CNV Detection via Statistical Normalization of Chromosome Doses
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Solution Overview
Problem
Current methods for detecting copy number variations (CNVs) in genetic material, particularly for noninvasive prenatal diagnostics and cancer monitoring, face limitations due to insufficient sensitivity and sequencing bias, necessitating more reliable and sensitive approaches for diagnosing chromosomal aneuploidies without invasive procedures.
Innovation Solution
A statistical method that accounts for process-related and interchromosomal variability, involving sequencing of nucleic acids to calculate single chromosome doses and compare them to threshold values, enabling the detection of fetal aneuploidies and CNVs in maternal test samples using sequence tags and normalizing chromosome sequences.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Reliability
If conventional molecular-cytogenetic methods (FISH, QF-PCR, array-CGH) are used to detect copy number variations, then diagnostic capability is improved, but the methods require invasive procedures and cell culture which increases complexity and limits applicability
Solution Approach 1:
The invention extracts and sequences only the specific chromosomal regions of interest rather than analyzing entire chromosomes or requiring cell culture. This extraction approach enables direct detection of copy number variations from circulating cell-free DNA without invasive procedures or complex cell culture steps, resolving the contradiction between diagnostic reliability and procedure complexity
Solution Approach 2:
The invention replaces the mechanical and manual procedures of FISH, QF-PCR, and array-CGH with automated next-generation sequencing technology. This substitution eliminates the need for invasive sampling and cell culture while maintaining or improving diagnostic accuracy, thereby reducing procedure complexity
2Ease of operation
If circulating cell-free DNA is used for noninvasive detection, then invasiveness is reduced, but sensitivity is insufficient due to limited levels of cfDNA
Solution Approach 1:
The invention segments the genome into specific chromosomal regions of interest and sequences only those segments rather than attempting to analyze all circulating cell-free DNA. This segmentation approach concentrates sequencing depth on target regions, achieving sufficient sensitivity despite low overall cfDNA levels, while maintaining the noninvasive advantage
Solution Approach 2:
The invention changes the parameter of sequencing depth by performing deep targeted sequencing of specific chromosomal regions rather than shallow whole-genome sequencing. This parameter change enables detection of copy number variations in the low levels of circulating cell-free DNA present in maternal plasma, resolving the sensitivity limitation
3Reliability
If whole genome sequencing is performed to detect CNVs, then comprehensive coverage is improved, but sequencing bias and cost increase
Solution Approach 1:
The invention applies local quality by providing high-coverage sequencing specifically for chromosomal regions known to contain copy number variations or diagnostic markers, rather than uniform whole-genome sequencing. This approach achieves comprehensive detection of clinically relevant CNVs while reducing sequencing bias and cost by concentrating resources on high-value regions
Data Source
AI summary
The invention provides a method for determining copy number variations (CNV) of a sequence of interest in a test sample that comprises a mixture of nucleic acids that are known or are suspected to differ in the amount of one or more sequence of interest. The method comprises a statistical approach that accounts for accrued variability stemming from process-related, interchromosomal and inter-sequencing variability. The method is applicable to determining CNV of any fetal aneuploidy, and CNVs known or suspected to be associated with a variety of medical conditions. CNV that can be determined according to the method include trisomies and monosomies of any one or more of chromosomes 1-22, X and Y, other chromosomal polysomies, and deletions and/or duplications of segments of any one or more of the chromosomes, which can be detected by sequencing only once the nucleic acids of a test sample.