Supplementary Figures S1-S13 from Cell-Free Urine and Plasma DNA Mutational Analysis Predicts Neoadjuvant Chemotherapy Response and Outcome in Patients with Muscle-Invasive Bladder Cancer

Christensen, Emil; Nordentoft, Iver; Birkenkamp-Demtröder, Karin; Elbæk, Sara K.; Lindskrog, Sia V.; Taber, Ann; Andreasen, Tine G.; Strandgaard, Trine; Knudsen, Michael; Lamy, Philippe; Agerbæk, Mads; Jensen, Jørgen B.; Dyrskjøt, Lars

doi:10.1158/1078-0432.22490092.v1

ccr-22-3250_supplementary_figures_s1-s13_suppfs1-fs13.pdf (5.69 MB)

Supplementary Figures S1-S13 from Cell-Free Urine and Plasma DNA Mutational Analysis Predicts Neoadjuvant Chemotherapy Response and Outcome in Patients with Muscle-Invasive Bladder Cancer

journal contribution

posted on 2023-04-01, 00:22 authored by Emil Christensen, Iver Nordentoft, Karin Birkenkamp-Demtröder, Sara K. Elbæk, Sia V. Lindskrog, Ann Taber, Tine G. Andreasen, Trine Strandgaard, Michael Knudsen, Philippe Lamy, Mads Agerbæk, Jørgen B. Jensen, Lars Dyrskjøt

Supplementary Fig. 1. Overview of sample availability and in-silico analysis of the limit of detection for the applied methodology.Supplementary Fig. 2. Comparison between mutations observed in primary and residual tumors at radical cystectomy (RC).Supplementary Fig. 3. Mean sample VAF levels for plasma, urine pellet and urine supernatant split by tdDNA call status in other paired sample types from the same clinical visit.Supplementary Fig. 4. Association between mean sample VAF.Supplementary Fig. 5. Association between mean VAFSupplementary Fig. 6. Urine dipstick measurements compared to the amount of DNA extracted from urine supernatants collected at the same time (+/- 1 day).Supplementary Fig. 7. Urine dipstick measurements compared to the urinary tumor DNA (utDNA) call status in urine supernatants collected at the same time (+/- 1 day).Supplementary Fig. 8. Urine dipstick measurements compared to the urinary tumor DNA (utDNA) level in urine supernatants collected at the same time (+/- 1 day).Supplementary Fig. 9. Mean sample VAF levels for all patients with detectable tumor DNA split by sample type. Tumor stage evaluation was based on the TURB specimen and tumor DNA level based on samples collected before TURB.Supplementary Fig. 10. Kaplan-Meier survival analysis of recurrence-free survival (RFS) stratified by tumor DNA status for all sample types.Supplementary Fig. 11. Tumor DNA levels compared to NAC response.Supplementary Fig. 12. Kaplan-Meier survival analysis of recurrence-free survival (RFS) stratified by tumor DNA level for all sample types.Supplementary Fig. 13. a-b) Association between tumor DNA dynamics and neoadjuvant chemotherapy (NAC) response for a) urine pellet and b) urine supernatant. Tumor DNA clearance was defined as tumor DNA going from detectable to non-detectable and tumor DNA remains was defined as tumor DNA remaining detectable. c-d) Kaplan-Meier survival analysis of tumor DNA dynamics groups and recurrence-free survival (RFS) for c) urine pellet and d) urine supernatant.

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ARTICLE ABSTRACT

To investigate the use of plasma and urine DNA mutation analysis for predicting neoadjuvant chemotherapy (NAC) response and oncological outcome in patients with muscle-invasive bladder cancer. Whole-exome sequencing of tumor and germline DNA was performed for 92 patients treated with NAC followed by radical cystectomy (RC). A custom NGS-panel capturing approximately 50 mutations per patient was designed and used to track mutated tumor DNA in plasma and urine. A total of 447 plasma samples, 281 urine supernatants, and 123 urine pellets collected before, during, and after treatment were analyzed. Patients were enrolled from 2013 to 2019, with a median follow-up time of 41.3 months after RC. We identified tumor DNA before NAC in 89% of urine supernatants, 85% of urine pellets, and 43% of plasma samples. Tumor DNA levels were higher in urine supernatants and urine pellets compared with plasma samples (P < 0.001). In plasma, detection of circulating tumor DNA (ctDNA) before NAC was associated with a lower NAC response rate (P < 0.001). Detection of tumor DNA after NAC was associated with lower response rates in plasma, urine supernatant, and urine pellet (P < 0.001, P = 0.03, P = 0.002). Tumor DNA dynamics during NAC was predictive of NAC response and outcome in urine supernatant and plasma (P = 0.006 and P = 0.002). A combined measure from plasma and urine supernatant tumor DNA dynamics stratified patients by outcome (P = 0.003). Analysis of tumor DNA in plasma and urine samples both separately and combined has a potential to predict treatment response and outcome.