Single Circulating-Tumor-Cell-Targeted Sequencing to Identify Somatic Variants in Liquid Biopsies in Non-Small-Cell Lung Cancer Patients

Abstract

Non-small-cell lung cancer (NSCLC) accounts for most cancer-related deaths worldwide. Liquid biopsy by a blood draw to detect circulating tumor cells (CTCs) is a tool for molecular profiling of cancer using single-cell and next-generation sequencing (NGS) technologies. The aim of the study was to identify somatic variants in single CTCs isolated from NSCLC patients by targeted NGS. Thirty-one subjects (20 NSCLC patients, 11 smokers without cancer) were enrolled for blood draws (7.5 mL). CTCs were identified by immunofluorescence, individually retrieved, and DNA-extracted. Targeted NGS was performed to detect somatic variants (single-nucleotide variants (SNVs) and insertions/deletions (Indels)) across 65 oncogenes and tumor suppressor genes. Cancer-associated variants were classified using OncoKB database. NSCLC patients had significantly higher CTC counts than control smokers (p = 0.0132; Mann–Whitney test). Analyzing 23 CTCs and 13 white blood cells across seven patients revealed a total of 644 somatic variants that occurred in all CTCs within the same subject, ranging from 1 to 137 per patient. The highest number of variants detected in ≥1 CTC within a patient was 441. A total of 18/65 (27.7%) genes were highly mutated. Mutations with oncogenic impact were identified in functional domains of seven oncogenes/tumor suppressor genes (NF1, PTCH1, TP53, SMARCB1, SMAD4, KRAS, and ERBB2). Single CTC-targeted NGS detects heterogeneous and shared mutational signatures within and between NSCLC patients. CTC single-cell genomics have potential for integration in NSCLC precision oncology.

Accurate isolation and detection of circulating tumor cells using enrichment-free multiparametric high resolution imaging

“…Several studies have utilized the AccuCyte-CyteFinder platform to detect CTCs in various cancers such as prostate, breast, and lung ( 9 , 10 , 26 , 28 , 29 , 31 – 33 ). This study is the first to examine CTCs using this platform in thyroid cancer.…”

“…Furthermore, the platform has a single-cell retrieval device, CellPicker™, where identified cells can be picked for single-cell molecular characterization. This includes genomic analysis such as TP53 mutations in breast cancer cells ( 27 ), APC mutations in colorectal cancer ( 36 ), targeted mutational panels in prostate ( 31 ) and lung cancer ( 33 ); and single-cell RNA sequencing as demonstrated in breast cancer ( 29 ).…”

Targeting immune cell types of tumor microenvironment to overcome resistance to PD-1/PD-L1 blockade in lung cancer

“…NSCLC patients with overexpression of PTCH1 displayed a better outcome ( Herreros-Pomares et al, 2022 ). Moreover, PTCH1 expression was found to be correlated with NSCLC development ( Barbirou et al, 2022 ). One circulating tumor cell NGS assay in early-stage lung cancer patients showed that more than 50% of lung cancer patients presented four common mutations, including Notch1, EGFR, IGF2, and PTCH1 ( Wan et al, 2021 ).…”

Molecules at Play in Cancer

“…The 24 chapters present the contributions of 166 researchers from 16 countries: Brazil (14), China (12), Ecuador (2), Greece (10), Hungary (8), Italy (4), Japan (25), Korea (29), Mexico (11), Poland (5), Portugal (2), Russia (2), Switzerland (7), the UK (1) and the USA (27). The articles reported new molecular findings regarding the following types of cancer: blood [ 1 , 2 ], breast [ 3 , 4 , 5 ], kidney [ 6 ], liver [ 7 , 8 ], lung [ 9 , 10 , 11 , 12 ], ovary [ 10 , 13 , 14 ], prostate [ 15 ], soft tissue [ 16 ], gastric [ 17 , 18 ], testicular [ 19 ], cervical [ 20 , 21 ], and adrenal gland [ 22 ].…”

“…For instance, Armakolas et al [ 8 ] compared the proteome profiles, expression of selected miRNAs and abundance of circulating tumor cells among a group of 56 people with advanced and 33 people with early hepatocellular carcinoma, 28 with cirrhosis and 5 healthy controls. Barbirou et al sequenced circulating tumor cells to identify somatic variants in people with non-small-cell lung cancer [ 9 ]. An interesting study by Bel’skaya et al [ 5 ] compared the saliva “omic” characteristics of 487 females with breast cancer with those of 298 healthy controls.…”