RECOMBINANT ADENOVIRUS COMPRISING RECOMBINANT khp53 GENE AND THE PREPARATION METHOD AND USES THEREOF

• US 20100034775A1
• Filed: 12/20/2006
• Published: 02/11/2010
• Est. Priority Date: 12/20/2006
• Status: Abandoned Application

• Current Assignee: Shenzhen Well D. Genetic Engineering Co., Ltd.
• Original Assignee: Shenzhen Well D. Genetic Engineering Co., Ltd.
• Inventors: Xie, Qingjun
• Application Number:
  US12/520,615
• Publication Number:
  US 20100034775A1
• Time in Patent Office: Days
• US Class Current: 424/93.200
• CPC Class Codes: A61K 38/00 Medicinal preparations containing peptides peptides containing beta-lactam rings A61K31/00; cyclic dipeptides not having in their molecule any other peptide link than those which form their ring, e.g. piperazine-2,5-diones, A61K31/00; ergot alkaloids of the cyclic peptide type A61K31/48; containing macromolecular compounds having statistically distributed amino acid units A61K31/74; medicinal preparations containing antigens or antibodies A61K39/00; medicinal preparations characterised by the non-active ingredients, e.g. peptides as drug carriers, A61K47/00
• CPC Class Codes: A61P 35/00 Antineoplastic agents
• CPC Class Codes: C07K 14/4746 p53
• CPC Class Codes: C12N 15/86 Viral vectors
• CPC Class Codes: C12N 2710/10332 Use of virus as therapeutic agent, other than vaccine, e.g. as cytolytic agent
• CPC Class Codes: C12N 2710/10343 viral genome or elements thereof as genetic vector
• CPC Class Codes: C12N 2830/005 repressible enhancer/promoter combination, e.g. KRAB
• CPC Class Codes: C12N 2840/105 enhancing translation
• CPC Class Codes: C12N 7/00 Viruses; Bacteriophages; Compositions thereof; Preparation or purification thereof preparing medicinal viral antigen or antibody compositions, e.g. virus vaccines, A61K39/00

Abstract

A recombinant adenovirus vector comprises eukaryotic cell promoter, operator, Kozak sequence, exogenous gene and PolyA signal sequence from 5′ to 3′ in the adenovirus vector backbone. Optionally, introns are inserted between the operator and Kozak sequence, and the exogenous gene is p53 gene. A recombinant adenovirus obtained by co-transfection of the adenovirus vector and adenovirus backbone is also included in the invention. Also disclosed are the preparation methods and uses of the recombinant adenovirus vector and the recombinant adenovirus, respectively. The present invention further discloses a pharmaceutical composition for treating cancers, which comprises the recombinant adenovirus.

Claims

1. A recombinant adenovirus shuttle vector, wherein a Kozak sequence and an exogenous gene are inserted between an operator and a PolyA signal sequence in the adenovirus shuttle vector pDC315(io) or pDC316(io), said adenovirus shuttle vector pDC315(io) is shown as FIG. 11(A), and said adenovirus shuttle vector pDC316(io) is shown as FIG. 11(B).

3. The recombinant adenovirus shuttle vector according to claim 1 or 2, wherein said Kozak sequence is CCACCATGG (SEQ ID NO:

4).

4. The recombinant adenovirus shuttle vector according to claim 1 or 2, wherein said exogenous gene is human p53 gene which comprises the nucleotide sequence set forth in SEQ ID NO:

3.

11. A method for producing the recombinant adenovirus shuttle vector of claim 1 or 2, comprising inserting an exogenous gene linked with a Kozak sequence between an operator and a PolyA signal sequence of an adenovirus shuttle vector in a forward direction, wherein said adenovirus shuttle vector is selected from the group consisting of pDC315(io), pDC316(io), pDC515(io) and pDC516(io).

14. A recombinant adenovirus obtained by co-transfecting 293IQ cells with the recombinant adenovirus shuttle vector of claim 1 and an adenoviral backbone, said adenovirus backbone is pBHGlox(delta)E1,3Cre shown as FIG. 12(A).

15. A recombinant adenovirus obtained by co-transfecting 293IQ cells with the recombinant adenovirus shuttle vector of claim 1 and an adenoviral backbone, said adenovirus backbone is pBHGfrt(del)E1,3FLP shown as FIG. 12(B).

16. The recombinant adenovirus of claim 14, wherein said Kozak sequence in said recombinant adenovirus shuttle vector is CCACCATGG (SEQ ID NO:

4).

17. The recombinant adenovirus of claim 15, wherein said Kozak sequence in said recombinant adenovirus shuttle vector is CCACCATGG (SEQ ID NO:

4).

18. The recombinant adenovirus of claim 14, wherein said exogenous gene in said recombinant adenovirus shuttle vector is human p53 gene which comprises the nucleotide sequence set forth in SEQ ID NO:

3.

19. The recombinant adenovirus of claim 15, wherein said exogenous gene in said recombinant adenovirus shuttle vector is human p53 gene which comprises the nucleotide sequence set forth in SEQ ID NO:

3.

21. A medicament for treating cancers, comprising the recombinant adenovirus according to any one of claims 14, 15 or 20, and a pharmaceutically acceptable excipient.

22. A method for producing a recombinant adenovirus, comprising co-transfecting 293IQ cells with the recombinant adenovirus shuttle vector according to claim 1 or claim 2 and an adenoviral backbone, wherein if said adenovirus shuttle vector is pDC315(io) or pDC316(io), said adenovirus backbone is pBHGlox(delta)E1,3Cre shown as FIG. 12(A), and if said adenovirus shuttle vector is pDC515(io) or pDC516(io), said adenovirus backbone is pBHGfrt(del)E1,3FLP shown as FIG. 12(B).

23. A method for treating cancers, comprising administering a therapeutic effective amount of the recombinant adenovirus according to any one of claim 14, claim 15 or claim 20, to a patient.

24. The method of claim 23, wherein said cancer is selected from the group consisting of lung cancer, liver cancer, breast cancer, nasopharyngeal cancer, ovarian cancer and cervical cancer.

2. A recombinant adenovirus shuttle vector, wherein a Kozak sequence and an exogenous gene are inserted between an operator and a PolyA signal sequence in the adenovirus shuttle vector pDC515(io) or pDC516(io), said adenovirus shuttle vector pDC515(io) is shown as FIG. 11(C), and said adenovirus shuttle vector pDC516(io) is shown as FIG. 11(D).

5. An eukaryotic gene expression vector, comprising the sequence of SEQ ID NO:

11.

6-10. -10. (canceled)

12-13. -13. (canceled)

20. A recombinant adenovirus, which is the adenovirus with the deposite number of CCTCC V200508.