Abstract 5871
Background
Tumor mutation burden (TMB) predicts durable benefit from immune checkpoint blockade in several cancer types. We demonstrate the ability of a targeted panel with fast turn-around time and low input needs to estimate TMB from research samples.
Methods
We developed a single sample analysis workflow to estimate mutation burden (TMB; nonsynonymous mutations/Mb) from FFPE and fresh frozen tumor research samples. The assay utilizes a PCR-based targeted panel that covers 409 genes and 1.7 Mb of genomic regions. The workflow requires only 10 ng of input DNA and enables a 2.5-day turn-around time from sample to the final report. Sequencing is performed on high throughput semiconductor sequencing platform to achieve sufficient depth (∼500x coverage) and accuracy. The workflow is tumor sample only, with no matched normal sample required; germ-line variants, along with background noise, are removed through filters based on population databases. The assay is research use only, not for diagnostic procedures.
Results
A comparison with whole exome sequencing (WES) on 12 FFPE tumors, where WES was performed on tumors and their matched normal using Agilent’s exome enrichment kit (∼150x coverage for tumor; ∼100x coverage for normal) on illumina platform and our assay ran on tumors only, showed high correlation (r2=0.83) between TMB estimates by our assay with that from WES. To assess reproducibility, we compared raw somatic mutations/Mb in library replicates for a cohort of 21 FFPE research samples (19 CRC, 2 Melanoma) and observed high correlation (r2=0.97). Our pipeline identifies mutation signatures consistent with specific mechanisms such as UV and tobacco damage, as well as substitutions from FFPE processing.
Conclusions
A simple workflow has been developed on the Ion Torrent sequencing platform with an AmpliSeq panel to estimate TMB from FFPE and fresh frozen tumor research samples. This solution will advance research in immuno-oncology.
Clinical trial identification
Legal entity responsible for the study
Thermo Fisher Scientific.
Funding
Thermo Fisher Scientific.
Editorial Acknowledgement
Disclosure
R. Chaudhary, D. Cyanam, V. Mittal, W. Tom, J. Au-Young, C. Allen, S. Sadis, F. Hyland: Employment: Thermo Fisher Scientific.
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