Digital gene expression analysis of NSCLC-patients reveals strong immune pressure, resulting in an immune escape under immunotherapy

Abstract

Background Immune checkpoint inhibitors (ICIs) are currently one of the most promising therapy options in the field of oncology. Although the first pivotal ICI trial results were published in 2011, few biomarkers exist to predict their therapy outcome. PD-L1 expression and tumor mutational burden (TMB) were proven to be sometimes-unreliable biomarkers. We have previously suggested the analysis of processing escapes, a qualitative measurement of epitope structure alterations under immune system pressure, to provide predictive information on ICI response. Here, we sought to further validate this approach and characterize interactions with different forms of immune pressure. Methods We identified a cohort consisting of 48 patients with advanced non-small cell lung cancer (NSCLC) treated with nivolumab as ICI monotherapy. Tumor samples were subjected to targeted amplicon-based sequencing using a panel of 22 cancer-associated genes covering 98 mutational hotspots. Altered antigen processing was predicted by NetChop, and MHC binding verified by NetMHC. The NanoString nCounter® platform was utilized to provide gene expression data of 770 immune-related genes. Patient data from 408 patients with NSCLC were retrieved from The Cancer Genome Atlas (TCGA) as a validation cohort. Results The two immune escape mechanisms of PD-L1 expression (TPS score) (n = 18) and presence of altered antigen processing (n = 10) are mutually non-exclusive and can occur in the same patient (n = 6). Both mechanisms have exclusive influence on different genes and pathways, according to differential gene expression analysis and gene set enrichment analysis, respectively. Interestingly, gene expression patterns associated with altered processing were enriched in T cell and NK cell immune activity. Though both mechanisms influence different genes, they are similarly linked to increased immune activity. Conclusion Pressure from the immune system will lay the foundations for escape mechanisms, leading to acquisition of resistance under therapy. Both PD-L1 expression and altered antigen processing are induced similarly by pronounced immunoactivity but in different context. The present data help to deepen our understanding of the underlying mechanisms behind those immune escapes.

Learning From the Past: Optimizing Future Use of Immune Checkpoint Inhibitors in Advanced Non-Small Cell Lung Cancer

“…Notably, EGFR-mutation frequency varies across different ethnicities and are more frequent in never smokers. In the past six years, immune-checkpoint inhibitors (ICIs) have changed the treatment paradigm for patients with advanced NSCLC without specific druggable molecular alterations, owing to improved response rates and survival compared to CTx in randomized controlled trials (RCTs) (Table 2.2) [46][47][48][49][50] . Table 2.2 Clinical outcomes in the pivotal RCTs with nivolumab or pembrolizumab monotherapy [46][47][48][49][50] .…”

“…In the past six years, immune-checkpoint inhibitors (ICIs) have changed the treatment paradigm for patients with advanced NSCLC without specific druggable molecular alterations, owing to improved response rates and survival compared to CTx in randomized controlled trials (RCTs) (Table 2.2) [46][47][48][49][50] . Table 2.2 Clinical outcomes in the pivotal RCTs with nivolumab or pembrolizumab monotherapy [46][47][48][49][50] . The ICI-revolution in NSCLC was commenced by the approval of the antiprogrammed cell death-1 (PD-1) antibody, nivolumab in previously treated, squamous NSCLC patients in September 2015 in Denmark, following the results from the Checkmate 017 trial 38,47 .…”

“…Patient and treatment characteristics in the pivotal RCTs with nivolumab or pembrolizumab monotherapy [46][47][48][49][50] .…”

“…Most NSCLC gene expression studies rely upon surgically resected early-stage tumors with large tissue resections, and thus may not be comparable to gene expressions in ICI-treated patients with advanced stage disease [19,50]. Only few GEP studies have included patients with advanced NSCLC treated with ICI in routine clinical care.…”

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All patients PD-L1 ≥50% Checkmate 017 47 Nivolumab 9.2 (7.3 -13.3) 42 (34 -50) 3.5 (2.1 -4.9) 20 (14 -28) Docetaxel 6.0 (5.1 -7.3) 24 (17 -31) 2.8 (2.1 -3.5) 9 (5 -15) Checkmate 057 46 Nivolumab 12.2 (9.7 -15.0) 51 (45 -56) 2.3 (2.2 -3.3) 19 (15 -24) Docetaxel 9.4 (8.1 -10.7) 39 (33 -45) 4.2 (3.5 -4.9) 12 (9 -17) Keynote 010 48 Pembro 2 mg/kg/3w 10.4 (9.4 -11.9) 43 (NR) 3.9 (3.1 -4.1) 18 (NR) 30 (NR) Pembro 10 mg/kg/3w 12.7 (10.0 -17.3) 52 (NR) 4.0 (2.7 -4.3) 18 (NR) 29 (NR) Docetaxel 8.5 (7.5 -9.8) 35 (NR) 4.0 (3.1 -4.2) 9 (NR) 8 (NR)Keynote 024 49,50 Pembro 200 mg/3w 26.

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