selected from:
increased activation of CD8+ T cells;
increased activation of CD4+ T cells; and
increased CD103+ dendritic cells.

### Claim 4 (depends on 1)

The method of claim 1, wherein the recombinant vaccinia virus is delivered to the subject parenterally, intravenously, intratumorally, intraperitoneally, intrathecally, or intracranially.

### Claim 5 (depends on 1)

The method of claim 1, wherein the recombinant vaccinia virus is heat-inactivated.

### Claim 6

A method for treating a malignant solid tumor in a subject in need thereof, the method comprising delivering to the cells of the tumor a recombinant vaccinia virus in replicative or inactivated form selected from the group consisting of: (i) E3LΔ83N-TK−-hFlt3L; (ii) E3LΔ83N-TK−; (iii) E3LΔ83N-TK−-GM-CSF; and (iv) combinations thereof, in an amount effective to induce the immune system of the subject to mount an immune response against the tumor.

### Claim 7 (depends on 6)

The method of claim 6, wherein the recombinant vaccinia virus comprises E3LΔ83N-TK−-hFlt3L in replicative or inactivated form.

### Claim 8 (depends on 6)

The method of claim 6 wherein the tumor is primary or metastatic melanoma, breast carcinoma, or colon carcinoma.

### Claim 9 (depends on 6)

The method of claim 6, wherein the immune response is systemic.

### Claim 10 (depends on 6)

The method of claim 6, wherein the immune response is effective to accomplish one or more of the following: reduction of the size of the tumor; eradication of the tumor; inhibition of growth of the tumor; inhibition of metastasis of the tumor; or reduction or eradication of metastatic tumor.

### Claim 11 (depends on 6)

The method of claim 6, wherein the tumor includes tumor located at the site of delivery, or tumor located at the site of delivery and elsewhere in the body of the subject.

### Claim 12 (depends on 6)

The method of claim 6, wherein the immune response comprises one or more of the following:
increase in cytotoxic CD8+ T cells within the tumor and/or in tumor-draining lymph nodes;
induction of maturation of dendritic cells infiltrating the tumor through induction of type I IFN;
induction of activated CD4+ effector T cells in the subject recognizing tumor cells within the tumor or systemically; and
increase of CD103+ dendritic cells in non-injected tumors of the subject.

### Claim 13 (depends on 6)

The method of claim 6, further comprising conjointly administering an immune checkpoint blocking agent in an amount effective to block immune suppressive mechanisms within the tumor elicited by tumor cells, stromal cells, or tumor infiltrating immune cells.

### Claim 14 (depends on 13)

The method of claim 13, wherein the immune checkpoint blocking agent comprises CTLA-4 inhibitors, CD80 inhibitors, CD86 inhibitors, PD-1 inhibitors, PD-L1 inhibitors, PD-L2 inhibitors, LAG3 inhibitors, B7-H3 inhibitors, B7-H4 inhibitors, TIM3 inhibitors, ICOS inhibitors, II DLBCL inhibitors, BTLA inhibitors, ipilimumab, nivolumab, pembrolizumab, pidilizumab, AMP-224, MPDL3280A, BMS-936559, MEDI4736, MSB 00107180, or any combination thereof.

### Claim 15 (depends on 6)

The method of claim 6, wherein the subject has been previously treated or dosed with an amount of an immune checkpoint blocking agent effective to block immune suppressive mechanisms within the tumor elicited by tumor cells, stromal cells, or tumor infiltrating immune cells.

### Claim 16 (depends on 15)

The method of claim 15, wherein the immune checkpoint blocking agent comprises CTLA-4 inhibitors, CD80 inhibitors, CD86 inhibitors, PD-1 inhibitors, PD-L1 inhibitors, PD-L2 inhibitors, LAG3 inhibitors, B7-H3 inhibitors, B7-H4 inhibitors, TIM3 inhibitors, ICOS inhibitors, II DLBCL inhibitors, BTLA inhibitors, ipilimumab, nivolumab, pembrolizumab, pidilizumab, AMP-224, MPDL3280A, BMS-936559, MEDI4736, MSB 00107180, or any combination thereof.

### Claim 17 (depends on 6)

The method of claim 6, wherein the recombinant vaccinia virus is delivered to the subject parenterally, intravenously, intratumorally, intraperitoneally, intrathecally, or intracranially.

### Claim 18 (depends on 6)

The method of claim 6, further comprising combining the treatment with an additional treatment modality comprising surgery, chemotherapy, targeted therapy, and/or radiation.

### Claim 19 (depends on 6)

The method of claim 6, wherein the recombinant vaccinia virus is heat-inactivated.