Abstract 2750: Detecting resistance mechanisms in patients on EGFR TKI treatment by liquid biopsy
Authors: Laure Sorber, Karen Zwaenepoel, An Wouters, Janssens Annelies, Birgitta Hiddinga, Jan Van Meerbeeck, Filip Lardon, Christian Rolfo, Patrick Pauwels
Journal: Cancer Research (2017)

Abstract The rise of liquid biopsy, including the molecular analysis of circulating cell-free tumor DNA (ctDNA), has enabled real-time follow up of cancer patients who receive targeted therapy. Here, we report the case of a lung cancer patient (adenocarcinoma), initially harboring the activating EGFR L858R mutation. Blood and tissue samples were collected during treatment with the EGFR tyrosine kinase inhibitor (TKI) gefitinib, and during the appearance of resistance. Analysis of the tissue biopsies consisted of EGFR mutational analysis by high-resolution melting analysis (HRMA) (all tissue samples), immunohistochemistry (IHC) of cMET (sample 2); FISH analysis of the cMET gene (sample 1 and 3); and next-generation sequencing (NGS) (sample 3). We performed digital droplet PCR (ddPCR) to detect EGFR mutations and cMET amplification in both tissue and plasma samples. To the best of our knowledge, this is the first time that ddPCR is used to screen for cMET amplification in ctDNA. The HRMA analysis of the biopsy during progression under EGFR TKI treatment revealed the EGFR T790M resistance mutation. Progression was also seen by radiological examination and this resulted in discontinuation of gefitinib treatment. As a third generation EGFR TKI was not available by that time, the patient was retreated with carboplatin plus pemetrexed, which resulted in a mixed response. In the second biopsy, only the EGFR activating L858R mutation was detected by HRMA. IHC analysis revealed a strong membrane positivity for cMET in 100% of the tumor cells. After progression upon rechallenging the patient with erlotinib, treatment with osimertinib (third generation EGFR TKI) was initiated. This resulted in only a partial response, with some strongly progressing lesions. NGS analysis of the third tissue biopsy indicated the presence of both the EGFR L858R and T790M mutations, while ddPCR analysis of the plasma sample only detected the EGFR L858R mutation. Due to the fast progression, it was opted to also screen for cMET amplification. DdPCR cMET analysis of the tissue and plasma resulted in a copy number variation of 10.0 and 3.7, respectively. FISH analysis confirmed the cMET amplification with a ratio of 3.1 and an average of 8.4 cMET-signals per nucleus. The results of the ddPCR EGFR and cMET analysis of both the plasma and tissue samples are now being processed to determine the course of the mutational load and the best course of treatment. As such, real-time follow-up of cancer patients is crucial to detect the underlying mechanisms of resistance to targeted therapies. Correlation of molecular analysis with the disease course is key. Our case study highlights the potential of ctDNA analysis for detection of both the EGFR T790M resistance mutation and cMET amplification. Hence, tissue biopsies can be supplemented by liquid biopsies in the screening for resistance mechanisms. Citation Format: Laure Sorber, Karen Zwaenepoel, An Wouters, Janssens Annelies, Birgitta Hiddinga, Jan Van Meerbeeck, Filip Lardon, Christian Rolfo, Patrick Pauwels. Detecting resistance mechanisms in patients on EGFR TKI treatment by liquid biopsy [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 2750. doi:10.1158/1538-7445.AM2017-2750