Background: Neoadjuvant chemoimmunotherapy for non-small cell lung cancer (NSCLC) has improved pathological responses and survival rates compared with chemotherapy alone, leading to Food and Drug Administration (FDA) approval of nivolumab plus chemotherapy for resectable stage IB-IIIA NSCLC (AJCC 7th edition) without ALK or EGFR alterations. Unfortunately, a considerable percentage of tumors do not completely respond to therapy, which has been associated with early disease progression. So far, it is impossible to predict these events due to lack of knowledge. In this study, we characterized the gene expression profile of tumor samples to identify new biomarkers and mechanisms behind tumor responses to neoadjuvant chemoimmunotherapy and disease recurrence after surgery. Methods: Tumor bulk RNA sequencing was performed in 16 pretreatment and 36 post-treatment tissue samples from 41 patients with resectable stage IIIA NSCLC treated with neoadjuvant chemoimmunotherapy from NADIM trial. A panel targeting 395 genes related to immunological processes was used. Tumors were classified as complete pathological response (CPR) and non-CPR, based on the total absence of viable tumor cells in tumor bed and lymph nodes tested at surgery. Differential-expressed genes between groups and pathway enrichment analysis were assessed using DESeq2 and gene set enrichment analysis. CIBERSORTx was used to estimate the proportions of immune cell subtypes. Results: CPR tumors had a stronger pre-established immune infiltrate at baseline than non-CPR, characterized by higher levels of IFNG, GZMB, NKG7, and M1 macrophages, all with a significant area under the receiver operating characteristic curve (ROC) >0.9 for CPR prediction. A greater effect of neoadjuvant therapy was also seen in CPR tumors with a reduction of tumor markers and IFNγ signaling after treatment. Additionally, the higher expression of several genes, including AKT1, BST2, OAS3, or CD8B; or higher dendritic cells and neutrophils proportions in post-treatment non-CPR samples, were associated with relapse after surgery. Also, high pretreatment PD-L1 and tumor mutational burden levels influenced the post-treatment immune landscape with the downregulation of proliferation markers and type I interferon signaling molecules in surgery samples. Conclusions: Our results reinforce the differences between CPR and non-CPR responses, describing possible response and relapse immune mechanisms, opening the possibility of therapy personalization of immunotherapy-based regimens in the neoadjuvant setting of NSCLC.

Survival data support the use of first-line osimertinib as the standard of care for epidermal growth factor receptor (EGFR)-positive non-small cell lung cancer (NSCLC). However, it remains unclear whether upfront osimertinib is superior to sequential first- or second-generation tyrosine kinase inhibitors (TKIs) followed by osimertinib for all patients. It is impossible to predict which patients are at high risk of progression, and this constitutes a major limitation of the sequential TKI approach. A total of 830 plasma samples from 228 patients with stage IV, EGFR-positive NSCLC who were treated with first-line TKIs were analysed by digital polymerase chain reaction (dPCR). The circulating tumour DNA (ctDNA) levels helped to identify patients with significantly improved survival rate, regardless of the treatment. Patients treated with first- or second-generation TKIs (N = 189) with EGFR mutations in plasma at a mutant allele frequency (MAF) <7% before treatment initiation (low-risk patients) or who were ctDNA negative after 3 or 6 months of treatment and with an MAF <7% at diagnosis (high responders) had two-thirds lower risk of death than patients in the opposite situation (adjusted hazard ratio [HR] = 0.38; 95% confidence interval [CI]: 0.23-0.64 and HR = 0.22; 95% CI: 0.12-0.42, respectively). The median overall survival (OS) for low-risk patients and high responders treated with first- or second-generation TKIs was 34.2 months and not reached, respectively, regardless of second-line treatment. There were no significant difference in OS between low-risk or high-responder patients treated upfront with osimertinib (N = 39) and those treated under a sequential approach with osimertinib (N = 60). Median OS was not reached in both cases. Pre-treatment ctDNA levels identify low-risk patients, who may benefit from sequential TKI treatment. Information regarding EGFR mutation clearance can help to improve patient selection.

Purpose: Perioperative chemoimmunotherapy is the standard of care for resectable, locally advanced non–small cell lung cancer (NSCLC). Although pathologic complete response (pCR) correlates with excellent survival outcomes, some patients without pCR still exhibit long-term survival. In this study, we evaluate the added value of minimal residual disease (MRD). Experimental Design: MRD was assessed in 60 patients from the NADIM II trial (NCT03838159) using the Guardant Reveal assay. In NADIM II, patients with NSCLC without EGFR or ALK alterations were randomly assigned to receive neoadjuvant nivolumab plus chemotherapy (experimental arm) or chemotherapy alone, followed by surgery. Patients in the experimental arm with R0 resection received adjuvant nivolumab. Results: The MRD detection rate was 9.6%. MRD after surgery or during adjuvant treatment was associated with inferior event-free survival (EFS) and overall survival [OS; hazard ratio (HR): 10.2; 95% confidence interval (CI), 3.7–28.3 and HR: 10.0; 95% CI, 2.0–49.9, respectively]. All patients with MRD-negative plasma samples in at least two time points were alive [HR: not estimable (NE), P < 0.001], with only one relapse (HR: 41.6; 95% CI, 5.0–348.8), corresponding to a patient relapsing with a single brain metastasis. MRD enhanced the prognostic value of pCR for both EFS (P < 0.001) and OS (P = 0.015). Among non-pCR patients, MRD remained a significant prognostic marker (HR: 6.2; 95% CI, 2.2–17.1 and HR: 6.5; 95% CI, 1.3–32.5, for EFS and OS respectively). All non-pCR patients with MRD-negative results in at least two time points were alive (HR: NE, P = 0.025), with one relapse (HR: 19.9; 95% CI, 2.4–165.6), corresponding to the aforementioned case. Conclusions: MRD may refine prognostic evaluation beyond pCR in resectable NSCLC undergoing perioperative chemoimmunotherapy.