Background

Patients across a range of disease types have demonstrated robust and durable responses using checkpoint inhibitor therapies (CPITs). Given the limitations of immuno-histochemical based testing, identifying a unified, quantitative metric to determine potential response to CPITs remains an urgent need. Tumor mutational burden (TMB) measures the number of somatic protein coding mutations per target sequence in a tumor specimen. This measure has been associated with response and survival for multiple CPITs across an array of indications. In this study we describe Foundation Medicine’s (FMI) work to develop and validate a TMB result as part of our comprehensive genomic profiling assays and summarize clinical feasibility in NSCLC, melanoma and bladder cancer.

Methods

We developed an analysis method to determine TMB based on data from our comprehensive genomic profiling assays. TMB is calculated by counting all synonymous and non-synonymous somatic variants across 315 or 405 genes, excluding germline alterations and known or likely driver alterations. The mutation count is normalized by the coding target territory to achieve a mutation density of mutations per megabase (mut/Mb). To determine accuracy, we compared TMB values from our comprehensive genomic profiling assay against a CLIA-validated whole-exome sequencing (WES) method on 29 patients. Precision was assessed over 10 clinical samples replicated 4-6 times. Lower limit of sample tumor purity was determined through dilutions of tumor/normal pairs from 80% to 5% tumor. Clinical feasibility was assessed by analyzing TMB versus immunotherapy-based survival in a cohort of 65 metastatic melanoma patients, 150 urothelial carcinoma patients and 463 NSCLC patients. Additionally, we examined the relationship between TMB and microsatellite instability status (MSI), an independent biomarker associated with response to CPITs.

Results

Foundation Medicine’s TMB measure provides accurate and precise results across a range of tumor mutational burden values on samples with as little as 20% tumor purity. Using cohort specific thresholds, TMB was significantly associated with improved survival to CPITs in NSCLC, melanoma and bladder cancer. Using data from over 40,000 patient samples, we also show significant overlap between high TMB and high MSI samples and show that MSI-High specimens represent a subset of TMB-High specimens.

Conclusions

We have developed and validated the tumor mutational burden (TMB) biomarker as part of our comprehensive cancer genomic profiling assays. Initial clinical feasibility results demonstrate that TMB can be used to predict the likely response to anti-PD-1/PD-L1 CPITs across a growing number of indications including NSCLC, melanoma and bladder cancer.

Citation Format: Daniel S. Lieber, Mark R. Kennedy, Douglas B. Johnson, Jonathan E. Rosenberg, Marcin Kowanetz, Joel R. Greenbowe, Garrett M. Frampton, Caitlin F. Connelly, Alexa B. Schrock, Jeffrey S. Ross, Philip J. Stephens, Siraj M. Ali, Vincent A. Miller, David A. Fabrizio. Validation and clinical feasibility of a comprehensive genomic profiling assay to identify likely immunotherapy responders through tumor mutational burden (TMB) [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 2987. doi:10.1158/1538-7445.AM2017-2987