Journal: Annals of Oncology

Article Title: Distinct subclonal tumour responses to therapy revealed by circulating cell-free DNA

doi: 10.1093/annonc/mdw278

Figure Lengend Snippet: Levels of KIT p.L576P in the circulating cell-free DNA (cfDNA) respond to imatinib. (A) Overview of patient's treatment history, including information on initial diagnosis (35 weeks before follow-up) and surgery. Organ denominations indicate sites of metastatic disease as detected by computed tomography (CT) scans. Red font, progression; green font, response; DTIC, dacarbazine; Ipi., ipilimumab; Pembro., pembrolizumab. (B) Routine CT-generated images of the liver (top panels) and inguinal lymph node (bottom panels) at the indicated times. DTIC, dacarbazine; carbo./pacli., carboplatin/paclitaxel. (C) RECIST 1.1 measurements of a segment II liver metastasis and the inguinal (ing.) lymph node lesion during treatment with dacarbazine (D), imatinib (Im), ipilimumab (Ip), pembrolizumab (Pem) and carboplatin/paclitaxel (C/P) corresponding to (B). (D) Prospective quantification of KIT p.L576P VAF in cfDNA up to week 37. DTIC, dacarbazine.

Article Snippet: After adding 90 μl of water, 8.8 μl of the reaction was combined with 11 μl ddPCR Supermix for Probes (No dUTP) (Bio-rad, Hercules, CA), 1.1 μl of KIT (Hs02812715_cn) or NLGN4X (Hs02584007_cn)-specific probe and 1.1 μl of TERT TaqMan Copy Number Reference Assay (all Thermo Fisher Scientific, Waltham, MA).

Techniques: Biomarker Discovery, Computed Tomography, Generated

Journal: Annals of Oncology

Article Title: Distinct subclonal tumour responses to therapy revealed by circulating cell-free DNA

doi: 10.1093/annonc/mdw278

Figure Lengend Snippet: Circulating cell-free DNA (cfDNA) reveals two tumour subclones with distinct responses to therapy. (A) Venn diagram showing single-nucleotide variations identified by whole-exome sequencing (WES) of cfDNA collected at week 37. The diagram shows the mutations detected in the 25 or 10 ng input samples, with common mutations in the intersection. KIT p.L576P is highlighted in red to confirm its identification in both WES runs. (B) Correlation of variant allele frequencies (VAFs) detected by WES or targeted sequencing in the week 37 cfDNA sample. WES-based VAFs represent average values of the two input DNA amounts. (C) VAFs of 14 mutations (see legend within the figure) in cfDNA from samples collected at the indicated times. VAFs of mutations in clusters 1 and 2 are connected by black and blue lines, respectively, and the treatments administered are indicated above the graph. DTIC, dacarbazine; Ipi., ipilimumab. (D) Copy number variation by chromosome (chr.) based on WES from the two different input cfDNA amounts isolated at week 37 of follow-up. Grey and red dots indicate allele loss (both and one, respectively), green dots indicate normal copy number state and blue and light blue dots indicate copy number gains (three and four or more copies, respectively).

Article Snippet: After adding 90 μl of water, 8.8 μl of the reaction was combined with 11 μl ddPCR Supermix for Probes (No dUTP) (Bio-rad, Hercules, CA), 1.1 μl of KIT (Hs02812715_cn) or NLGN4X (Hs02584007_cn)-specific probe and 1.1 μl of TERT TaqMan Copy Number Reference Assay (all Thermo Fisher Scientific, Waltham, MA).

Techniques: Sequencing, Variant Assay, Isolation

Journal: Annals of Oncology

Article Title: Distinct subclonal tumour responses to therapy revealed by circulating cell-free DNA

doi: 10.1093/annonc/mdw278

Figure Lengend Snippet: Analysis of the primary tumour reveals tumour heterogeneity. (A) Single-nucleotide variations in chromosomal DNA from the diagnostic biopsy of the primary tumour and the radical surgery 2 months after the initial biopsy. n.d., not detected. (B) Copy number determination of KIT and NLGN4X in the diagnostic biopsy and in tissue from the subsequent surgical excision when compared with germline DNA by using droplet digital PCR. *** P < 0.001.

Article Snippet: After adding 90 μl of water, 8.8 μl of the reaction was combined with 11 μl ddPCR Supermix for Probes (No dUTP) (Bio-rad, Hercules, CA), 1.1 μl of KIT (Hs02812715_cn) or NLGN4X (Hs02584007_cn)-specific probe and 1.1 μl of TERT TaqMan Copy Number Reference Assay (all Thermo Fisher Scientific, Waltham, MA).

Techniques: Diagnostic Assay, Digital PCR