Vascular invasion-associated gene expression is detectable in pre-surgical biopsies of stage I lung adenocarcinoma [Spatial Transcriptomics]

Microscopic vascular invasion (VI) predicts recurrence and benefit from lobectomy in stage I lung adenocarcinoma (LUAD) but cannot be accurately predicted before surgery. Thus, biomarkers that identify this aggressive tumor subset are needed. Here, we show that VI in stage I LUAD is associated with reproducible gene expression programs detectable beyond the invasive focus. Using bulk RNA sequencing of 162 resected tumors and spatial transcriptomics in a subset, we identify and characterize a VI-associated gene signature. A predictor derived from this signature is associated with VI and recurrence in an independent validation cohort and is robust to intra-tumor heterogeneity in multiregional sampling data. In a cohort of pre-surgical biopsies, predictor scores correlate with matched resections and show promising discrimination of VI. These findings indicate that VI-associated transcriptional changes extend across the tumor and are detectable in limited biopsy material, supporting further validation for preoperative risk stratification in stage I LUAD. Overall design: 16 samples from 14 patient tumors within the validation cohort were selected for 10x Genomics Visium for FFPE spatial whole-transcriptome profiling. All samples were selected to be stage IA/IB at the time of collection except for one that was upstaged to stage IIA under the 8th TNM edition (this sample was excluded from the bulk RNA-seq validation cohort when evaluating the performance of VI predictor scores). Samples were chosen based on 1) the presence of pathological features of interest (e.g., VI foci, representative LUAD histologic patterns) and 2) more than 50% of RNA fragments being greater than 200 nucleotides (DV200) after extraction with the AllPrep DNA/RNA Universal Kit (Qiagen) during bulk RNA-seq library preparation.