Processes of identifying and characterizing X-linked disorders

Patent metadata

Field	Value
Publication number	US-10385394-B2
Application number	US-201414216618-A
Country	US
Kind code	B2
Filing date	Mar 17, 2014
Priority date	Mar 15, 2013
Publication date	Aug 20, 2019
Grant date	Aug 20, 2019

Abstract

The present invention relates to processes for characterizing and screening for the existence or predisposition to X-linked disorders associated with changes in X-chromosome inactivation. The present invention also relates to processes of reducing a disease phenotype associated with an X-linked disorder in a female subject.

First claim

What is claimed is:

1. A process of characterizing X chromosome inactivation (XCI) in a female subject, the process comprising:
(a) obtaining a first biological sample comprising nucleic acids from the female subject, a second biological sample comprising nucleic acids from the biological mother of the female subject, and a third biological sample comprising nucleic acids from the biological father of the female subject;
(b) isolating the nucleic acids from the first biological sample, the second biological sample, and the third biological sample;
(c) performing whole genome sequencing or whole exome sequencing by next generation sequencing of the isolated nucleic acids from the second biological sample, and the third biological sample;
(d) identifying from the whole genome sequencing or whole exome sequencing a plurality of X-linked variant alleles, wherein the female subject is heterozygous for the X-linked variant alleles;
(e) performing whole transcriptome sequencing by next generation sequencing of the isolated nucleic acids from the first biological sample;
(f) measuring from the whole transcriptome sequencing allelic ratios of the X-linked variant alleles at each heterozygous locus; and
(g) characterizing an XCI ratio in the female subject by determining a combined mean allelic ratio for the X-linked variant alleles from the allelic ratios at each heterozygous locus.

2. The process of claim 1 , further comprising performing whole genome sequencing or whole exome sequencing by next generation sequencing of the isolated nucleic acids from the first biological sample.

3. The process of claim 1 , wherein the whole transcriptome sequencing is performed simultaneously with step (c) and comprises whole mRNA sequencing.

4. The process of claim 1 , wherein the isolated nucleic acids in step (c) comprises genomic DNA and the isolated nucleic acids in step (e) comprise total RNA or mRNA.

5. A process of identifying an X-linked disorder in a female subject, the process comprising:
(a) characterizing X chromosome inactivation (XCI) in the female subject according to the process of claim 1 ; and
(b) identifying an X-linked disorder in the female subject if the combined mean allelic ratio for the X-linked variant alleles is substantially skewed, wherein the combined mean allelic ratio is substantially skewed if expression of the X-linked variant alleles from the biological mother of the female subject compared to expression of the X-linked variant alleles from the biological father of the female subject differs by at least a ratio of 65:35.

6. The process of claim 5 , wherein the expression of at least one of the X-linked variant alleles differs by at least 80:20.

7. The process of claim 5 , wherein the X-linked disorder is at least partially caused by a change in mode and magnitude of XCI.

8. The process of claim 7 , wherein the mode of XCI is determined by phasing of the chromosome-wide X-linked heterozygous SNP alleles of the female subject.

9. The process of claim 7 , wherein the magnitude of XCI is determined in silico by computer simulation of phased or unphased chromosome-wide X-linked heterozygous SNP alleles of the female subject or the magnitude of XCI is determined by allele-specific expression analysis of phased or unphased chromosome-wide X-linked heterozygous SNP alleles of the female subject that are transcribed into mRNA.

10. The process of claim 9 , wherein the XCI ratio is estimated from phased heterozygous SNPs by a beta parametric model and the XCI ratio is estimated from unphased heterozygous SNPs by a semi-parametric model.

11. The process of claim 7 , wherein a combination of the change in mode and magnitude of XCI and the presence or absence of at least one of the X-linked variant alleles is directly associated with a phenotype for the X-linked disorder.

12. The process of claim 11 , wherein the X-linked disorder is a neurobehavioral condition manifesting at least an emotional instability, attention deficit, or delays in development and learning.

13. The process of claim 5 , wherein the X-linked variant alleles comprise a chromosomal deletion, amplification, or both.

14. The process of claim 13 , wherein a chromosomal deletion is detected by plotting log 2 differences,

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Assignees

• Translational Genomics Res Inst

Inventors

• Szelinger Szabolcs
• Craig David W
• Huentelman Matthew
• Narayanan Vinodh

Classifications

• C12Q2600/156

  Polymorphic or mutational markers · CPC title

• G16B30/00

  ICT specially adapted for sequence analysis involving nucleotides or amino acids · CPC title

• C12Q1/6883 Primary

  for diseases caused by alterations of genetic material · CPC title

• G16B30/10

  Sequence alignment; Homology search · CPC title