Characterizing Response to Neoadjuvant Anti-PD-1 Therapy in Non-Small Cell Lung Carcinoma

Abstract

Purpose: Approximately 40% of non-small cell lung carcinoma (NSCLC) patients are resistant to anti-PD-1 treatment. While neoadjuvant application has improved efficacy, current predictive biomarkers for therapeutic response demonstrate limited efficacy in early-stage disease. Developing predictive biomarkers for neoadjuvant anti-PD-1 treatment and gaining a better understanding of mechanisms of resistance is crucial to ensure patient quality of life. Hypothesis and Aims: We hypothesize that a high degree of PD-L1+ immune cell infiltration within pre-treatment specimens will be associated with response to neoadjuvant anti-PD-1, while low immune infiltration and separate immunosuppressive mechanisms in post-treatment specimens will associate with non-response. Aim 1: Assess pre-treatment tumour microenvironment (TME) for evidence of pre-existing PD-1 and PD-L1 expression and observe relationship with response. Aim 2: Determine effect of therapy on TME through comparison of paired pre- and post-treatment specimens. Aim 3: Characterize features of response and resistance to treatment through examination of post-treatment TME. Methodology: Pre- and post-treatment samples from 34 NSCLC patients were stained with a six-marker multiplex immunofluorescence assay to characterize cell subsets impacted by the PD-1/PD-L1 axis. Cell density was calculated following digital image analysis classifying each cell as positive or negative for markers within the multiplex immunofluorescence panel. Response was designated as ≥90% tumour regression following treatment and surgical resection. Single-cell RNA sequencing was performed on select subsets to explore cell function. Results: Intratumoural density of CD8+ T cells within pre-treatment specimens was significantly associated with anti-PD-1 response, with the CD8+FoxP3+ population showing the strongest predictive capacity (p=0.01). These CD8+FoxP3+ cells demonstrate distinct transcriptional profiles from regulatory T cells, with their immediate immune milieu serving as hallmarks of an effective anti-tumour immune response. Patterns among immune cell density over neoadjuvant anti-PD-1 treatment were inconclusive. Significance: CD8+FoxP3+ T cells present a potential predictive biomarker for neoadjuvant anti-PD-1 therapy. Further study will elucidate the exact function of these cells within the tumour microenvironment, as well as identify mechanisms of resistance that could be targeted to improve treatment efficacy.