National Real-life Study of Tumor Mutational Burden Assessment in First-line Lung Cancer

1. Current knowledge about the field under investigation

Clinical evidence demonstrates that treatment with immune checkpoint blocker (ICB) agents benefit to patients across multiple tumor type. However, development of predictive biomarkers is needed to identify patients who are most likely to respond to immunotherapy.

An emerging biomarker for response to immunotherapy is the total number of mutations present in a tumor specimen. This biomarker is named mutation load or tumor mutational burden (TMB). It is hypothesized that highly mutated tumors are more likely to harbor neoantigens targeted by activated immune cells. This metric, allowed by recent advances in next-generation sequencing (NGS) technologies, notably whole-exome sequencing (WES) and RNA-sequencing (RNA-seq), has been shown, in several tumor types, to correlate with patient response to ICB. Indeed, the TMB has been correlated with clinical benefits of anti-PD-1 and anti-CTLA-4 therapy in various tumor types, including malignant melanoma (Snyder et al., 2014; Van Allen et al., 2015) with a threshold of more than 100 nonsynonymous single-nucleotide variants (nsSNV) per exome, non-small cell lung cancers (NSCLC) (Rizvi et al., 2015) with a threshold defined as superior to 178 nsSNVs per exome, and several DNA repair-deficient tumors (Howitt et al., 2015; Le et al., 2015, 2017). A recent study prospectively confirmed that the PFS among patients with a high tumor mutational burden was significantly longer with nivolumab plus ipilimumab than with chemotherapy in NSCLC (Hellmann et al., 2018). Overall, a direct link between DNA repair deficiency, mutational landscape, predicted neoantigen load, and clinical activity of ICB is suggested.

TMB was defined as the number of somatic, coding, base substitution, and indel mutations per megabase of genome examined. All base substitutions and indels in the coding region of targeted genes. It has been shown that TMB calculated using cancer gene panel (CGP) assay agrees well with whole exome measures of mutation burden (Chalmers et al., 2017). This indicates that CGP, targeting the entire coding region of several hundred genes, covers sufficient genomic size to accurately assess WES mutational burden. It was found that filtering out germline alterations and rare variants was important to obtaining accurate measurements of TMB, and this will especially be important in patients from ethnic backgrounds not well represented in sequencing datasets. These findings indicate that CGP is an accurate, cost-effective, and clinically available tool for measuring TMB. The results of down sampling analysis show that the variation in measurement due to sampling when sequencing 1.1 Mb is acceptably low, resulting in highly accurate calling of TMB at a range of TMB levels. This sampling variation increases as the number of megabases sequenced decreases, especially at lower levels of TMB. While targeted CGP can be used to accurately assess TMB, it is not currently suited for identification of neoantigens, which might occur in any gene. Nevertheless, the theragnostic impact of TMB has been also determined with a targeted CGP by the FoundationOne CDx assay (Hellmann et al., 2018).

The failure to assess the TMB may occur at different steps in the analytical process. It can be at the sample acquisition with a low level of DNA (5-10%). The NGS processing depending on the method can lead to a process failure around 2-3%. And finally, the bioinformatic curation will lead to a 3-5% of failure. At the end, the TMB will be undetermined with an attrition rate at 15% of the samples (Fondation Medicine Personal communication).