Cell free DNA diagnostic testing standards

• US 9,499,870 B2
• Filed: 09/26/2014
• Issued: 11/22/2016
• Est. Priority Date: 09/27/2013
• Status: Active Grant

• Current Assignee: Natera Incorporated (Natera)
• Original Assignee: Natera Incorporated (Natera)
• Inventors: Babiarz, Joshua, Zimmermann, Bernhard
• Primary Examiner(s): Wallenhorst, Maureen
• Application Number:
  US14/498,629
• Publication Number:
  US 20150147815A1
• Time in Patent Office: 788 Days
• Field of Search: 436/8, 436/63, 436/94, 436/174, 435/4, 435/6.1, 435/6.14, 435/29, 435/325, 435/366, 435/372
• US Class Current: 1/1
• CPC Class Codes: C12N 15/10 Processes for the isolation, preparation or purification of DNA or RNA chemical preparation of DNA or RNA C07H21/00; preparation of non-structural polynucleotides from microorganisms or with enzymes C12P19/34
• CPC Class Codes: C12Q 1/6806 Preparing nucleic acids for analysis, e.g. for polymerase chain reaction [PCR] assay C12Q1/6804 takes precedence
• 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 2521/301 Endonuclease
• CPC Class Codes: C12Q 2537/16 Assays for determining copy number or wherein the copy number is of special importance
• CPC Class Codes: C12Q 2545/113 with an external standard/control, i.e. control reaction is separated from the test/target reaction
• CPC Class Codes: C12Q 2600/16 Primer sets for multiplex assays
• CPC Class Codes: C12Q 2600/166 Oligonucleotides used as internal standards, controls or normalisation probes
• CPC Class Codes: Y10T 436/10 Composition for standardization, calibration, simulation, stabilization, preparation or preservation; processes of use in preparation for chemical testing

Abstract

Embodiments of the invention include methods and compositions for producing proficiency testing standards for noninvasive prenatal genetic diagnostics and for the detection and monitoring of cancer. The compositions can include a plurality of different nucleosomal DNA fragments derived from either primary cells or cell lines. The amount of the different nucleosomal DNA fragments can be varied so as to simulate naturally occurring cell free DNA samples obtained from the blood of the pregnant woman or naturally occurring cell free DNA samples obtained from the blood of cancer patients.

Claims

1. A cell-free nucleic acid diagnostic proficiency testing standard composition for analyzing non-invasive prenatal genetic tests, comprising

a set of at least two nucleosomal nucleic acid preparations,

wherein the nucleosomal nucleic acid preparations in the set have a quantity of a first nucleosomal nucleic acid preparation from a first cell line derived from a first individual and a quantity of a second nucleosomal nucleic acid preparation from a second cell line derived from a second individual,

wherein the first individual is a mother or father of the second individual and the first and second individuals have different copy numbers of at least one chromosome,

and wherein the quantity of the second nucleosomal nucleic acid preparation as a percentage of the quantity of the first nucleosomal nucleic acid preparation plus the quantity of the second nucleosomal nucleic acid preparation is different for each of the nucleosomal nucleic acid preparations in the set.

2. The composition according to claim 1, wherein the first nucleosomal nucleic acid preparation and the second nucleosomal nucleic acid preparation comprise mononucleosomal ladder fractions.

3. The composition according to claim 2, wherein the first nucleosomal nucleic acid preparation further comprises dinucleosomal ladder fractions.

4. The composition of claim 1, wherein the quantity of the second nucleosomal nucleic acid preparation in each of the nucleosomal nucleic acid preparations in the set is less than 40% as a percentage of the quantity of the first nucleosomal nucleic acid preparation plus the quantity of the second nucleosomal nucleic acid preparation.

5. The composition of claim 1, wherein the first nucleosomal nucleic acid preparation is obtained after apoptosis has been induced in the first cell line and/or the second nucleosomal nucleic acid preparation is obtained after apoptosis has been induced in the second cell line.

6. The composition of claim 1, wherein the quantity of the second nucleosomal nucleic acid preparation in each of the nucleosomal nucleic acid preparations in the set is less than 5% as a percentage of the quantity of the first nucleosomal nucleic acid preparation plus the quantity of the second nucleosomal nucleic acid preparation.

7. The composition of claim 1, wherein the quantity of the second nucleosomal nucleic acid preparation in each of the nucleosomal nucleic acid preparations in the set is between about 1% and about 70% as a percentage of the quantity of the first nucleosomal nucleic acid preparation plus the quantity of the second nucleosomal nucleic acid preparation.

8. The composition of claim 1, wherein the quantity of the second nucleosomal nucleic acid preparation in each of the nucleosomal nucleic acid preparations in the set is between about 4% and about 40% as a percentage of the quantity of the first nucleosomal nucleic acid preparation plus the quantity of the second nucleosomal nucleic acid preparation.

9. The composition of claim 1, wherein the first individual or the second individual is aneuploid.

10. A method of making a cell-free nucleic acid diagnostic testing standard composition for analyzing non-invasive prenatal genetic tests, comprising

generating a set of at least two nucleosomal nucleic acid preparations,

wherein the nucleosomal nucleic acid preparations in the set are made by mixing a quantity of a first nucleosomal nucleic acid preparation derived from a first cell line derived from a first individual and a quantity of a second nucleosomal nucleic acid preparation derived from a second cell line derived from a second individual,

wherein the first individual is a mother or father of the second individual and the first and second individuals have different copy numbers of at least one chromosome,

and wherein the quantity of the second nucleosomal nucleic acid preparation as a percentage of the quantity of the first nucleosomal nucleic acid preparation plus the quantity of the second nucleosomal nucleic acid preparation is different for each of the nucleosomal nucleic acid preparations in the set.

11. The method according to claim 10, wherein the first nucleosomal nucleic acid preparation and the second nucleosomal nucleic acid comprise mononucleosomal ladder fractions.

12. The method according to claim 11, wherein the first nucleosomal nucleic acid preparation further comprises dinucleosomal ladder fractions.

13. The method of claim 10, wherein the quantity of the second nucleosomal nucleic acid preparation in each of the nucleosomal nucleic acid preparations in the set is less than 40% as a percentage of the quantity of the first nucleosomal nucleic acid preparation plus the quantity of the second nucleosomal nucleic acid preparation.

14. The method of claim 10, wherein the first nucleosomal nucleic acid preparation is obtained after apoptosis has been induced in the first cell line and/or the second nucleosomal nucleic acid preparation is obtained after apoptosis has been induced in the second cell line.

15. The method of claim 10, wherein the quantity of the second nucleosomal nucleic acid preparation in each of the nucleosomal nucleic acid preparations in the set is less than 5% as a percentage of the quantity of the first nucleosomal nucleic acid preparation plus the quantity of the second nucleosomal nucleic acid preparation.

16. The method of claim 10, wherein the quantity of the second nucleosomal nucleic acid preparation in each of the nucleosomal nucleic acid preparations in the set is between about 1% and about 70% as a percentage of the quantity of the first nucleosomal nucleic acid preparation plus the quantity of the second nucleosomal nucleic acid preparation.

17. The method of claim 10, wherein the quantity of the second nucleosomal nucleic acid preparation in each of the nucleosomal nucleic acid preparations in the set is between about 4% and about 40% as a percentage of the quantity of the first nucleosomal nucleic acid preparation plus the quantity of the second nucleosomal nucleic acid preparation.

18. The method of claim 10, wherein the first individual or the second individual is aneuploid.