Abstract 3375: Ultrasensitive ctDNA minimal residual disease monitoring in early NSCLC with PhasED-Seq
Authors: Isbell James M. , Li Bob T. , Razavi Pedram , Reis-Filo Jorge , Liu Si-Yang , Selenica Pier , Adusumilli Prasad , Bott Matthew , Jones David R. , Rusch Valerie W. , Sihag Smita , Buonocore Darren J. , Jee Justin , Lebow Emily , Gomez Daniel , Rimner Andreas , Santini Fernando C. , Rudin Charles M. , Eichholz Jordan E. , Martinez Andres , Alerte Daphne , Hogan Gregory J. , Schultz Andre , Schuyler Ronald P. , Roff Alanna , Hite Dustin , Chabon Jacob J. , Kurtz David M. , Alizadeh Ash A. , Diehn Maximilian 
Published: April 2023

Abstract
Background: Circulating tumor DNA (ctDNA) minimal residual disease (MRD) detection is a promising approach for personalization of adjuvant therapy in non-small cell lung cancer (NSCLC). First generation ctDNA MRD assays that employ tumor-informed approaches to track single nucleotide variants (SNVs) have limits of detection (LOD95) of ~1E-4 and have high positive predictive values for recurrence. However, they have suboptimal clinical sensitivity, with false negative results at the completion of therapy in most patients who will ultimately recur. PhasED-Seq is a novel ctDNA MRD method that tracks multiple “phased” variants (PVs) within individual DNA fragments with a LOD95 ~100-fold better than first generation assays. Here we report PhasED-Seq ctDNA MRD results for the first prospective cohort of early stage NSCLC patients.
Methods: Tumor tissues (n=46), PBMCs (n=46) and plasma samples (n=169) from 46 Stage I-III NSCLC patients treated with curative intent were prospectively collected at Memorial Sloan Kettering Cancer Center. All patients underwent resection and received neoadjuvant +/- adjuvant therapy (n=14), adjuvant therapy only (n=17), or neither (n=15). Samples were analyzed in Foresight Diagnostics' CLIA laboratory (Aurora, CO) using personalized PhasED-Seq. Briefly, PVs were identified via whole genome sequencing of tumors and matched blood germline DNA. Custom capture panels targeting PVs were synthesized and used to assess MRD status in pre-, on- and post-treatment plasma samples. Detection of ctDNA MRD was assessed at a post-treatment landmark, defined as the first post-therapy sample or when not available the last post-surgical sample taken during therapy. To enable comparisons, the same plasma samples were analyzed using an SNV-based ctDNA MRD approach.
Results: PVs were identified in tumor tissue from all 46 patients. Across all plasma samples PhasED-Seq achieved a median LOD95 of 1.3E-6 and as low as 2.5E-7. Of 74 plasma samples with detectable ctDNA, 38 (51%) contained concentrations below 1E-4 and the lowest level of ctDNA MRD detected was 1.7E-7. For post-treatment landmark samples (n=45), the median time from end of therapy was 2 months. Cancer recurred in all patients (n=10) with detectable MRD at the landmark. Furthermore, PhasED-Seq better stratified freedom from recurrence (log-rank p=3E-8, Cox HR=10.8) than the SNV-based approach (log-rank p=0.08, Cox HR=2.5) and detected MRD at the landmark in more patients who ultimately recurred (56% vs 28%). PhasED-Seq also achieved longer lead times, including detecting MRD in 66% of samples collected 12 to 24 months prior to recurrence versus only 33% using SNV-based monitoring.
Conclusion: PhasED-Seq achieves ctDNA detection below 1 part per million and appears to be significantly more sensitive than SNV-based MRD monitoring. These results suggest that PhasED-Seq is a promising approach for use in risk adapted trials in early stage NSCLC.
Citation Format: James M. Isbell, Bob T. Li, Pedram Razavi, Jorge Reis-Filo, Si-Yang Liu, Pier Selenica, Prasad Adusumilli, Matthew Bott, David R. Jones, Valerie W. Rusch, Smita Sihag, Darren J. Buonocore, Justin Jee, Emily Lebow, Daniel Gomez, Andreas Rimner, Fernando C. Santini, Charles M. Rudin, Jordan E. Eichholz, Andres Martinez, Daphne Alerte, Gregory J. Hogan, Andre Schultz, Ronald P. Schuyler, Alanna Roff, Dustin Hite, Jacob J. Chabon, David M. Kurtz, Ash A. Alizadeh, Maximilian Diehn. Ultrasensitive ctDNA minimal residual disease monitoring in early NSCLC with PhasED-Seq [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 3375.