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Last Updated: January 18, 2020

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Claims for Patent: 9,677,071

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Summary for Patent: 9,677,071
Title:Multiple interferon and virus response element cell-based fluorescence system
Abstract: The present disclosure refers to a method for a specific, versatile and sensitive detection of IFN-/virus-induced genes, a method for quantifying IFN potency and activity in a pharmaceutical preparation or biological sample, a method for distinguishing between IFN- and viral induction, and/or for distinguishing between different viruses, and a method for the quantification of virus activity. Also, the invention provides the necessary molecular tools like expression active response constructs, suitable cell lines, an array to perform the method and a kit.
Inventor(s): Khabar; Khalid S. A. (Riyadh, SA), Al-Majhdi; Fahad Nasser (Riyadh, SA)
Assignee: KING FAISAL SPECIALIST HOSPITAL AND RESEARCH CENTRE (Riyadh, SA) KING SAUD UNIVERSITY (Riyadh, SA)
Application Number:14/112,773
Patent Claims:1. An expression active reporter construct, comprising a response element, a transcriptional control element, a reporter DNA sequence, and a termination sequence, wherein the response element is an interferon-stimulated response element (ISRE) or a virus response element (VRE) comprising any one of SEQ ID NOs: 4-24.

2. The reporter construct according to claim 1, wherein the response element is attached to a flanking region of 20-100 nucleotides.

3. The reporter construct according to claim 1, wherein the transcriptional control element comprises a minimal promoter which comprises at least a TATAA or TATAA-like signal, a GC-Box, CAAT signal, and/or an AP-1 site.

4. The reporter construct according to claim 3, wherein the minimal promoter comprises a minimal CMV promoter, a HSV TK promoter, a SV40 promoter, a synthetic minimal promoter, a viral or cellular promoter, or an inducible promoter.

5. The reporter construct according to claim 4, wherein the transcriptional control element comprises a minimal CMV IE promoter from position -36, -53, or -74 from the transcriptional start site.

6. The reporter construct according to claim 1, wherein the reporter DNA sequence encodes a reporter protein that is an enhanced green fluorescent protein (EGFP), an EGFP-MODC fusion protein, or luciferase.

7. The reporter construct according to claim 1, wherein the termination sequence comprises a SV40 polyadenylation signal, and/or wherein the termination sequence is the termination sequence of bovine growth hormone (BGH).

8. The reporter construct according to claim 1, wherein the expression active reporter construct comprises an intron or enhancer.

9. A stable cell line expressing a reporter protein from an expression active reporter construct according to claim 1.

10. The stable cell line according to claim 9, which is a Vero, 293T, K562, MDCK, HT1080, or HepGR, or a liver cell line.

11. An array comprising at least one expression active response reporter construct, wherein the expression active response reporter construct comprises a response element, a transcriptional control element, a reporter DNA sequence, and a termination sequence, wherein the response element is an interferon-stimulated response element (ISRE) or a virus response element (VRE) selected from SEQ ID NO: 4 to SEQ ID NO: 109.

12. The array according to claim 11, comprising at least two expression active reporter constructs, wherein at least two reporter constructs have different response elements, and wherein the sequences of the response elements are selected from SEQ ID NO: 4 to SEQ ID NO:109.

13. The array according to claim 11, comprising at least thirteen different expression active reporter constructs, wherein at least thirteen reporter constructs have different response elements, and wherein the sequences of the response elements are selected from SEQ ID NO: 4 to SEQ ID NO:109.

14. The array according to claim 11, wherein the response elements of the reporter constructs comprise at least one sequence selected from SEQ ID NO: 4, SEQ ID NO:5, SEQ ID NO: 6, SEQ ID NO:7, SEQ ID NO: 8, SEQ ID NO: 9, SEQ ID NO:10, and SEQ ID NO: 11.

15. The array according to claim 11, wherein the response elements of the reporter constructs comprise at least one sequence selected from SEQ ID NO: 12, SEQ ID NO: 13,SEQ ID NO:14, and SEQ ID NO: 58.

16. The array according to claim 11, wherein the reporter constructs have response elements that comprise SEQ ID NO: 4 (VREL-1), SEQ ID NO: 11 (PARP10), SEQ ID NO:16 (OAS3V2), and SEQ ID NO: 9 (USB18-M).

17. The array according to claim 11, wherein the reporter constructs have reporter constructs with response elements that comprise SEQ ID NO: 16 (OAS3V2), SEQ ID NO: 10 (IFIT3-2), SEQ ID NO: 4 (VREL-1), SEQ ID NO: 53 (GIP3-6-16), SEQ ID NO: 5 (VREL-2), SEQ ID NO: 12 (IFIT3-1), SEQ ID NO: 78 (GPB1-V), SEQ ID NO: 14 (VRE Con), SEQ ID NO: 58 (AB-VRE), SEQ ID NO: 62 (IFNA-V), SEQ ID NO: 33 (MX-1), SEQ ID NO: 15 (OAS3-V), and SEQ ID NO: 11 (PARP 10).

18. The array according to claim 11, wherein the reporter constructs have response elements that comprise SEQ ID NO: 104 (AB-VRE-M2), SEQ ID NO: 5 (VREL-2), SEQ ID NO: 4 (VREL-1), SEQ ID NO: 100 (VRE-G1), SEQ ID NO: 9 (USB18-M), SEQ ID NO: 101 (SYN-ISRE-2R), SEQ ID NO: 19 (PARP10-S), SEQ ID NO: 11 (PARP10), SEQ ID NO: 16 (OAS3V2), SEQ ID NO: 15 (OAS3-V), SEQ ID NO:18 (MX1-2-2), SEQ ID NO: 33 (MX1), SEQ ID NO: 62 (IFNA-V), SEQ ID NO: 23 (IFIT3-2S), SEQ ID NO: 10 (IFIT3-2), SEQ ID NO: 12 (IFIT3-1), SEQ ID NO: 24 (IFIT1), SEQ ID NO: 78 (GPB1-V), SEQ ID NO: 53 (GIP3-6-16), SEQ ID NO: 105 (AB-VRE-M), SEQ ID NO: 58 (AB-VRE), SEQ ID NO: 31 (HERC5), SEQ ID NO: 102 (SYN-ISRE-2), SEQ ID NO: 103 (B-VRE-3X), and SEQ ID NO: 74 (PSMP9-V).

19. The array according to claim 11, wherein the expression active response reporter construct is transfected into a stable cell line.

20. A method for detection of recombinant interferon induction, comprising the steps of: providing an array with expression active reporter constructs according to claim 1, transfection of the expression active reporter constructs into cells, exposing cells to a recombinant interferon, and detection of reporter activity.

21. The method of claim 20, wherein the reporter construct is in a 96-well plate or a 384-well plate.

22. A kit comprising an array according to claim 11, a buffer, and a stable cell line.

23. The kit of claim 22, further comprising an instruction sheet.

Summary for Patent:   Start Trial

PCT Information
PCT FiledApril 20, 2011PCT Application Number:PCT/EP2011/002029
PCT Publication Date:October 26, 2012PCT Publication Number:WO2012/143020

Details for Patent 9,677,071

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 KING FAISAL SPECIALIST HOSPITAL AND RESEARCH CENTRE (Riyadh, SA) KING SAUD UNIVERSITY (Riyadh, SA) 2039-02-26 RX search
Schering INTRON A interferon alfa-2b VIAL 103132 002 1986-06-04   Start Trial KING FAISAL SPECIALIST HOSPITAL AND RESEARCH CENTRE (Riyadh, SA) KING SAUD UNIVERSITY (Riyadh, SA) 2039-02-26 RX search
Schering INTRON A interferon alfa-2b VIAL 103132 003 1986-06-04   Start Trial KING FAISAL SPECIALIST HOSPITAL AND RESEARCH CENTRE (Riyadh, SA) KING SAUD UNIVERSITY (Riyadh, SA) 2039-02-26 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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