Blood-based assessment of patients with mismatch repair-deficient tumors enrolled in NCI-MATCH Arm Z1D (nivolumab).
Authors: Karlovich Chris Alan , Silhavy Jennifer , LoCoco Jennifer S. , Das Biswajit , Rahmanian Sorena , Chang Ting-Chia , Peach Amanda , Harrington Robin , Chen Li , Harris Lyndsay , Fu Yao , Patton David R. , Li Jessica , Chen Alice P. , Takebe Naoko , Pawlowski Traci L. , McShane Lisa , Febbo Phillip G. , Doroshow James H. , Williams P. Mickey 
Published: June 2023

2560 Background: NCI-MATCH was the largest precision medicine initiative conducted to date. Arm Z1D was a study of nivolumab in patients (pts) whose tumors were mismatch repair deficient (MMRd) by immunohistochemistry (IHC). Plasma was available for analysis of circulating tumor DNA (ctDNA), a component of cell-free DNA (cfDNA), in 39 of 47 pts enrolled to treatment. Large ctDNA gene panels that interrogate microsatellite instability (MSI), tumor mutational burden (TMB) and other complex biomarkers may provide an important complement to tumor-based assessments. Methods: 10 – 30 ng of cfDNA extracted from Streck tubes was tested with a modified version of the 523-gene TruSightä Oncology 500 ctDNA RUO assay (Illumina, San Diego, CA). Blood MSI (bMSI) status was calculated by interrogating ~2400 microsatellite sites and applying an experimentally validated cut-off. Blood TMB (bTMB) was calculated as total number of nonsynonymous and synonymous SNVs and Indels identified per Mb. Tumor sequencing was performed with the 143-gene Oncomine Comprehensive Assay v2. Overlap of tumor and plasma gene panels was evaluated for concordance. MMRd was evaluated by MLH1 and MSH2 IHC expression. MSI PCR testing was performed on 7 microsatellite sites. Clinical response was assessed by RECIST v1.1 criteria. Results: ORR was 31% (12/39) for 39 MMRd patients with plasma samples. Among these, 34 pts (87%) were classified as MSI-high (MSI-H) by ctDNA at baseline. The 5 MSS cases included 1 CR, 2 PRs and 2 unevaluable for response. The median bTMB was 56.4 mut/Mb (range: 1.65 – 435 mut/Mb). All pts had at least one oncogenic/likely oncogenic alteration identified in plasma. Of 136 of these mutations identified in tumor, 107 (78.6%) were detected in plasma. The most frequently mutated genes in plasma included TP53 (n = 26 pts), RNF43 (n = 16 pts, all G659Vfs*41) and PIK3CA (n = 16 pts). MLH1 (n = 5) and MSH2 (n = 5) mutations were also identified. RNF43 G659Vfs*41, which was not interrogated by the targeted tumor panel, is known to be strongly associated with MSI-H status. In this heavily pre-treated cohort (median prior therapies = 3), variants in genes commonly associated with clonal hematopoiesis of indeterminant potential were identified in all pts. Changes in cfDNA tumor fraction tracked with clinical response in 4 of 4 pts where on-treatment blood was collected for TSO 500 assessment. Conclusions: 78.6% of oncogenic/likely oncogenic alterations found in tumor where also identified in plasma. 87% of MMRd pts by tumor IHC were defined as MSI-H by baseline ctDNA, and median bTMB was 56.4 mut/Mb. These data suggest blood-based MSI and TMB should be further explored as biomarkers for the non-invasive identification of patients who may be candidates for immune checkpoint blockade, particularly when tissue is limiting.