Longitudinal tracking of ALK-positive lung cancer from plasma using circulating-tumor RNA and circulating-tumor DNA in the BRIGHTSTAR clinical trial.
Authors: Heeke Simon , Gandhi Saumil , Tran Hai T. , Lam Vincent K. , Byers Lauren Averett , Gibbons Don Lynn , Gay Carl Michael , Altan Mehmet , Antonoff Mara , Le Xiuning , Tu Janet Chen , Pek Michelle , Poh Jonathan , Ngeow Kao Chin , Tsao Anne S. , Cascone Tina , Negrao Marcelo Vailati , Blumenschein George R. , Heymach John , Elamin Yasir Y 
Published: June 2023

e21072

Background: While the administration of specific tyrosine-kinase inhibitors (TKIs) in ALK-fusion positive lung cancer has led to significant improvement in clinical outcomes, detection of gene fusions remains challenging, especially from liquid biopsies. In tissue biopsies, assays which incorporate RNA-based detection have demonstrated increased sensitivity for gene fusion detection. We therefore hypothesized that a liquid biopsy assay equally including assessment of gene fusions using circulating-tumor RNA (ctRNA) in addition to circulating-tumor DNA (ctDNA), will improve detection. Furthermore, we hypothesized that detection of gene fusions as well as mutations will also correlate with clinical treatment. Methods: We retrospectively analyzed 89 samples from 35 patients included in the BRIGHTSTAR clinical trial assessing local consolidative therapy (LCT) and brigatinib in patients with stage IV or recurrent Non-small Cell Lung Cancer and confirmed ALK rearrangement (NCT03707938). Samples were included at baseline (N = 31), prior to LCT after 8 weeks of brigatinib treatment (N = 29), after LCT ( < 3 weeks; N = 25) and at progression (N = 4). We used a targeted next generation sequencing (NGS) assay assessing both ctDNA as well as ctRNA (LiquidHALLMARK, Lucence Health, Palo Alto) to detect ALK fusions as well as mutations in 80 genes. Up to 5 ml of plasma was analyzed per sample. Results: At baseline, ALK fusions were detected in 15/31 patients (48%) of which 8 were detected using both ctDNA and ctRNA, while four were exclusively detected in ctDNA and three in ctRNA. Plasma ctDNA concentrations for patients with detectable ALK fusions at baseline were significantly higher than for those without detectable gene fusions (mean 26.1 ng/mL versus 16.6 ng/mL; p = 0.0044). ALK fusions were detected in two patients pre-LCT, exclusively in ctRNA, while ALK fusions cleared completely post-LCT. At progression, ALK fusions were detected in 2/4 samples (50%) in both ctDNA and ctRNA. Of the two negative samples tissue biopsies were available confirming the absence of an ALK rearrangement, including one transformation to squamous cell carcinoma. Overall, including analysis of ctRNA led to a 36% relative additional detection of ALK fusions compared to analyzing ctDNA alone. Furthermore, ctDNA mutation positivity in genes other than ALK was 17/31 (55%), 9/29 (31%), 8/25 (32%) and 3/4 (75%) for baseline, pre-LCT, post-LCT and progression, respectively. Conclusions: We highlight that ALK fusions can be reliably detected in plasma of lung cancer patients and detectability of ALK fusions correlate with exposure to treatment. Analyzing ctRNA in addition to ctDNA improves sensitivity of fusion detection and is a highly promising strategy in this setting.