[ Title ]
- Genomic variations in plasma cell free DNA differentiate early stage
lung cancers from normal controls
[ Abstract ]
- Objectives: Cell free tumor DNA (cfDNA) circulating in blood has a great
potential as biomarker for cancer clinical management. The objective of
this study is to evaluate if cfDNA in blood plasma is detectable in
early stage lung cancer patients.
Materials and methods: We extracted cfDNAs and tumor tissue DNAs from 8
lung adenocarcinoma patients. We also extracted cfDNAs from 8 normal
controls. To evaluate copy number variations (CNV) and identify
potential mutations, we performed low pass whole genome sequencing and
targeted sequencing of 50 cancer genes. To accurately reflect the
tumor-associated genomic abnormality burden in plasma, we developed a
new scoring algorithm, plasma genomic abnormality (PGA) score, by
summarizing absolute log2 ratios in most variable genomic regions. We
performed digital PCR and allele-specific PCR to validate mutations
detected by targeted sequencing.
Results and conclusions: The median yield of cfDNA in 400ul plasma was
4.9ng (range 2.25-26.98 ng) in patients and 232 ng (range 1.30-2.81 ng)
in controls (p=0.003). The whole genome sequencing generated
approximately 20 million mappable sequence reads per subject and 5303
read counts per 1 Mb genomic region. Log2 ratio-based CNV analysis
showed significant chromosomal abnormality in cancer tissue DNAs and
subtle but detectable differences in cfDNAs between patients and
controls. Genomic abnormality analysis showed that median PGA score was
9.28 (7.38-11.08) in the 8 controls and 19.50 (5.89-64.47) in the 8
patients (p=0.01). Targeted deep sequencing in tumor tissues derived
from the 8 patients identified 14 mutations in 12 different genes. The
PCR-based assay confirmed 3 of 6 selected mutations in cfDNAs. These
results demonstrated that the PGA score and cfDNA mutational analysis
could be useful tool for the early detection of lung cancer. These
blood-based genomic and genetic assays are noninvasive and may
sensitively distinguish early stage disease when combined with other
existing screening strategies including
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