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Claims for Patent: 7,598,362

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Summary for Patent: 7,598,362
Title:Hepatitis C virus vaccine
Abstract: The present invention features Ad6 vectors and a nucleic acid encoding a Met-NS3-NS4A-NS4B-NS5A-NS5B polypeptide containing an inactive NS5B RNA-dependent RNA polymerase region. The nucleic acid is particularly useful as a component of an adenovector or DNA plasmid vaccine providing a broad range of antigens for generating an HCV specific cell mediated immune (CMI) response against HCV.
Inventor(s): Emini; Emilio A. (Wayne, PA), Kaslow; David C. (Rancho Santa Fe, CA), Bett; Andrew J. (Lansdale, PA), Shiver; John W. (Chalfont, PA), Nicosia; Alfredo (Rome, IT), Lahm; Armin (Rome, IT), Luzzago; Alessandra (Rome, IT), Cortese; Riccardo (Rome, IT), Colloca; Stefano (Rome, IT)
Assignee: Merck & Co., Inc. (Rahway, NJ) Istituto Di Ricerche Di Biologia Molecolare P. Angeletti S.p.A. (Rome, IT)
Application Number:10/492,178
Patent Claims:1. A nucleic acid comprising a nucleotide sequence encoding a Met-NS3-NS4A-NS4B-NS5A-NS5B polypeptide substantially similar to SEQ ID NO: 1, provided that said polypeptide has sufficient protease activity to process itself to produce an NS5B protein and said NS5B protein is enzymatically inactive, wherein said polypeptide consists of SEQ ID NO: 1 or a sequence substantially similar to SEQ ID NO: 1, wherein said sequence substantially similar to SEQ ID NO: 1 differs from SEQ ID NO: 1 by 1-20 amino acids and maintains all T-cell antigen regions present in SEQ ID NO: 1.

2. The nucleic acid of claim 1, wherein said nucleic acid is an expression vector capable of expressing said polypeptide from said nucleotide sequence in a human cell.

3. A nucleic acid comprising a gene expression cassette able to express a Met-NS3-NS4A-NS4B-NS5A-NS5B polypeptide substantially similar to SEQ ID NO: 1 in a human cell, provided that said polypeptide can process itself to produce an NS5B protein and said NS5B protein is enzymatically inactive, said expression cassette comprising: a) a promoter transcriptionally coupled to a nucleotide sequence encoding said polypeptide; b) a 5' ribosome binding site functionally coupled to said nucleotide sequence, c) a terminator joined to the 3' end of said nucleotide sequence, and d) a 3' polyadenylation signal functionally coupled to said nucleotide sequence, wherein said polypeptide consists of SEQ ID NO: 1 or a sequence substantially similar to SEQ ID NO: 1, wherein said sequence substantially similar to SEQ ID NO: 1 differs from SEQ ID NO: 1 by 1-20 amino acids and maintains all T-cell eptitope regions present in SEQ ID NO: 1.

4. The nucleic acid of claim 3, wherein said nucleic acid is a shuttle vector further comprising a selectable marker, an origin of replication, a first adenovirus homology region and a second adenovirus homology region flanking said expression cassette, wherein said first homology region has at least about 100 base pairs substantially homologous to at least right end of a wild-type adenovirus region from about base pairs 1-425, and said second homology region has at least about 100 base pairs substantially homologous to at least the left end of a wild-type adenovirus region from about base pairs 3511-5792 of Ad5 or corresponding region of another adenovirus.

5. The nucleic acid of claim 3, wherein said nucleic acid is a plasmid suitable for administration into a human and further comprises a prokaryotic origin of replication and a gene coding for a selectable marker.

6. The nucleic acid of claim 5, wherein said nucleotide sequence encodes for a polypeptide of SEQ ID NO: 1.

7. The nucleic acid of claim 6, wherein said nucleotide sequence is the coding sequence of either SEQ ID NO: 2, SEQ ID NO: 3, SEQ ID NO: 10, or SEQ ID NO: 11.

8. The nucleic acid of claim 6, wherein said nucleotide sequence is the coding sequence of SEQ ID NO: 2 or SEQ ID NO: 3.

9. The nucleic acid of claim 6, wherein said promoter is the human intermediate early cytomegalovirus promoter (intron A), said 5' ribosome binding site consists of SEQ ID NO: 12, and said 3' polyadenylation is the bovine growth hormone (BGH) polyadenylation signal.

10. The nucleic acid of claim 3, wherein said nucleic acid is a adenovirus genome plasmid comprising a selectable marker, an origin of replication, and a recombinant adenovector genome containing an E1 deletion, an E3 deletion, and said expression cassette.

11. The nucleic acid of claim 3, wherein said nucleic acid is a adenovirus genome plasmid comprising a selectable marker, an origin of replication, and a) a first adenovirus region from about base pair 1 to about base pair 450 corresponding to either Ad5 or Ad6; b) said gene expression cassette in a E1 parallel or E1 anti-parallel orientation joined to said first region; c) a second adenovirus region from about base pair 3511 to about base pair 5548 corresponding to Ad5 or from about base pair 3508 to about base pair 5541 corresponding to Ad6, joined to said expression cassette; d) a third adenovirus region from about base pair 5549 to about base pair 28133 corresponding to Ad5 or from about base pair 5542 to about base pair 28156 corresponding to Ad6, joined to said second region; e) a fourth adenovirus region from about base pair 30818 to about base pair 33966 corresponding to Ad5 or from about base pair 30789 to about base pair 33784 corresponding to Ad6, joined to said third region; and f) a fifth adenovirus region from about base pair 33967 to about base pair 35935 corresponding to Ad5 or from about base pair 33785 to about base pair 35759 corresponding to Ad6, joined to said fourth region.

12. The nucleic acid of claim 3, wherein said nucleic acid is a adenovirus genome plasmid comprising an origin of replication, a selectable marker, and: a) a first adenovirus region from about base pair 1 to about base pair 450 corresponding to either Ad5 or Ad6; b) a second adenovirus region from about base pair 3511 to about base pair 5548 corresponding to Ad5 or from about base pair 3508 to about base pair 5541 corresponding to Ad6, joined to said first region; c) a third adenovirus region from about base pair 5549 to about base pair 28133 corresponding to Ad5 or from about base pair 5542 to about base pair 28156 corresponding to Ad6, joined to said second region; d) said gene expression cassette in a E3 parallel or E3 anti-parallel orientation joined to said third region; e) a fourth adenovirus region from about base pair 30818 to about base pair 33966 corresponding to Ad5 or from about base pair 30789 to about base pair 33784 corresponding to Ad6, joined to said gene expression cassette; and f) a fifth adenovirus region from about base pair 33967 to about base pair 35935 corresponding to Ad5 or from about base pair 33785 to about base pair 35759 corresponding to Ad6, joined to said fourth region.

13. An adenovector consisting of: a) a first adenovirus region from about base pair 1 to about base pair 450 corresponding to either Ad5 or Ad6; b) a gene expression cassette in a E1 parallel or E1 anti-parallel orientation joined to said first region; c) a second adenovirus region from about base pair 3511 to about base pair 5548 corresponding to Ad5 or from about base pair 3508 to about base pair 5541 corresponding to Ad6, joined to said expression cassette; d) a third adenovirus region from about base pair 5549 to about base pair 28133 corresponding to Ad5 or from about base pair 5542 to about base pair 28156 corresponding to Ad6, joined to said second region; e) a fourth adenovirus region from about base pair 30818 to about base pair 33966 corresponding to Ad5 or from about base pair 30789 to about base pair 33784 corresponding to Ad6, joined to said third region; and f) a fifth adenovirus region from about base pair 33967 to about base pair 35935 corresponding to Ad5 or from about base pair 33785 to about base pair 35759 corresponding to Ad6, joined to said fourth region, wherein said gene expression cassette is able to express a Met-NS3-NS4A-NS4B-NS5A-NS5B polypeptide substantially similar to SEQ ID NO: 1 in a human cell, provided that said polypeptide can process itself to produce an NS5B protein and said NS5B protein is enzymatically inactive, said expression cassette comprising: a) a promoter transcriptionally coupled to a nucleotide sequence encoding said polypeptide; b) a 5' ribosome binding site functionally coupled to said nucleotide sequence, c) a terminator joined to the 3' end of said nucleotide sequence, and d) a 3' polyadenylation signal functionally coupled to said nucleotide sequence, wherein said polypeptide consists of SEQ ID NO: 1 or a sequence substantially similar to SEQ ID NO: 1, wherein said substantially similar to SEQ ID NO: 1 differs from SEQ ID NO:1 by 1-20 amino acids and maintains all T-cell epitope regions present in SEQ ID NO: 1.

14. The nucleic acid of claim 13, wherein said first region corresponds to Ad5, said second region corresponds to Ad5, said third region corresponds to Ad5, said fourth region corresponds to Ad5, and said fifth region corresponds to Ad5.

15. The nucleic acid of claim 14, wherein said promoter is the human intermediate early cytomegalovirus promoter, said 5' ribosome binding site consists of SEQ ID NO: 12, and said 3' polyadenylation is the BGH polyadenylation signal.

16. The nucleic acid of claim 15, wherein said expression cassette is in an E1 anti parallel orientation and said nucleotide sequence is either SEQ ID NO: 2, SEQ ID NO: 3, SEQ ID NO: 10, or SEQ ID NO: 11.

17. The nucleic acid of claim 13, wherein said first region corresponds to Ad5 or Ad6, said second region corresponds to Ad5 or Ad6, said third region corresponds to Ad6, said fourth region corresponds to Ad6, and said fifth region corresponds to Ad5 or Ad6.

18. The nucleic acid of claim 17, where said promoter is the human intermediate early cytomegalovirus promoter, said 5' ribosome binding site consists of SEQ ID NO: 12, and said 3' polyadenylation is the BGH polyadenylation signal.

19. The nucleic acid of claim 18, wherein said expression cassette is in an E1 anti parallel orientation and said nucleotide sequence is SEQ ID NO: 2, SEQ ID NO: 3, SEQ ID NO: 10, or SEQ ID NO: 11.

20. The nucleic acid of claim 18, wherein said expression cassette is in an E1 anti parallel orientation and said nucleotide sequence is SEQ ID NO: 2 or SEQ ID NO: 3.

21. The nucleic acid of claim 3, wherein said nucleic acid is an adenovector consisting of: a) a first adenovirus region from about base pair 1 to about base pair 450 corresponding to either Ad5 or Ad6; b) a second adenovirus region from about base pair 3511 to about base pair 5548 corresponding to Ad5 or from about base pair 3508 to about base pair 5541 corresponding to Ad6, joined to said first region; c) a third adenovirus region from about base pair 5549 to about base pair 28133 corresponding to Ad5 or from about base pair 5542 to about base pair 28156 corresponding to Ad6, joined to said second region; d) said gene expression cassette in a E3 parallel or E3 anti-parallel orientation joined to said third region; e) a fourth adenovirus region from about base pair 30818 to about base pair 33966 corresponding to Ad5 or from about base pair 30789 to about base pair 33784 corresponding to Ad6, joined to said gene expression cassette; and f) a fifth adenovirus region from about base pair 33967 to about base pair 35935 corresponding to Ad5 or from about base pair 33785 to about base pair 35759 corresponding to Ad6, joined to said fourth region.

22. The nucleic acid of claim 21, wherein said first region corresponds to Ad5, said second region corresponds to Ad5, said third region corresponds to Ad5, said fourth region corresponds to Ad5, and said fifth region corresponds to Ad5.

23. The nucleic acid of claim 21, wherein said first region corresponds to Ad5 or Ad6, said second region corresponds to Ad5 or Ad6, said third region corresponds to Ad6, said fourth region corresponds to Ad6, and said fifth region corresponds to Ad5 or Ad6.

24. An adenovector produced by a process comprising the steps of: a) producing an adenovirus genome plasmid by homologous recombination between the shuttle vector of claim 4 and a nucleic acid comprising; a first adenovirus region from about base pair 1 to about base pair 450 corresponding to either Ad5 or Ad6; a second adenovirus region from about base pair 3511 to about base pair 5548 corresponding to Ad5 or from about base pair 3508 to about base pair 5541 corresponding to Ad6, joined to said first region; a third adenovirus region from about base pair 5549 to about base pair 28133 corresponding to Ad5 or from about base pair 5542 to about base pair 28156 corresponding to Ad6, joined to said second region; a fourth adenovirus region from about base pair 30818 to about base pair 33966 corresponding to Ad5 or from about base pair 30789 to about base pair 33784 corresponding to Ad6, joined to said third region; and a fifth adenovirus region from about base pair 33967 to about base pair 35935 corresponding to Ad5 or from about base pair 33785 to about base pair 35759 corresponding to Ad6, joined to said fourth region; and b) rescuing said adenovector from said adenovirus plasmid.

25. A cultured recombinant cell comprising the nucleic acid of claim 2.

26. A cultured recombinant cell comprising the nucleic acid of claim 4.

27. A method of making an adenovector comprising the steps of: a) producing an adenovirus genome plasmid comprising a gene expression cassette by homologous recombination between the nucleic acid of claim 4 and a nucleic acid comprising; a first adenovirus region from about base pair 1 to about base pair 450 corresponding to either Ad5 or Ad6; a second adenovirus region from about base pair 3511 to about base pair 5548 corresponding to Ad5 or from about base pair 3508 to about base pair 5541 corresponding to Ad6, joined to said first region; a third adenovirus region from about base pair 5549 to about base pair 28133 corresponding to Ad5 or from about base pair 5542 to about base pair 28156 corresponding to Ad6, joined to said second region; a fourth adenovirus region from about base pair 30818 to about base pair 33966 corresponding to Ad5 or from about base pair 30789 to about base pair 33784 corresponding to Ad6, joined to said third region; and a fifth adenovirus region from about base pair 33967 to about base pair 35935 corresponding to Ad5 or from about base pair 33785 to about base pair 35759 corresponding to Ad6, joined to the fourth region; and b) rescuing said recombinant adenovirus from said recombinant adenovirus plasmid.

28. The nucleic acid of claim 1, wherein said polypeptide consists of SEQ ID NO: 1.

29. The nucleic acid of claim 1, wherein said polypeptide differs from SEQ ID NO: 1 by 1-20 amino acids and maintains all T-cell antigen regions present in SEQ ID NO: 1.

30. The nucleic acid of claim 1, wherein said polypeptide differs from SEQ ID NO: 1 by 1-10 amino acids and maintains all T-cell antigen regions present in SEQ ID NO: 1.

31. The nucleic acid of claim 1, wherein said nucleotide sequence is the coding sequence of either SEQ ID NO: 2, SEQ ID NO: 3, SEQ ID NO: 10 or SEQ ID NO: 11.

32. The nucleic acid of claim 1, wherein said nucleotide sequence is the coding sequence of either SEQ ID NO: 2 or SEQ ID NO: 3.

33. The nucleic acid of claim 1, wherein said nucleotide sequence is the coding sequence for SEQ ID NO: 2 or differs from SEQ ID NO: 2 by 1 to 50 nucleotides.

34. The nucleic acid of claim 3, wherein said polypeptide differs from SEQ ID NO: 1 by 1-10 amino acids and maintains all T-cell antigen regions present in SEQ ID NO: 1.

35. The nucleic acid of claim 3, wherein said polypeptide consists of SEQ ID NO: 1.

36. A pharmaceutical composition comprising: an effective amount of a nucleic acid comprising a gene expression cassette able to express a Met-NS3-NS4A-NS4B-NS5A-NS5B polypeptide in a human cell, wherein said polypeptide consists of SEQ ID NO: 1 or a sequence substantially similar to SEQ ID NO: 1, provided that said polypeptide can process itself to produce an NS5B protein and said NS5B protein is enzymatically inactive, said expression cassette comprising: a) a promoter transcriptionally coupled to a nucleotide sequence encoding said polypeptide; b) a 5' ribosome binding site functionally coupled to said nucleotide sequence, c) a terminator joined to the 3' end of said nucleotide sequence, and d) a 3' polyadenylation signal functionally coupled to said nucleotide sequence; wherein said sequence substantially similar to SEQ ID NO: 1 differs from SEQ ID NO: 1 by 1-20 amino acids and maintains all T-cell antigen regions present in SEQ ID NO: 1; and a pharmaceutically acceptable carrier.

37. The pharmaceutical composition of claim 36, wherein said nucleic acid is a plasmid suitable for administration into a human and further comprises a prokaryotic origin of replication and a gene coding for a selectable marker.

38. The pharmaceutical composition of claim 37, wherein said nucleotide sequence is the coding sequence of either SEQ ID NO: 2, SEQ ID NO: 3, SEQ ID NO: 10, or SEQ ID NO: 11.

39. The pharmaceutical composition of claim 38, wherein said nucleotide sequence is the coding sequence of SEQ ID NO: 2 or SEQ ID NO: 3.

40. The pharmaceutical composition of claim 39, wherein said nucleotide sequence is the coding sequence of SEQ ID NO: 2.

41. The pharmaceutical composition of claim 37, wherein said promoter is the human intermediate early cytomegalovirus promoter (intron A), said 5' ribosome binding site consists of SEQ ID NO: 12, and said 3' polyadenylation is the bovine growth hormone (BGH) polyadenylation signal.

42. A pharmaceutical composition comprising: an effective amount of a nucleic acid comprising a gene expression cassette able to express a Met-NS3-NS4A-NS4B-NS5A-NS5B polypeptide in a human cell, wherein said polypeptide consists of SEQ ID NO: 1 or a sequence substantially similar to SEQ ID NO: 1, provided that said polypeptide can process itself to produce an NS5B protein and said NS5B protein is enzymatically inactive, said expression cassette comprising: a) a promoter transcriptionally coupled to a nucleotide sequence encoding said polypeptide; b) a 5' ribosome binding site functionally coupled to said nucleotide sequence, c) a terminator joined to the 3' end of said nucleotide sequence, and d) a 3' polyadenylation signal functionally coupled to said nucleotide sequence; wherein said substantially similar to SEQ ID NO: 1 differs from SEQ ID NO: 1 by 1 to amino acids and maintains all T-cell epitope regions present in SEQ ID NO: 1; and a pharmaceutically acceptable carrier; wherein said nucleic acid is an adenovector consisting of: a) a first adenovirus region from about base pair 1 to about base pair 450 corresponding to either Ad5 or Ad6; b) said gene expression cassette in a E1 parallel or E1 anti-parallel orientation joined to said first region; c) a second adenovirus region from about base pair 3511 to about base pair 5548 corresponding to Ad5 or from about base pair 3508 to about base pair 5541 corresponding to Ad6, joined to said expression cassette; d) a third adenovirus region from about base pair 5549 to about base pair 28133 corresponding to Ad5 or from about base pair 5542 to about base pair 28156 corresponding to Ad6, joined to said second region; e) a fourth adenovirus region from about base pair 30818 to about base pair 33966 corresponding to Ad5 or from about base pair 30789 to about base pair 33784 corresponding to Ad6, joined to said third region; and f) a fifth adenovirus region from about base pair 33967 to about base pair 35935 corresponding to Ad5 or from about base pair 33785 to about base pair 35759 corresponding to Ad6, joined to said fourth region.

43. The pharmaceutical composition of claim 42, wherein said first region corresponds to Ad5, said second region corresponds to Ad5, said third region corresponds to Ad5, said fourth region corresponds to Ad5, and said fifth region corresponds to Ad5.

44. The pharmaceutical composition of claim 43, wherein said promoter is the human intermediate early cytomegalovirus promoter, said 5' ribosome binding site consists of SEQ ID NO: 12, and said 3' polyadenylation is the BGH polyadenylation signal.

45. The pharmaceutical composition of claim 44, wherein said expression cassette is in an E1 anti parallel orientation and said nucleotide sequence is either SEQ ID NO: 2, SEQ ID NO: 3, SEQ ID NO: 10, or SEQ ID NO: 11.

46. The pharmaceutical composition of claim 42, wherein said first region corresponds to Ad5 or Ad6, said second region corresponds to Ad5 or Ad6, said third region corresponds to Ad6, said fourth region corresponds to Ad6, and said fifth region corresponds to Ad5 or Ad6.

47. The pharmaceutical composition of claim 46, wherein said promoter is the human intermediate early cytomegalovirus promoter, said 5' ribosome binding site consists of SEQ ID NO: 12, and said 3' polyadenylation is the BGH polyadenylation signal.

48. The pharmaceutical composition of claim 47, wherein said expression cassette is in an E1 anti parallel orientation and said nucleotide sequence is SEQ ID NO: 2, SEQ ID NO: 3, SEQ ID NO: 10, or SEQ ID NO: 11.

49. The pharmaceutical composition of claim 47, wherein said expression cassette is in an E1 anti parallel orientation and said nucleotide sequence is SEQ ID NO: 2 or SEQ ID NO: 3.

50. A pharmaceutical composition comprising: an effective amount of a nucleic acid comprising a gene expression cassette able to express a Met-NS3-NS4A-NS4B-NS5A-NS5B polypeptide in a human cell, wherein said polypeptide consists of SEQ ID NO: 1 or a sequence substantially similar to SEQ ID NO: 1, provided that said polypeptide can process itself to produce an NS5B protein and said NS5B protein is enzymatically inactive, said expression cassette comprising: a) a promoter transcriptionally coupled to a nucleotide sequence encoding said polypeptide; b) a 5' ribosome binding site functionally coupled to said nucleotide sequence, c) a terminator joined to the 3' end of said nucleotide sequence, and d) a 3' polyadenylation signal functionally coupled to said nucleotide sequence; wherein said sequence substantially similar to SEQ ID NO: 1 differs from SEQ ID NO: 1 by 1 to 20 amino acids and maintains all T-cell epitope regions present in SEQ ID NO: 1; and a pharmaceutically acceptable carrier; wherein said nucleic acid is an adenovector consisting of: a) a first adenovirus region from about base pair 1 to about base pair 450 corresponding to either Ad5 or Ad6; b) a second adenovirus region from about base pair 3511 to about base pair 5548 corresponding to Ad5 or from about base pair 3508 to about base pair 5541 corresponding to Ad6, joined to said first region; c) a third adenovirus region from about base pair 5549 to about base pair 28133 corresponding to Ad5 or from about base pair 5542 to about base pair 28156 corresponding to Ad6, joined to said second region; d) said gene expression cassette in a E3 parallel or E3 anti-parallel orientation joined to said third region; e) a fourth adenovirus region from about base pair 30818 to about base pair 33966 corresponding to Ad5 or from about base pair 30789 to about base pair 33784 corresponding to Ad6, joined to said gene expression cassette; and f) a fifth adenovirus region from about base pair 33967 to about base pair 35935 corresponding to Ad5 or from about base pair 33785 to about base pair 35759 corresponding to Ad6, joined to said fourth region.

Summary for Patent:   Start Trial

PCT Information
PCT FiledOctober 10, 2002PCT Application Number:PCT/US02/32512
PCT Publication Date:April 17, 2003PCT Publication Number:WO03/031588

Details for Patent 7,598,362

Applicant Tradename Biologic Ingredient Dosage Form BLA Number Approval Date Patent No. Assignee Estimated Patent Expiration Status Orphan Source
Schering INTRON A interferon alfa-2b VIAL 103132 001 1986-06-04   Start Trial Merck & Co., Inc. (Rahway, NJ) Istituto Di Ricerche Di Biologia Molecolare P. Angeletti S.p.A. (Rome, IT) 2021-10-11 RX search
Schering INTRON A interferon alfa-2b VIAL 103132 002 1986-06-04   Start Trial Merck & Co., Inc. (Rahway, NJ) Istituto Di Ricerche Di Biologia Molecolare P. Angeletti S.p.A. (Rome, IT) 2021-10-11 RX search
Schering INTRON A interferon alfa-2b VIAL 103132 003 1986-06-04   Start Trial Merck & Co., Inc. (Rahway, NJ) Istituto Di Ricerche Di Biologia Molecolare P. Angeletti S.p.A. (Rome, IT) 2021-10-11 RX search
>Applicant >Tradename >Biologic Ingredient >Dosage Form >BLA >Number >Approval Date >Patent No. >Assignee >Estimated Patent Expiration >Status >Orphan >Source

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