Multitargeting onocolytic adenovirus, methods of use, and methods of making
AssigneeThe UAB Research Foundation
InventorGeorge Clement Dobbins
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Abstract
To increase the therapeutic potential of these oncolytic viruses based on a 24 base pair deletion in the viral E1 A gene (D24), a conditionally replicating adenovirus targeting multiple receptors upregulated on tumors was generated by incorporating an Ad5/3 fiber with a carboxyl terminus RGD ligand. The virus displayed full cytopathic effect in tumor lines assayed at low titers with improved cytotoxicity over Ad5-RGD D24, Ad5/3 D24 and an HSV oncolytic virus. The virus was further engineered to deliver immunotherapeutic agents such as GMCSF while maintaining enhanced heterogenic oncolysis.
Claims36
A nucleotide sequence encoding a genetically modified adenovirus, said nucleotide sequence comprising:
- a nucleotide sequence encoding an amino acid sequence comprising an N-terminal Ad5 serotype fiber shaft domain, a Ad3 serotype fiber knob domain, and an arginine-glycine-aspartate (RGD) peptide wherein the peptide RGD is at the C-terminus of the Ad3 serotype fiber knob;
- and
- a nucleotide sequence encoding an adenovirus E1A polypeptide having a 24 base pair deletion compared to a wild-type E1A polypeptide, wherein the 24 base pair deletion in E1A-encoding nucleotide sequence renders the expressed E1A polypeptide unable to bind a pRb transcription factor, and wherein the adenovirus E1A gene encoding the E1A polypeptide is operably linked to an adenovirus E1A promoter.
A nucleotide sequence encoding a genetically modified shuttle adenovirus vector, wherein the nucleotide sequence is selected from the group consisting of SEQ ID NOs.:
- 3, 6, 9, 10, 11, 12, and 13.
A genetically modified adenovirus of the Ad5 serotype, wherein said adenovirus comprises a nucleotide sequence, wherein the nucleotide sequence encodes the adenovirus and comprises:
- a region encoding an amino acid sequence comprising an N-terminal Ad5 serotype fiber shaft domain, a Ad3 serotype fiber knob domain, and an arginine-glycine-aspartate (RGD) peptide wherein the peptide RGD is at the C-terminus of the Ad3 serotype fiber knob;
- and
- a region encoding an adenovirus E1A polypeptide having a 24 base pair deletion compared to a wild-type E1A polypeptide, wherein the 24 base pair deletion in E1A-encoding nucleotide sequence renders the expressed E1A polypeptide unable to bind a pRb transcription factor, and wherein the adenovirus E1A gene encoding the E1A polypeptide is operably linked to an adenovirus E1A promoter.
A method of monitoring the progress of delivery of a genetically modified adenovirus to a tumor in a patient, said method comprising the steps of:
- (a) administering to the patient a pharmaceutically acceptable composition comprising a genetically modified adenovirus of the Ad5 serotype, wherein said adenovirus is encoded by a nucleotide sequence comprising:
- a region encoding an amino acid sequence comprising an N-terminal Ad5 serotype fiber shaft domain, a Ad3 serotype fiber knob domain, and an arginine-glycine-aspartate (RGD) peptide wherein the peptide RGD is at the C-terminus of the Ad3 serotype fiber knob;
- a region encoding an adenovirus E1A polypeptide having a 24 base pair deletion compared to a wild-type E1A polypeptide, wherein the 24 base pair deletion in E1A-encoding nucleotide sequence renders the expressed E1A polypeptide unable to bind a pRb transcription factor, and wherein the adenovirus E1A gene encoding the E1A polypeptide is operably linked to an adenovirus E1A promoter;
- and
- a region encoding a reporter protein,
- wherein the nucleotide sequence region encoding the reporter protein is operably linked to a nucleotide sequence of the adenovirus encoding an adenovirus pIX polypeptide;
- and
- (b) detecting a signal from the reporter protein, wherein the detected signal indicates the presence of the genetically modified adenovirus in cells of a tumor in the patient.
A method of modulating an immune response to a tumor in a patient, said method comprising:
- administering to the patient a pharmaceutically acceptable composition comprising a genetically modified adenovirus of the Ad5 serotype, wherein said adenovirus is encoded by a nucleotide sequence comprising:
- a region encoding an amino acid sequence comprising an N-terminal Ad5 serotype fiber shaft domain, a Ad3 serotype fiber knob domain, and an arginine-glycine-aspartate (RGD) peptide wherein the peptide RGD is at the C-terminus of the Ad3 serotype fiber knob;
- a region encoding an adenovirus E1A polypeptide having a 24 base pair deletion compared to a wild-type E1A polypeptide, wherein the 24 base pair deletion in E1A-encoding nucleotide sequence renders the expressed E1A polypeptide unable to bind a pRb transcription factor, and wherein the adenovirus E1A gene encoding the E1A polypeptide is operably linked to an adenovirus E1A promoter;
- and
- a region encoding a immunomodulator protein,
- wherein the nucleotide sequence region encoding the reporter protein is operably linked to a nucleotide sequence encoding an adenovirus E3 polypeptide or a fragment thereof.
A nucleotide sequence encoding a genetically modified adenovirus, said nucleotide sequence comprising:
- a nucleotide sequence encoding an amino acid sequence comprising an N-terminal Ad5 serotype fiber shaft domain, a Ad3 serotype fiber knob domain, and an arginine-glycine-aspartate (RGD) peptide wherein the peptide RGD is at the C-terminus of the Ad3 serotype fiber knob;
- and
- a nucleotide sequence encoding an adenovirus E1A polypeptide having a 24 base pair deletion compared to a wild-type E1A polypeptide, wherein the 24 base pair deletion in E1A-encoding nucleotide sequence renders the expressed E1A polypeptide unable to bind a pRb transcription factor, wherein the nucleotide sequence is SEQ ID NO:
- 1.
A genetically modified adenovirus of the Ad5 serotype, wherein said adenovirus is encoded by a nucleotide sequence comprising:
- a region encoding an amino acid sequence comprising an N-terminal Ad5 serotype fiber shaft domain, a Ad3 serotype fiber knob domain, and an arginine-glycine-aspartate (RGD) peptide wherein the peptide RGD is at the C-terminus of the Ad3 serotype fiber knob;
- and
- a region encoding an adenovirus E1A polypeptide having a 24 base pair deletion compared to a wild-type E1A polypeptide, wherein the 24 base pair deletion in E1A-encoding nucleotide sequence renders the expressed E1A polypeptide unable to bind a pRb transcription factor, wherein the nucleotide sequence is SEQ ID NO:
- 1.
Dependencies are inferred from the claim text.