Figure S1-S7 from Perioperative Dynamic Changes in Circulating Tumor DNA in Patients with Lung Cancer (DYNAMIC)

Figure S1. (A) Strategy for plasma detection of oncogenic mutations. The exonic positions and nature of the hotspot mutations are shown. Two sets of bidirectional reverse primers (arrows) were positioned to robust sequences on either side of each mutation site to target and amplify each allele by inverse PCR. For detection of ALK fusions, targeting primers were progressively spaced across exon 19, intron 19, and exon 20, spanning the known ALK breakpoint region. (B) Hot Spot Gene Mutations and Diagnostic cSMART Primers. Figure S2. Comparison of the distributions of driver mutations identified in 26 matched tDNA and plasma ctDNA samples. Thirty-two mutations were detected in the plasma at time A, including 14 TP53 mutations, 8 EGFR mutations, 6 PIK3CA mutations, 3 KRAS mutations and 1 ALK mutation. Five patients had two mutations simultaneously in the plasma ctDNA: case 3 had PIK3CA and EGFR mutations; case 6 had EGFR and TP53 mutations; case 15 had two different TP53 mutations, namely, a base substitution mutation in exon 5 and a base substitution mutation in exon 8; case 17 had EGFR and TP53 mutations; and case 20 had KRAS and PIK3CA mutations simultaneously. A single mutation was detected in the other 21 cases. For the corresponding tumor tissues, a total of 29 gene mutations were detected: 14 TP53 mutations, 8 EGFR mutations, 5 PIK3CA mutations and 2 KRAS mutation. More than one mutation was detected in the tumors of 5 patients, which was completely consistent with the ctDNA results. (A) Numbers of gene mutations in tDNA vs. plasma ctDNA from matched sample pairs. (B) The inner circle shows the distribution of gene mutations, and the outer ring shows specific alterations in amino acids. Figure S3. Excluded two positive plasma A patients with CHIP validated by normal lung tissue and white blood cell. Figure S4. The plasma ctDNA concentration at each time point during the perioperative period (from time A to time P2). The X-axis represents the logarithmic value of time. Figure S5. Longitudinal ctDNA profiles of non-relapse or non-progression cases. Red curve represents the fluctuation of ctDNA concentration over time. The X-axis represents the days of post-operation. The Y-axis represents the ctDNA MAF (%). In this figure, rectangular frames represent the treatment that patients have received, as shown at the bottom, green frame: surgical treatment; yellow frame: chemotherapy; blue frame: radiotherapy and red frame: target therapy. Figure S6. The RFS (left) and OS (right）between different ctDNA statuses at time P3 (Three cases with no blood samples at this time point were excluded). Figure S7. The RFS between patients who received postoperative therapy or not based on different strategy. (A) Patients with stage II, III and above lung cancer according to TNM stage in our study. (B) Patients with positive MRD detection in our study. (C) Patient with positive ctDNA at time P2.