Recombinant adenovirus-based interferon biotherapeutics in swine
Inventors
Zhu, James J. • Bishop, Elizabeth A. • Ramanathan, Palaniappan
Assignees
US Department of Agriculture USDA
Publication Number
US-11582956-B2
Publication Date
2023-02-21
Expiration Date
2039-09-03
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Abstract
Disclosed herein is a recombinant adenovirus genome, said adenovirus genome comprising a heterologous nucleic acid inserted into a cloning site of said genome, said heterologous nucleic acid comprising: (a) a first nucleic acid sequence comprising an adenovirus tripartite sequence (e.g., SEQ ID NO:1) operably linked to a second nucleic acid sequence encoding an interferon (e.g., SEQ ID NO:2); (b) a third nucleic acid sequence comprising a bovine growth hormone polyA termination sequence operably linked to said second nucleic acid sequence (e.g., SEQ ID NO:3); (c) a fourth nucleic acid sequence comprising a porcine elongation factor 1-alpha (EF1α) promoter (e.g., SEQ ID NO:4); (d) a fifth nucleic acid sequence operably linked to said fourth nucleic acid sequence, said fifth nucleic acid sequence encoding a suppressor of cytokine signaling 1 (SOCS1) protein (e.g., SEQ ID NO:5). Furthermore, there is disclosed a method of producing interferon in an animal (e.g., swine).
Core Innovation
The invention disclosed is a recombinant adenovirus genome that comprises a heterologous nucleic acid inserted into a cloning site. This nucleic acid includes an adenovirus tripartite sequence operably linked to a nucleic acid sequence encoding an interferon, a bovine growth hormone polyA termination sequence linked to the interferon sequence, a porcine elongation factor 1-alpha (EF1α) promoter, and a nucleic acid sequence encoding a suppressor of cytokine signaling 1 (SOCS1) protein linked to the EF1α promoter. The genome may be codon-optimized for bacterial expression and can be incorporated into host cells for interferon production both in vitro and in vivo. The invention further discloses methods of producing interferon in animals, especially swine, by introducing an effective amount of the recombinant adenovirus, as well as immunomodulatory compositions containing the recombinant virus.
The problem addressed arises from the foot-and-mouth disease virus (FMDV), which causes acute disease in various cloven-hoofed animals and poses serious economic threats to livestock industries. Although commercial vaccines are available, they require about a week to induce protective immunity, creating a need for countermeasures with rapid onset of immunity. Previous adenovirus-based biotherapeutics expressing type I interferons protected pigs but required doses approximately 100 times higher than vaccines and showed protection lasting less than a week, limiting field applicability. There is thus a need for feasible biotherapeutics that induce both rapid and long-lasting protection against FMDV.
The invention builds on identifying the most potent porcine interferon gene for therapeutic use, incorporating the adenovirus tripartite non-coding sequence to enhance expression, and utilizing the porcine EF1α promoter combined with SOCS1 gene to increase and sustain interferon expression. Experimental results demonstrate that the new recombinant adenovirus exhibits more than 20-fold increased potency in inducing anti-FMDV activity relative to earlier versions, both in vitro and in vivo. This integrated approach addresses the shortcomings of high dose requirements and short duration of protection present in prior adenovirus-based interferon therapies, providing a significantly improved biotherapeutic for controlling FMDV in swine.
Claims Coverage
The patent includes one independent claim for a recombinant adenoviral vector and multiple claims directed to methods and compositions involving this vector. The main inventive features relate to the vector's genetic composition and its functional application in interferon production.
Recombinant adenoviral vector comprising interferon and SOCS1 genes
A recombinant adenoviral vector whose genome comprises a nucleic acid sequence encoding interferon (IFN) and a nucleic acid sequence encoding suppressor of cytokine signaling 1 (SOCS1) operably linked to a porcine elongation factor 1-alpha (EF1α) promoter.
Codon optimization of nucleic acid sequences for bacterial expression
The nucleic acid sequences encoding interferon and SOCS1 proteins are codon-optimized for expression in bacteria, specifically Escherichia coli.
Genomic organization placing SOCS1 downstream of interferon
The nucleic acid sequence encoding SOCS1 operably linked to the porcine EF1α promoter is positioned 3′ to the nucleic acid sequence encoding interferon in the adenoviral genome.
Use of specific nucleic acid sequences
The adenoviral genome comprises the nucleic acid sequence of SEQ ID NO:6, encoding interferon (SEQ ID NO:2), the EF1α promoter (SEQ ID NO:4), and SOCS1 (SEQ ID NO:5).
Replication-competent vector elements and host cells
The recombinant adenoviral vector includes vector sequences allowing adenovirus replication in host cells, including bacterial cells such as E. coli, and isolated host cells comprising the vector.
Methods of interferon production in animals and tissue culture
Methods involve introducing the adenoviral vector into swine or cells in tissue culture to produce interferon, with administration routes including intramuscular, subcutaneous, oral, or intranasal inoculation and use with veterinary or pharmaceutical carriers.
Immunomodulatory composition
An immunomodulatory composition comprising the adenoviral vector in a veterinary or pharmaceutically acceptable carrier is included.
The claims cover a recombinant adenoviral vector engineered to express interferon alongside SOCS1 under a porcine EF1α promoter, optimized for bacterial expression and adenoviral replication, and methods and compositions to produce interferon in animals and culture using this vector.
Stated Advantages
The recombinant adenovirus biotherapeutic induces a significantly higher anti-FMDV activity—more than 20-fold—compared to prior adenovirus-based interferon therapies.
It provides rapid onset and longer-lasting protection against foot-and-mouth disease virus in swine.
Inclusion of the adenovirus tripartite non-coding sequence and SOCS1 gene enhances interferon expression and antiviral potency.
Use of a porcine EF1α promoter increases transcriptional activity in porcine cells.
Documented Applications
Rapid induction of antiviral interferon in swine to protect against foot-and-mouth disease virus infection during outbreaks.
Production of interferon protein in animals, especially pigs, by administration of recombinant adenovirus-based vectors.
In vitro production of interferon using host cells comprising the recombinant adenovirus genome.
Use of the recombinant adenovirus vector in immunomodulatory compositions for veterinary or pharmaceutical applications targeting FMDV.
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