Nucleic acid amplification primers for pcr-based clonality studies

• US 20060234234A1
• Filed: 10/13/2003
• Published: 10/19/2006
• Est. Priority Date: 10/11/2002
• Status: Active Grant

• Current Assignee: Antonio Parreira, Christian Bastard, Eduardus Maria Dominicus Schuuring, Elizabeth Anne Macintyre, Frances Louise Lavender, Gareth John Morgan, Jacobus Johannes Maria Van Dongen, Jesus Fernando San Miquel, John Lewis Smith, Michael Hummel, Michael Kneba, Paul Anthony Stuart Evans, Ramon Garzia Sanz
• Original Assignee: Antonio Parreira, Christian Bastard, Eduardus Maria Dominicus Schuuring, Elizabeth Anne Macintyre, Frances Louise Lavender, Gareth John Morgan, Jacobus Johannes Maria Van Dongen, Jesus Fernando San Miquel, John Lewis Smith, Michael Hummel, Michael Kneba, Paul Anthony Stuart Evans, Ramon Garzia Sanz
• Inventors: Hummel, Michael, Parreira, Antonio, Schuuring, Eduardus Maria Dominicus, Evans, Paul Anthony Stuart, Smith, John Lewis, San Miquel, Jesus Fernando, Lavender, Frances Louise, Macintyre, Elizabeth Anne, Van Dongen, Jacobus Johannes Maria, Kneba, Michael, Garzia Sanz, Ramon, Morgan, Gareth John, Bastard, Christian
• Granted Patent: US 8,859,748 B2
• Time in Patent Office: Days
• US Class Current: 435/6
• CPC Class Codes: C12Q 1/6883 for diseases caused by alterations of genetic material
• CPC Class Codes: C12Q 1/6886 for cancer immunoassay for cancer G01N33/574
• CPC Class Codes: C12Q 2600/112 Disease subtyping, staging or classification
• CPC Class Codes: C12Q 2600/156 Polymorphic or mutational markers
• CPC Class Codes: C12Q 2600/16 Primer sets for multiplex assays

Abstract

The invention relates to PCR-based clonality studies for among others early diagnosis of lymphoproliferative disorders. Provided is a set of nucleic acid amplification primers comprising a forward primer, or a variant thereof, and a reverse primer, or a variant thereof, capable of amplifying a rearrangement selected from the group consisting of a VH-JH IGH rearrangement, a DH-JH IGH rearrangement, a VK-JK IGK rearrangement, a VK/intron-Kde IGK rearrangement, a Vλ-Jλ IGL rearrangement, a Vβ-Jβ TCRB rearrangement, a Dβ-Jβ TCRB rearrangement, a Vγ-Jγ TCRG rearrangement, a Vδ-Jδ TCRD rearrangement, a Dδ-Dδ TCRD rearrangement, a Dδ-Jδ TCRD rearrangement, a Vδ-Dδ TCRD rearrangement, or a translocation selected from t(11; 14)(BCL1-IGH) and t(14;18)(BCL2-IGH). The primers can be used in PCR-based clonality studies for early diagnosis of lymphoproliferative disorders and detection of minimal residual disease (MRD). Also provided is a kit comprising at least one set of primers of the invention.

Claims

1. A set of nucleic amplification primers capable of amplifying a V_H-J_H IGH rearrangement comprising a forward primer and a reverse primer, wherein said forward primer is selected from the V_H family primers shown in FIG. 3B, or a variant thereof, and wherein said reverse primer is the J_H consensus primer shown in FIG. 3B, or a variant thereof.

20. A nucleic acid amplification assay, preferably a PCR assay, more preferably a multiplex PCR assay, using at least one set of primers according to any one of claims 1 to 19.

21. A method for detecting a V_H-J_H IGH rearrangement, comprising using one or more sets of primers according to claim 1 in a nucleic acid amplification assay according to claim 20.

23. A method for detecting a V_K-J_K IGK rearrangement, comprising using one or more sets of primers according to claim 3 in a nucleic acid amplification assay according to claim 20.

24. A method for detecting a V_K/intron-Kde IGK rearrangement, comprising using one or more sets of primers according to claim 4 in a nucleic acid amplification assay according to claim 20.

25. A method for detecting a Vλ

-Jλ

IGL rearrangement, comprising using one or more sets of primers according to claim 5 in a nucleic acid amplification assay according to claim 20.

26. A method for detecting a Vβ

-Jβ

TCRB rearrangement, comprising using one or more sets of primers according to claim 6 in a nucleic acid amplification assay according to claim 20.

27. A method for detecting a Dβ

-Jβ

TCRB rearrangement, comprising using one or more sets of primers according to claim 7 in a nucleic acid amplification assay according to claim 20.

28. A method for detecting a Vγ

-Jγ

TCRG rearrangement, comprising using one or more sets of primers according to claim 8 in a nucleic acid amplification assay according to claim 20.

29. A method for detecting a Vδ

-Jδ

TCRD rearrangement, comprising using one or more sets of primers according to claim 9 in a nucleic acid amplification assay according to claim 20.

37. A method for detecting two or more rearrangements, two or more translocations or at least one rearrangement and at least one translocation selected from the group consisting of a V_H-J_H IGH rearrangement, a D_H-J_H IGH rearrangement, a V_K-J_K IGK rearrangement, a V_K/intron-Kde IGK rearrangement, a Vλ

-Jλ

IGL rearrangement, a Vβ

-Jβ

TCRB rearrangement, a Dβ

-Jβ

TCRB rearrangement, a Vγ

-Jγ

TCRG rearrangement, a Vδ

-Jδ

TCRD rearrangement, a Dδ

-Dδ

TCRD rearrangement, a Dδ

-Jδ

TCRD rearrangement, a Vδ

-Dδ

TCRD rearrangement, a t(11;

14)(BCL1-IGH) translocation and t(14;

18)(BCL2-IGH) translocation, using at least two sets of primers according to any one of claims 1 or 14.

38. A method for assessing clonal rearrangements and/or chromosome aberrations using at least one set of primers according to any one of claims 1 to 14, and optionally at least one set of primers according to any one of claims 15 to 19.

39. A method according to claim 38 for the detection of minimal residual disease (MRD) or for identification of PCR targets to be used for MRD detection via real-time quantitative PCR.

40. A method according to claim 38 or 39, wherein an amplified nucleic acid is detected using automated high resolution PCR fragment analysis.

41. A kit for the detection of at least one rearrangement selected from the group consisting of a V_H-J_H IGH rearrangement, a D_H-J_H IGH rearrangement, a V_K-J_K IGK rearrangement, a V_K/intron-Kde IGK rearrangement, a Vλ

-Jλ

IGL rearrangement, a Vβ

-Jβ

TCRB rearrangement, a Dβ

-Jβ

TCRB rearrangement, a Vγ

-Jγ

TCRG rearrangement, a Vδ

-Jδ

TCRD rearrangement, a Dδ

-Dδ

TCRD rearrangement, a Dδ

-Jδ

TCRD rearrangement, a Vδ

-Dδ

TCRD, and/or at least one translocation selected from t(11;

14)(BCL1-IGH) and t(14;

18)(BCL2-IGH), comprising at least one set of primers according to any one of claims 1 to 14.

2. A set of nucleic amplification primers capable of amplifying a D_H-J_H IGH rearrangement comprising a forward primer and a reverse primer, wherein said forward primer is selected from the D_H family primers shown in FIG. 4A, or a variant thereof, and wherein said reverse primer is the J_H consensus primer shown in FIG. 4A, or a variant thereof.

22. A method for detecting a D_H-J_H IGH rearrangement, comprising using one or more sets of primers according to claim 2 in a nucleic acid amplification assay according to claim 20.

3. A set of nucleic amplification primers capable of amplifying a V_K-J_K IGK rearrangement comprising a forward primer and a reverse primer, wherein said forward primer is selected from the V_K family primers shown in FIG. 5B, or a variant thereof, and wherein said reverse primer is a J_K primer shown in FIG. 5B, or a variant thereof.

4. A set of nucleic amplification primers capable of amplifying a V_K/intron-Kde IGK rearrangement comprising a forward primer and a reverse primer, wherein said forward primer is selected from the Vκ

primers or the INTR primer shown in FIG. 5B, or a variant thereof, and wherein said reverse primer is the Kde primer shown in FIG. 5B, or a variant thereof.

5. A set of nucleic amplification primers capable of amplifying a Vλ

-Jλ

IGL rearrangement comprising a forward primer and a reverse primer, wherein said forward primer is selected from the Vλ

primers shown in FIG. 6B, or a variant thereof, and wherein said reverse primer is the Jλ

primer shown in FIG. 6B, or a variant thereof.

6. A set of nucleic amplification primers capable of amplifying a Vβ

-Jβ

TCRB rearrangement comprising a forward primer and a reverse primer, wherein said forward primer is selected from the Vβ

family primers shown in FIG. 7B, or a variant thereof, and wherein said reverse primer is selected from the Jβ

A en Jβ

B primers shown in FIG. 7B, or a variant thereof.

7. A set of nucleic amplification primers capable of amplifying a Dβ

-Jβ

TCRB rearrangement comprising a forward primer and a reverse primer, wherein said forward primer is selected from the DB primers shown in FIG. 7B, or a variant thereof, and wherein said reverse primer is selected from the Jβ

A en Jβ

B primers shown in FIG. 7B, or a variant thereof.

8. A set of nucleic amplification primers capable of amplifying a Vγ

-Jγ

TCRG rearrangement comprising a forward primer and a reverse primer, wherein said forward primer is selected from the Vγ

family primers shown in FIG. 8B, or a variant thereof, and wherein said reverse primer is selected from the Jγ

primers shown in FIG. 8B, or a variant thereof.

9. A set of nucleic amplification primers capable of amplifying a Vδ

-Jδ

TCRD rearrangement comprising a forward primer and a reverse primer, wherein said forward primer is selected from the Vδ

primers shown in FIG. 9B, or a variant thereof, and wherein said reverse primer is selected from the Jδ

primers shown in FIG. 9B, or a variant thereof.

10. A set of nucleic amplification primers capable of amplifying a Dδ

-Dδ

TCRD rearrangement comprising a forward primer and a reverse primer, wherein said forward primer is the Dδ

2 primer shown in FIG. 9B, or a variant thereof, and wherein said reverse primer is the Dδ

3 primer shown in FIG. 9B, or a variant thereof.

30. A method for detecting a Dδ

-Dδ

TCRD rearrangement, comprising using one or more sets of primers according to claim 10 in a nucleic acid amplification assay according to claim 20.

11. A set of nucleic amplification primers capable of amplifying a Dδ

-Jδ

TCRD rearrangement comprising a forward primer and a reverse primer, wherein said forward primer is the Dδ

2 primer shown in FIG. 9B, or a variant thereof, and wherein said reverse primer is selected from the Jδ

primers shown in FIG. 9B, or a variant thereof.

31. A method for detecting a Dδ

-Jδ

TCRD rearrangement, comprising using one or more sets of primers according to claim 11 in a nucleic acid amplification assay according to claim 20.

12. A set of nucleic amplification primers capable of amplifying a Vδ

-Dδ

TCRD rearrangement comprising a forward primer and a reverse primer, wherein said forward primer is selected from the Vδ

primers shown in FIG. 9B, or a variant thereof, and wherein said reverse primer is the Dδ

3 primer shown in FIG. 9B, or a variant thereof.

32. A method for detecting a Vδ

-Dδ

TCRD rearrangement, comprising using one or more sets of primers according to claim 12 in a nucleic acid amplification assay according to claim 20.

13. A set of nucleic amplification primers capable of amplifying a chromosomal translocation (11;

14)(BCL1-IGH) comprising a forward primer and a reverse primer, wherein said forward primer is the BCL1/MTC primer as shown in FIG. 10A, or a variant thereof, and wherein said reverse primer is the JH consensus primer shown in FIG. 10A, or a variant thereof.

33. A method for detecting a chromosomal translocation (11;

14)(BCL1-IGH), comprising using one or more sets of primers according to claim 13 in a nucleic acid amplification assay according to claim 20.

14. A set of nucleic amplification primers capable of amplifying a chromosomal translocation t(14;

18)(BCL2-IGH), comprising a forward primer and a reverse primer, wherein said forward primer is selected from the MBR primers, the 3′

MBR primers and the mcr primers shown in FIG. 11A, or a variant thereof, and wherein said reverse primer is the JH consensus primer shown in FIG. 11A, or a variant thereof.

34. A method for detecting a chromosomal translocation t(14;

18)(BCL2-IGH), comprising using one or more sets of primers according to claim 14 in a nucleic acid amplification assay according to claim 20.

15. A set of nucleic amplification primers capable of amplifying the human TBXAS1 gene comprising a forward and a reverse primer, wherein said forward primer is the TBXAS1/X9U primer of FIG. 12A, or a variant thereof, and wherein said reverse primer is the TBXAS1/X9L primer of FIG. 12A, or a variant thereof.

35. A method for detecting a gene selected from the group consisting of the human AF4 gene (exon 3), the human AF4 gene (exon 11), the human PLZF1 gene, the human RAG1 gene and the human TBXAS1 gene, comprising using one or more sets of primers according to any one of claims 15 to 19 in a nucleic acid amplification assay according to claim 20.

36. Use of a method according to claim 35 to assess the quality of a DNA sample extracted from a biological sample, preferably a paraffin-embedded biological sample.

42. A kit according to claim 41, further comprising at least one set of primers according to any one of claims 15 to 19.

16. A set of nucleic amplification primers capable of amplifying the human recombination activating protein (RAG1) gene comprising a forward and a reverse primer, wherein said forward primer is the RAG1/X2U primer of FIG. 12A, or a variant thereof, and wherein said reverse primer is the RAG1/X2L primer of FIG. 12A, or a variant thereof.

17. A set of nucleic amplification primers capable of amplifying human promyelocytic leukemia zinc finger protein (PLZF) comprising a forward and a reverse primer, wherein said forward primer is the PLZF/X1U primer of FIG. 12A, or a variant thereof, and wherein said reverse primer is the PLZF/X1L primer of FIG. 12A, or a variant thereof.

18. A set of nucleic amplification primers capable of amplifying gene the human AF4 gene (Exon 3) comprising a forward and a reverse primer, wherein said forward primer is the AF4/X3U primer of FIG. 12A, or a variant thereof, and wherein said reverse primer is the AF4/X3L primer of FIG. 12A, or a variant thereof.

19. A set of nucleic amplification primers capable of amplifying gene the human AF4 gene (Exon11) comprising a forward and a reverse primer, wherein said forward primer is the AF4/X11U primer of FIG. 12A, or a variant thereof, and wherein said reverse primer is the AF4/X11L primer of FIG. 12A, or a variant thereof.