Claims for Patent: 7,850,957

Summary for Patent: 7,850,957

Title:	Adenovirus/alphavirus hybrid vector for the effective administration and expression of therapeutic genes in tumour cells
Abstract:	The present invention relates to a genic expression adenoviral hybrid vector characterized in that it contains at least the following elements, oriented in the direction 5\' to 3\': i. a first chain of adenoviral origin comprising a first inverted terminal repeat (ITR) sequence and a signal sequence for packaging of the adenovirus; ii. a first non-encoding stuffer sequence; iii. a sequence corresponding to a tissue specific promoter; iv. a chain of cDNA derived from an alphavirus, the sequence of which is partly complementary to an alphaviral RNA sequence, comprising at least a sequence encoding for at least one exogenous gene of interest; v. a polyadenylation sequence; and vi. a second adenoviral inverted terminal repeat (ITR) sequence, it preferably relates to an adenoviral hybrid vector comprising as exogenous gene of interest the therapeutic gene of mammalian interleukin IL-12 and even more preferably human interleukin hIL-12; and to the use of the hybrid vector in a process for transferring genetic material to a cell, particularly a tumor cell that preferably expresses alpha-fetoprotein (AFP), and to its use for inducing an immune response against foreign antigens.
Inventor(s):	Qian; Cheng (Gorraiz, ES), Guan; Min (Arcadia, CA), Picazo; Cristian Smerdou (Pamplona, ES), Valtuena; Jes s Prieto (Pamplona, ES)
Assignee:	Proyecto De Biomecdicina Cima, S.L. (Pamplona (Navarra), ES)
Application Number:	11/569,303
Patent Claims:	1. A genic expression adenoviral hybrid vector for expressing a gene of interest directly in an adenovirus-permissive cell in which a tissue-specific promoter is active, wherein said tissue specific promoter is the tumor specific promoter AFP, characterized in that the genic expression adenoviral hybrid vector comprises at least the following elements, oriented in the direction 5' to 3': i. a first chain of adenoviral origin comprising a first inverted terminal repeat (ITR) sequence and a signal sequence for packaging of the adenovirus; ii. a first non-encoding stuffer sequence; iii. a sequence corresponding to a tissue specific promoter, wherein said tissue specific promoter is the tumor specific promoter AFP; iv. a chain of cDNA derived from an alphavirus, the sequence of which is partly complementary to an alphaviral RNA sequence, comprising at least a sequence encoding for at least one exogenous gene of interest; said chain of cDNA derived from an alphavirus comprising: (a) a 5' sequence necessary for replication of the alphavirus, (b) a sequence encoding for the non-structural proteins required for replication of the alphaviral RNA, (c) at least one subgenomic promoter of the alphavirus, and (d) a 3' sequence necessary for replication of the alphavirus; wherein said sequence encoding for at least one exogenous gene of interest is under the control of said at least one subgenomic promoter of the alphavirus; and said chain of cDNA derived from an alphavirus is functionally controlled by the promoter iii; v. a polyadenylation sequence; and vi. a second adenoviral inverted terminal repeat (ITR) sequence. 2. A genic expression adenoviral hybrid vector according to claim 1, further comprising an element vii which is a second non-encoding stuffer sequence located between element v and element vi. 3. A genic expression adenoviral hybrid vector according to claim 1, characterized in that element ii is a human non-encoding stuffer sequence. 4. A genic expression adenoviral hybrid vector according to claim 3, characterized in that element ii is the intron region of human genomic hypoxanthine phosphoribosyltransferase, HPRT. 5. A genic expression adenoviral hybrid vector according to claim 1, characterized in that element i comprises the sequence SEQ ID NO: 1. 6. A genic expression adenoviral hybrid vector according to claim 1, characterized in that element iii is a tumor specific promoter having the sequence SEQ ID NO: 7, corresponding to AFP p+e. 7. A genic expression adenoviral hybrid vector characterized in that the genic expression adenoviral hybrid vector comprises at least the following elements, oriented in the direction 5' to 3': i. a first chain of adenoviral origin comprising a first inverted terminal repeat (ITR) sequence and a signal sequence for packaging of the adenovirus; ii. a first non-encoding stuffer sequence; iii. a sequence corresponding to a tissue specific promoter, wherein said tissue specific promoter is the tumor specific promoter AFP; iv. a chain of cDNA derived from an alphavirus, the sequence of which is partly complementary to an alphaviral RNA sequence, comprising at least a sequence encoding for at least one exogenous gene of interest; said chain of cDNA derived from an alphavirus comprising: (a) a 5' sequence necessary for replication of the alphavirus, (b) a sequence encoding for the non-structural proteins required for replication of the alphaviral RNA, (c) at least one subgenomic promoter of the alphavirus, and (d) a 3' sequence necessary for replication of the alphavirus; wherein said sequence encoding for at least one exogenous gene of interest is under the control of said at least one subgenomic promoter of the alphavirus; and said chain of cDNA derived from an alphavirus is functionally controlled by the promoter iii; v. a polyadenylation sequence; and vi. a second adenoviral inverted terminal repeat (ITR) sequence; wherein said at least one exogenous gene of interest is expressed directly in an adenovirus-permissive cell in which a tissue-specific promoter is active, and wherein said tissue specific promoter is the tumor specific promoter AFP. 8. The genic expression adenoviral hybrid vector according to claim 1, wherein said element iv comprises a sequence derived from the Semliki Forest Virus (SFV). 9. A genic expression adenoviral hybrid vector according to claim 8, characterized in that element iv a), b) and c) comprises, as a whole, a sequence selected from the group consisting of SEQ ID NO: 3 and SEQ ID NO: 4. 10. A genic expression adenoviral hybrid vector according to claim 9, characterized in that element iv d) has the sequence SEQ ID NO: 5. 11. A genic expression adenoviral hybrid vector according to claim 1, characterized in that the exogenous gene of interest is selected from the group consisting of one or more therapeutic genes, one or more reporter genes, and combinations thereof. 12. A genic expression adenoviral hybrid vector according to claim 11, characterized in that the exogenous gene of interest is the mammalian interleukin IL-12 therapeutic gene. 13. A genic expression adenoviral hybrid vector according to claim 11, characterized in that the exogenous gene of interest is the human interleukin hIL-12 therapeutic gene. 14. A genic expression adenoviral hybrid vector according to claim 9, characterized in that element iv comprises (subgenomic promoter+exogenous gene of interest) in series or in several subsets. 15. A genic expression adenoviral hybrid vector according to claim 1, characterized in that element iv forms a replicon functionally controlled by the promoter iii, and in that the alphaviral subgenomic promoter in iv.c) functionally controls the expression of the exogenous gene of interest. 16. A genic expression adenoviral hybrid vector according to claim 1, characterized in that element v is a polyadenylation sequence of SV40. 17. A genic expression adenoviral hybrid vector according to claim 16, characterized in that element v comprises the sequence SEQ ID NO: 6. 18. A genic expression adenoviral hybrid vector according to claim 2, characterized in that the second non-encoding stuffer sequence is C346. 19. A genic expression adenoviral hybrid vector comprising: i. a first chain of adenoviral origin comprising a first inverted terminal repeat (ITR) sequence and a signal sequence for packaging of the adenovirus; ii. a first non-encoding stuffer sequence, which is the intron region of human genomic hypoxanthine phosphoribosyltransferase (HPRT); iii. a sequence corresponding to a tissue specific promoter, which is the AFP promoter, iv. a cDNA chain derived from an alphavirus, the sequence of which is partly complementary to an alphaviral RNA derived from the SFV virus, which comprises a sequence encoding for an exogenous gene of interest which is hIL-12, v. a polyadenylation sequence of SV40, vi. a second adenoviral inverted terminal repeat (ITR) sequence and vii. a second non-encoding stuffer sequence, which is human genomic C346, located between element v and element vi. 20. A genic expression adenoviral hybrid vector according to claim 1, characterized in that said vector has a length comprised between 27 and 38 kilobases. 21. A genic expression adenoviral hybrid vector characterized in that the genic expression adenoviral hybrid vector comprises at least the following elements, oriented in the direction 5' to 3': i. a first chain of adenoviral origin comprising a first inverted terminal repeat (ITR) sequence and a signal sequence for packaging of the adenovirus; ii. a first non-encoding stuffer sequence; iii. a sequence corresponding to a tissue specific promoter; iv. a cDNA chain derived from an alphavirus, the sequence of which is partly complementary to an alphaviral RNA sequence derived from the SFV virus, comprising at least a sequence encoding for at least one exogenous gene of interest; v. a polyadenylation sequence; vi. a second adenoviral inverted terminal repeat (ITR) sequence; AND vii. a second non-encoding stuffer sequence located between element v and element vi, wherein the vector comprises the sequence of SEQ ID NO:8. 22. A method of transferring genetic material to a cell comprising the steps of administering the adenoviral hybrid vector of claim 1 to a subject. 23. The method according to claim 22 wherein said cell is a tumor cell. 24. The method according to claim 23 wherein said cell is a tumor cell expressing AFP. 25. A method of treating hepatocarcinoma tumors in a subject comprising the step of directly administering the adenoviral vector of claim 1 comprising a nucleotide sequence encoding IL-12 to said tumor in a subject. 26. A method for inducing an immune response against foreign antigens comprising the steps of administering said adenoviral hybrid vector of claim 1 to a subject. 27. A pharmaceutical composition comprising at least one adenoviral hybrid vector according to claim 1. 28. The pharmaceutical composition according to claim 27, characterized in that said composition comprises at least one adenoviral hybrid vector defined according to claim 1, wherein the exogenous gene of interest is hIL-12. 29. A method of inducing an immune response against foreign antigens in a subject comprising the steps of administering to said subject the pharmaceutical composition of claim 27, the hybrid vector further comprising a sequence encoding a foreign antigen of interest.

Details for Patent 7,850,957

Applicant	Tradename	Biologic Ingredient	Dosage Form	BLA	Approval Date	Patent No.	Expiredate
Merck Sharp & Dohme Corp.	INTRON A	interferon alfa-2b	For Injection	103132	06/04/1986		2024-05-20
Merck Sharp & Dohme Corp.	INTRON A	interferon alfa-2b	For Injection	103132			2024-05-20
Merck Sharp & Dohme Corp.	INTRON A	interferon alfa-2b	Injection	103132			2024-05-20