Investigating ecDNA Dynamics in a Lapatinib-Resistant NCI-H2170 Lung Cancer Model

Extrachromosomal DNA (ecDNA) is an oncogene amplification mechanism that leads to intratumoral heterogeneity, therapeutic resistance, and poor clinical outcomes in various cancers, including glioblastoma and breast, lung, colon, and stomach cancer. Unlike chromosomal amplifications, ecDNAs are 1-3 Mbp circular structures that lack centromeres. Thus, ecDNAs undergo uneven segregation during mitosis. This stochastic segregation results in high copy number variations within tumor cell populations. This study utilizes the NCI-H2170 non-small cell lung cancer (NSCLC) cell line, a model with MYC and ERBB2 co-amplification on ecDNA and high ecDNA counts, to investigate the longitudinal dynamics of ecDNAs under pressure from the HER2/EGFR inhibitor lapatinib. By establishing a lapatinib-resistant subline through an 8-week treatment protocol and a subsequent 8-week lapatinib-withdrawal group, this research quantifies the shifts in ecDNA copies using fluorescence in situ hybridization (FISH) and an ecDNA identification pipeline previously developed by the Brunk Lab using Microscopic Image Analyzer (MIA). Median ecDNA counts fell from 295.50 in the 72-hour DMSO control to 66.50 in the lapatinib-resistant group and decreased to 57.00 after withdrawal. While both experimental groups differed significantly from the control (p < 0.01), no significant difference was observed between the resistant and withdrawn groups. This persistence of decreased ecDNA counts after lapatinib removal suggests that a long-term genomic alteration was introduced by lapatinib pressure. This report argues that cancer cells use ecDNA dynamics as a strategy to optimize oncogenic dosage to balance signaling requirements with metabolic fitness, ensuring long-term survival even after therapy removal.