Applicability of routine targeted next generation sequencing to estimate tumour mutational burden (TMB) in patients treated with immune-checkpoint inhibitor therapy


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Heather Urus1,David Pinato2,Thomas Newsom- Davis3,Persephone Du Parcq4,Katherine Belessiotis2,Leah Mapara2,Nandita Gupta5,Danielle Power2,Justin Weir5,Ching N Wong1,Ragu P Ratnakumaran2,Jamshid Khorashad4,Kathy Dominy4,Mark Bower3
1Department of Surgery & Cancer, Imperial College London,2Department of Oncology, Imperial College NHS Trust,3Department of Oncology, Chelsea & Westminster Hospital,4Molecular Pathology Laboratory, Hammersmith Hospital,5Department of Histopathology, Imperial College NHS Trust

Abstract

Background

Tumour Mutational Burden (TMB) is an emerging biomarker of response to Immune- Checkpoint inhibitors (ICPI). Routine Targeted Next Generation Sequencing (tNGS) was used to test for actionable mutations from solid tumours to establish the precision of estimates of patients’ TMB. The ability of tNGS platforms to reflect the overall mutational status of the corresponding tumour has never been prospectively validated.

Method

We sequenced 79 archival samples of immune checkpoint inhibitor (ICPI) recipients (57% lung cancer, 43% melanoma) using an Ion Ampliseq Cancer Hotspot Panel. Samples were obtained from 4 cohorts: Primary Lung Adenocarcinoma (47%), Primary Lung Squamous Cell Carcinoma (2%), Primary Lung Carcinoma of other histotypes (8%) and Malignant melanoma (43%). On ICPI commencement, the majority of samples (84%) had metastatic disease, with loco- regional lymph node spread (16%) in the remaining samples. Liver (8%), Lung (46%), Bone (23%), Central Nervous System (26%) and Adrenal gland (15%) were the main sites of metastasis. Prior to ICPI treatment, the majority of patients received chemotherapy (42%). 34% received surgery, 14% had received radical radiotherapy, 24% had received palliative radiotherapy and 9% were given targeted therapy.

Anti- PD(L)-1 monotherapy was received by 93% of patients (71 patients) with 7% of patients (5) receiving Anti- PD-1/ CTLA-4 combination therapy.

Results

Using multiple cut-off values, we verified that TMB by tNGS did not correlate with response or survival following ICPI. We found enrichment of ATM mutations in ICPI-refractory tumours (p=0.01) to correlate with worse survival (4.2 months versus 10 months, p=0.03).

Conclusion

Limited-coverage tNGS delivers an imprecise estimate of patients’ TMB but may help identify somatic variants of predictive/prognostic significance. This highlights adequate coverage as a key factor affecting the reliability of TMB testing. Increasing the coverage of tNGS platforms will enable the development of TMB quantification as a predictive marker of response to ICPI.