Molecular subtyping of stage I lung adenocarcinoma via molecular alterations in pre-invasive lesion progression

Patients with adenocarcinoma in situ (AIS) and minimally invasive (MIA) lung adenocarcinoma (LUAD) are curable by surgery, whereas 20% stage I patients die within five years after surgery. We hypothesize that poor-prognosis stage I patients may exhibit key molecular characteristics deviating from AIS/MIA. Therefore, we tried to reveal molecularly and prognostically distinct subtypes of stage I LUAD by applying key molecular alterations from AIS/MIA to invasive LUAD progression.

We successfully revealed two molecularly and prognostically distinct subtypes of stage I LUAD by applying key molecular alterations from AIS/MIA to invasive LUAD progression. Our model may help reliably identify high-risk stage I patients for more intensive post-surgery treatment.

The RNA and whole-exome sequencing data of 197 tumor-normal matched samples from patients with AIS, MIA, and invasive LUAD were analyzed. ddPCR quantified 202 samples from 182 patients at the absolute expression level. Immunohistochemical quantified the protein expression levels of ACTA2. RNA-seq data from 954 LUAD patients, including 541 stage I patients, along with 12 published datasets comprising 1,331 stage I LUAD patients, were used to validate our findings.

Focal adhesion (FA) was identified as the only pathway significantly perturbed at both genomic and transcriptomic levels by comparing 98 AIS/MIA and 99 LUAD. Then, two FA genes (COL11A1 and THBS2) were found strongly upregulated from AIS/MIA to stage I while steadily expressed from normal to AIS/MIA. Furthermore, unsupervised clustering separated stage I patients into two molecularly and prognostically distinct subtypes (S1 and S2) based on COL11A1 and THBS2 expressions (FA2). Subtype S1 resembled AIS/MIA, whereas S2 exhibited more somatic alterations and activated cancer-associated fibroblast. Immunohistochemistry on 73 samples also observed that CAF was more active in S2 compared to S1 and AIS/MIA. The prognostic value of these two genes identified from our knowledge-driven process was confirmed by 541 stage I patients in a prospective dataset, ddPCR and 12 published datasets. Conclusions: We successfully revealed two molecularly and prognostically distinct subtypes of stage I LUAD by applying key molecular alterations from AIS/MIA to invasive LUAD progression. Our model may help reliably identify high-risk stage I patients for more intensive post-surgery treatment.