# US 8,785,611 B2 · Compositions and methods related to mRNA translational enhancer elements

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- Number: US 8,785,611 B2
- Document ID: US8785611B2
- Status: Granted
- Kind: B2
- Application number: 12/316,581
- Filed: 2008-12-11
- Published: 2009-09-10
- Granted: 2014-07-22
- Earliest claimed: 2007-12-11
- Assignees: The Scripps Research Institute
- Applicants: Vincent P. Mauro; Gerald M. Edelman; Wei Zhou
- Inventors: Vincent P. Mauro; Gerald M. Edelman; Wei Zhou
- Web page: https://patentgrep.com/patent/US8785611B2

## CPC

- C12N 15/85: for animal cells
- A61P 43/00: Drugs for specific purposes, not provided for in groups A61P1/00-A61P41/00
- C12N 15/1051: {Gene trapping, e.g. exon-, intron-, IRES-, signal sequence-trap cloning, trap vectors}
- C12N 15/67: General methods for enhancing the expression
- C12N: MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA (microbiological testing media C12Q1/00)
- A61P: SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS

## Abstract

Provided are mRNA translational enhancer elements (TEEs), e.g., SEQ ID NOs:1-35. Also provided are translational enhancer polynucleotides that comprise one or more of the specific TEEs exemplified herein or their variants, homologs or functional derivatives. Further provided are expression vectors comprising such TEEs or translational enhancer polynucleotides, as well as host cells and expression systems that harbor such vectors.

## Claims

### Claim 1

An isolated polynucleotide sequence comprising at least two mRNA translational enhancer elements (TEE), wherein at least one TEE consists of a full-length sequence that is selected from the group consisting of 5′-CGCGGCTGA-3′ (SEQ ID NO: 31), 5′-AGCCGCCGCA-3′ (SEQ ID NO: 34) and 5′-ACGCCGCCGA-3′ (SEQ ID NO: 35).

### Claim 2 (depends on 1)

The polynucleotide sequence of claim 1 which comprises at least 5 copies of the TEE.

### Claim 3 (depends on 1)

The polynucleotide sequence of claim 1 which comprises at least 10 copies of the TEE.

### Claim 4

A vector for recombinantly expressing a polypeptide in a eukaryotic cell, comprising a eukaryotic promoter operably linked to a polynucleotide sequence comprising at least one mRNA translational enhancer element (TEE), wherein the TEE is selected from the group consisting of 5′-CGCGGCTGA-3′ (SEQ ID NO: 31), 5′-AGCCGCCGCA-3′ (SEQ ID NO: 34) and 5′-ACGCCGCCGA-3′ (SEQ ID NO: 35).

### Claim 5 (depends on 4)

The vector of claim 4, wherein the mRNA translational enhancer element is located 3 to the promoter and adapted for directional ligation of a second polynucleotide sequence encoding a polypeptide of interest.

### Claim 6 (depends on 4)

The vector of claim 4, further comprising a second polynucleotide sequence encoding a polypeptide of interest which is operably linked to the mRNA translational enhancer element.

### Claim 7 (depends on 6)

The vector of claim 6, wherein the mRNA translational enhancer element is located in the 5′ leader sequence of the second polynucleotide sequence.

### Claim 8 (depends on 4)

A DNA vaccine comprising the vector of claim 4.

### Claim 9 (depends on 4)

An isolated eukaryotic host cell transfected with the vector of claim 4.

### Claim 10 (depends on 9)

The host cell of claim 9, wherein the host cell is a CHO cell.

### Claim 11

A method for recombinantly producing a polypeptide comprising:
(i) constructing an expression vector which comprises a eukaryotic promoter and a mRNA translational enhancer element that are each operably linked to a polynucleotide encoding the polypeptide, wherein the TEE is selected from the group consisting of 5′-CGCGGCTGA-3′ (SEQ ID NO: 31), 5′-AGCCGCCGCA-3′ (SEQ ID NO: 34) and 5′-ACGCCGCCGA-3′ (SEQ ID NO: 35);
(ii) transfecting the expression vector into a eukaryotic host cell; and
(iii) culturing the host cell transfected with the expression vector; thereby producing the polypeptide.

### Claim 12 (depends on 11)

The method of claim 11, further comprising purifying the recombinant polypeptide.

### Claim 13 (depends on 11)

The method of claim 11, wherein the host cell is CHO cell.

### Claim 14 (depends on 11)

The method of claim 11, wherein the polypeptide is a therapeutic protein.