# US 12,112,832 B2 · Methods and processes for non-invasive assessment of genetic variations

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- Number: US 12,112,832 B2
- Document ID: US12112832B2
- Status: Granted
- Kind: B2
- Application number: 16/573,161
- Filed: 2019-09-17
- Published: 2020-03-05
- Granted: 2024-10-08
- Earliest claimed: 2012-10-04
- Assignees: Sequenom, Inc.
- Applicants: Sequenom, Inc.
- Inventors: Cosmin Deciu; Zeljko Dzakula; Amin Mazloom
- Web page: https://patentgrep.com/patent/US12112832B2

## CPC

- G16B 20/10: Ploidy or copy number detection
- C12Q 1/6869: Methods for sequencing
- C12Q 1/6883: for diseases caused by alterations of genetic material
- G16B 30/00: ICT specially adapted for sequence analysis involving nucleotides or amino acids
- G16B 30/10: Sequence alignment; Homology search
- G16B 30/20: Sequence assembly
- G16B 40/00: ICT specially adapted for biostatistics; ICT specially adapted for bioinformatics-related machine learning or data mining, e.g. knowledge discovery or pattern finding
- C12Q 2545/101: with an internal standard/control
- C12Q 2545/114: involving a quantitation step
- G16B: BIOINFORMATICS, i.e. INFORMATION AND COMMUNICATION TECHNOLOGY [ICT] SPECIALLY ADAPTED FOR GENETIC OR PROTEIN-RELATED DATA PROCESSING IN COMPUTATIONAL MOLECULAR BIOLOGY
- C12Q: MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS (immunoassay G01N33/53); COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES

## Abstract

Provided herein are methods, processes and apparatuses for non-invasive assessment of genetic variations.

## Claims

### Claim 1

A system comprising:
I. a sequencing module that performs a non-targeted massively parallel sequencing process to generate nucleic acid sequence reads from a test sample from a pregnant subject, wherein the sequence reads are reads of circulating cell-free nucleic acid from (i) a maternal genome and (ii) a fetal genome; and
II. one or more microprocessors and memory, which memory comprises instructions executable by the one or more microprocessors, and which instructions executable by the one or more microprocessors are configured to:
(a) map the nucleic acid sequence reads to genomic sections of a reference genome, count the nucleic acid sequence reads mapped to the genomic sections of the reference genome, and normalize the counts of the nucleic acid sequence reads mapped to the genomic sections of a reference genome, thereby providing normalized counts for the genomic sections, wherein the sequence reads that are mapped, counted, and normalized comprise reads of the maternal genome and the fetal genome;
(b) identify a first elevation of the normalized counts for a first set of genomic sections significantly different than a second elevation of the normalized counts for a second set of genomic sections;
(c) determine a copy number variation in the maternal genome for the first set of genomic sections based on the identified first and second elevations;
(d) determine a categorization for the copy number variation in the maternal genome by comparing the first elevation to expected elevations for predetermined copy number variation categorizations; and
(e) determine a fetal fraction of the circulating cell-free nucleic acid according to the first elevation and a deviation from an expected elevation of normalized counts for the categorization.

### Claim 2 (depends on 1)

The system of claim 1, wherein the categorization of the copy number variation in the maternal genome is determined in (d) by comparing the first elevation to an expected elevation range for the respective predetermined copy number variation categorizations.

### Claim 3 (depends on 2)

The system of claim 2, wherein each expected elevation range is determined according to an expected elevation for each of the predetermined copy number variation categorizations and an uncertainty value.

### Claim 4 (depends on 3)

The system of claim 3, wherein the expected elevation for the respective predetermined copy number variation categorizations is determined according to a normalized reference value (NRV) and an expected elevation constant of each of the predetermined copy number variation categorizations.

### Claim 5 (depends on 4)

The system of claim 4, wherein the NRV is determined according to a reference elevation.

### Claim 6 (depends on 5)

The system of claim 5, wherein the reference elevation is the second elevation.

### Claim 7 (depends on 6)

The system of claim 6, wherein the second elevation is an average, mean or median elevation for the second set of genomic sections.

### Claim 8 (depends on 4)

The system of claim 4, wherein the expected elevation constant for a maternal homozygous duplication is 2, a maternal heterozygous duplication is 1.5, a maternal heterozygous deletion is 0.5, and a maternal homozygous deletion is zero.

### Claim 9 (depends on 3)

The system of claim 3, wherein the uncertainty value is a standard deviation or median absolute deviation (MAD).

### Claim 10 (depends on 1)

The system of claim 1, wherein the copy number variation in the maternal genome determined in (c) is a maternal microduplication or a maternal microdeletion.

### Claim 11 (depends on 10)

The system of claim 10, wherein the categorization for the copy number variation in the maternal genome determined in (d) is chosen from a maternal homozygous microduplication, a maternal heterozygous microduplication, a maternal heterozygous microdeletion, and a maternal homozygous microdeletion.

### Claim 12 (depends on 1)

The system of claim 1, wherein the second set of genomic sections includes substantially no copy number variation.

### Claim 13 (depends on 1)

The system of claim 1, wherein the fetal fraction is determined as twice the difference between the first elevation and an expected elevation of the copy number variation categorization.

### Claim 14 (depends on 1)

The system of claim 1, wherein the instructions executable by the one or more microprocessors are further configured to output a classification of presence or absence of a fetal chromosome aneuploidy for the test sample according to the normalized counts in (a), when the fetal fraction determined in (e) is at or above a fetal fraction threshold.

### Claim 15 (depends on 14)

The system of claim 14, wherein the fetal chromosome aneuploidy is chosen from a chromosome 13 trisomy, a chromosome 18 trisomy, a chromosome 21 trisomy, or a combination thereof.

### Claim 16 (depends on 14)

The system of claim 14, wherein the fetal fraction threshold is about 4%.

### Claim 17 (depends on 1)

The system of claim 1, wherein the sequencing process is performed at about 1-fold coverage.

### Claim 18 (depends on 1)

The system of claim 1, wherein the sequencing process is performed at less than 1-fold coverage.

### Claim 19 (depends on 1)

The system of claim 1, wherein thousands to millions of nucleic acid sequence reads mapped to genomic sections of a reference genome are normalized in (a).

### Claim 20 (depends on 1)

The system of claim 1, wherein the non-targeted massively parallel sequencing process generates thousands to millions of sequence reads, and wherein the instructions executable by the one or more microprocessors are further configured to map the thousands to millions of sequence reads to genomic sections of the reference genome.