In:
Cancer Research, American Association for Cancer Research (AACR), Vol. 80, No. 4_Supplement ( 2020-02-15), p. P5-01-14-P5-01-14
Abstract:
Background: To gain comprehensive insights into the genomic complexity useful for therapy management in metastatic breast cancer (MBC), we aimed to isolate and analyze genomic DNA (gDNA) from circulating tumor cells (CTCs) and matched cell-free DNA (cfDNA) from a minimized blood volume. Methods: EDTA blood was drawn from 27 MBC patients with hormone receptor-positive and HER2 negative primary tumors at the time of disease progression. CTCs and CTC mRNA were isolated from 2 × 5 ml whole blood using the AdnaTest EMT2/StemCell Select/Detect. Plasma of CTC-depleted blood was used for cfDNA isolation. gDNA from CTCs was isolated from the mRNA-depleted CTC lysates using the AllPrep DNA/RNA Nano Kit prototype. CTC gDNA and cfDNA were analyzed with a customized QIAseq Targeted DNA Panel for Illumina with unique molecular indices (UMIs) analyzing AKT1, AR, BRCA1, BRCA2, EGFR, ERCC4, ERBB2, ERBB3, ESR1, FGFR1, KRAS, MUC16, PIK3CA, PIK3R1, PTEN, PTGFR and TGFB1. The library preparation protocol was slightly modified for the usage of CTC gDNA using the entire amount of the CTC gDNA eluate as input without pre-quantification followed by fragmentation and ligation with an increased number of adapters. All libraries were analyzed by paired-end sequencing on the Illumina NextSeq Sequencer with a NextSeq 550 System High-Output Kit, 2 × 150 bp reads with a mean coverage of 11000 x (CTC gDNA) and 8000 x (cfDNA). Consumables: QIAGEN, Germany. Results: Isolation of CTC gDNA and cfDNA was successfully established in a condensed workflow. The UMI coverage observed for cfDNA (2000 x) differed from the UMI coverage for CTC gDNA (500 x), resulting in dramatically increased lowest detectable allele frequency (AF) called with a confidence of 90% in CTC gDNA compared to cfDNA. On average, 6 CTC gDNA variants and 2 cfDNA variants were detected in each patient. Most variants were found in the MUC16 gene in both analytes. BRCA2 variants were the second most prevalent variants in CTC gDNA and cfDNA. PIK3CA and ESR1 variants were less common in CTC gDNA compared with cfDNA, while ERBB2 variants were only detected in CTC gDNA. 57% of all cfDNA variants (29/51) were recovered in the matched CTC gDNA, while 89% of all variants were unique in either CTC gDNA (125 variants) or cfDNA (22 variants). On average, the cfDNA variants with low AF (mean 15%) were not detected in CTC gDNA, while the cfDNA variants that were also found in CTC gDNA exhibited a mean AF of 44%. Similarly, the unique CTC gDNA variants had a mean AF of 23%, while the shared variants were prevalent with a mean AF of 47% in the CTC gDNA fraction. Comparing the patients, 11%/14% of the single CTC gDNA / cfDNA variants were detected in more than one patient, hence the majority of variants was patient-specific. The portion of patients without detectable cfDNA variants or CTC gDNA variants was 15% (4/27) / 11% (3/27), but combined analysis of CTC gDNA and cfDNA identified variants in 26 of all 27 MBC patients (96%). Conclusion: cfDNA and CTC gDNA showed additive variant information. Thus, it is advised to assess cfDNA and CTC gDNA variants to receive a comprehensive genomic picture that might enable to identify the most suitable therapy regimen in each individual patient in the future. Citation Format: Corinna Keup, Ann-Kathrin Bittner, Oliver Hoffmann, Mitra Tewes, Markus Storbeck, Peter Hahn, Siegfried Hauch, Markus Sprenger-Haussels, Rainer Kimmig, Sabine Kasimir-Bauer. The combined blood analysis of cell-free DNA and genomic DNA of circulating tumor cells reveals additive value in hormone receptor-positive, HER2-negative metastatic breast cancer patients [abstract]. In: Proceedings of the 2019 San Antonio Breast Cancer Symposium; 2019 Dec 10-14; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2020;80(4 Suppl):Abstract nr P5-01-14.
Type of Medium:
Online Resource
ISSN:
0008-5472
,
1538-7445
DOI:
10.1158/1538-7445.SABCS19-P5-01-14
Language:
English
Publisher:
American Association for Cancer Research (AACR)
Publication Date:
2020
detail.hit.zdb_id:
2036785-5
detail.hit.zdb_id:
1432-1
detail.hit.zdb_id:
410466-3
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