Epidermal Growth Factor Receptor T790M Research Articles

Discovery of a Potent and Mutant-Selective EGFR Inhibitor that Overcomes T790M-Mediated Resistance in Lung Cancer.

Despite remarkable activity in epidermal growth factor receptor (EGFR)-mutant lung cancer patients, the clinical efficacy of EGFR tyrosine kinase inhibitors (TKIs) is limited by the emergence of acquired resistance, which is mostly caused by a secondary T790M mutation. Fortunately, newly developed, mutant-selective EGFR-TKIs against T790M have been proven as an effective therapeutic approach although only osimertinib has received the FDA approval until now. To determine the in vitro and in vivo efficacy of a new EGFR TKI, OBX1-012 in cells with mutant EGFR. Effects of OBX1-012 on cellular viability and EGFR-related signaling were determined in EGFR-mutant non-small cell lung cancer (NSCLC) cells, including cells harboring T790M mutations. In addition, in vivo efficacy of OBX1-012 was evaluated in xenograft models. We report the discovery and preclinical assessment of another novel, mutant-selective EGFR-TKI, OBX1-012. Compared with other mutant-selective EGFR-TKIs such as olumitinib and osimertinib, it showed similar potency and selectivity for mutant EGFR. OBX1-012 treatment was highly effective against human EGFR-mutant lung cancer models with or without EGFR T790M, not only in vitro but also in vivo. However, OBX1-012 like other EGFR-TKIs failed to exhibit efficacy for the exon 20 insertion mutation or C797S mutation, which was generated by site-directed mutagenesis and stable transfection of Ba/F3 cells. These results identify OBX1-012 as a highly effective, mutant-selective EGFR-TKI for the treatment of T790M-mediated resistance in NSCLC.

Xenograft tumors derived from malignant pleural effusion of the patients with non-small-cell lung cancer as models to explore drug resistance

BackgroundNon-small cell lung cancer (NSCLC) patients with epidermal growth factor receptor (EGFR) mutations or anaplastic lymphoma kinase (ALK) fusions show dramatic responses to specific tyrosine kinase inhibitors (TKIs); however, after 10–12 months, secondary mutations arise that confer resistance. We generated a murine xenograft model using patient-derived NSCLC cells isolated from the pleural fluid of two patients with NSCLC to investigate the mechanisms of resistance against the ALK- and EGFR-targeted TKIs crizotinib and osimertinib, respectively.MethodsGenotypes of patient biopsies and xenograft tumors were determined by whole exome sequencing (WES), and patients and xenograft-bearing mice received targeted treatment (crizotinib or osimertinib) accordingly. Xenograft mice were also treated for prolonged periods to identify whether the development of drug resistance and/or treatment responses were associated with tumor size. Finally, the pathology of patients biopsies and xenograft tumors were compared histologically.ResultsThe histological characteristics and chemotherapy responses of xenograft tumors were similar to the actual patients. WES showed that the genotypes of the xenograft and patient tumors were similar (an echinoderm microtubule-associated protein-like 4-ALK (EML4–ALK) gene fusion (patient/xenograft: CTC15035EML4–ALK) and EGFR L858R and T790M mutations (patient/xenograft: CTC15063EGFR L858R, T790M)). After continuous crizotinib or osimertinib treatment, WES data suggested that acquired ALK E1210K mutation conferred crizotinib resistance in the CTC15035EML4–ALK xenograft, while decreased frequencies of EGFR L858R and T790M mutations plus the appearance of v-RAF murine sarcoma viral oncogene homolog B (BRAF) G7V mutations and phosphatidylinositol-4-phosphate 3-kinase catalytic subunit type 2 alpha (PIK3C2A) A86fs frame shift mutations led to osimertinib resistance in the CTC15063EGFR L858R, T790M xenografts.ConclusionsWe successfully developed a new method of generating drug resistance xenograft models from liquid biopsies using microfluidic technology, which might be a useful tool to investigate the mechanisms of drug resistance in NSCLC.

Phase Ib Study of High-dose Intermittent Afatinib in Patients With Advanced Solid Tumors

BackgroundThe present phase Ib study assessed the maximum tolerated dose (MTD), safety, pharmacokinetics, and antitumor activity of high-dose intermittent (HDI) afatinib monotherapy for patients with advanced solid tumors. The planned focus was patients with epidermal growth factor receptor (EGFR) T790M+ non–small-cell lung cancer (NSCLC). Materials and MethodsEligible patients had histologically confirmed advanced solid tumors that were unsuitable for, or unresponsive to, standard therapy. The study used a 3+3 design with a starting dose of 90 mg/d for 3 days every 14 days (28-day cycles) and incremental dose escalations to 200 mg/d. ResultsThirty-five patients (18 with NSCLC) were treated (6 at 90 mg; 3 at 120 mg; 9 at 150 mg; 11 at 160 mg; and 6 at 200 mg). One patient in the 90-mg cohort (grade 3 rash) and 2 patients in the 200-mg cohort (grade 3 diarrhea; grade 3 mucositis) experienced a dose-limiting toxicity. The MTD was 160 mg. The most common treatment-related adverse events were diarrhea (total, 88.6%; grade 3, 14.3%), rash/acne (total, 62.9%; grade 3, 2.9%), and fatigue (total, 40.0; grade 3, 0%). The maximum afatinib plasma concentration at the MTD was 313 ng/mL, exceeding the in vitro IC50 (inhibitor concentration decreased by one half) range for T790M inhibition. The trough levels suggested no systematic change in afatinib plasma concentrations during long-term treatment at this dosing schedule. Of the 13 T790M+ NSCLC patients, 1 achieved an objective response (7.7%). ConclusionHDI afatinib was feasible and tolerable and could potentially be further explored for NSCLC indications, including patients with central nervous system disease, rare EGFR mutations, or T790M+ NSCLC intolerant of third-generation EGFR tyrosine kinase inhibitors.

Acquired EGFR T790M Mutation After Relapse Following EGFR-TKI Therapy: A Population-based Multi-institutional Study.

To describe the prevalence and determinants of acquired epidermal growth factor receptor (EGFR) T790M gene mutation in a clinical practice setting. We performed a retrospective chart review study between January 2013 and November 2017 across multiple institutes, covering a population of 3 million people. We reviewed the charts of 233 patients non-small cell lung cancer with EGFR mutations. Of them, 99 (42.5%) patients had acquired T790M mutations in EGFR. Patients ≥75 years old and patients with an exon 19 deletion had higher rates of acquired T790M mutation than did younger patients and those with an exon 21 L858R mutation. In 75 patients treated with afatinib, 34 (45.3%) patients had acquired T790M mutation. The sensitivity of T790M mutation detection was lower in plasma specimens than in biopsy specimens. This population-based study confirms previous studies and highlights potential determinants of acquired T790M mutation to be considered in clinical practice.

Diagnostic accuracy of droplet digital PCR for detection of EGFR T790M mutation in circulating tumor DNA.

ObjectivesAlthough different methods have been established to detect epidermal growth factor receptor (EGFR) T790M mutation in circulating tumor DNA (ctDNA), a wide range of diagnostic accuracy values were reported in previous studies. The aim of this meta-analysis was to provide pooled diagnostic accuracy measures for droplet digital PCR (ddPCR) in the diagnosis of EGFR T790M mutation based on ctDNA.Materials and methodsA systematic review and meta-analysis were carried out based on resources from Pubmed, Web of Science, Embase and Cochrane Library up to October 11, 2017. Data were extracted to assess the pooled sensitivity, specificity, positive likelihood ratio, negative likelihood ratio (NLR), diagnostic OR (DOR), and areas under the summary receiver-operating characteristic curve (SROC).ResultsEleven of 311 studies identified have met the including criteria. The sensitivity and specificity of ddPCR for the detection of T790M mutation in ctDNA ranged from 0.0% to 100.0% and 63.2% to 100.0%, respectively. For the pooled analysis, ddPCR had a performance of 70.1% (95% CI, 62.7%–76.7%) sensitivity, 86.9 % (95% CI, 80.6%–91.7%) specificity, 3.67 (95% CI, 2.33–5.79) PLR, 0.41 (95% CI, 0.32–0.55) NLR, and 10.83 (95% CI, 5.86–20.03) DOR, with the area under the SROC curve being 0.82.ConclusionThe ddPCR harbored a good performance for detection of EGFR T790M mutation in ctDNA.

Molecular Testing for Stage IV Non-Small Cell Lung Cancer Patients With Targetable Mutations Following Disease Progression.

Molecular Testing for Stage IV Non-Small Cell Lung Cancer Patients With Targetable Mutations Following Disease Progression.

Evidence-Based Best Practices for EGFR T790M Testing in Lung Cancer in Canada

Epidermal growth factor receptor (egfr) tyrosine kinase inhibitors (tkis) are recommended as first-line systemic therapy for patients with non-small-cell lung cancer (nsclc) having mutations in the EGFR gene. Resistance to tkis eventually occurs in all nsclc patients treated with such drugs. In patients with resistance to tkis caused by the EGFR T790M mutation, the third-generation tki osimertinib is now the standard of care. For optimal patient management, accurate EGFR T790M testing is required. A multidisciplinary working group of pathologists, laboratory medicine specialists, medical oncologists, a respirologist, and a thoracic radiologist from across Canada was convened to discuss best practices for EGFR T790M mutation testing in Canada. The group made recommendations in the areas of the testing algorithm and the pre-analytic, analytic, and post-analytic aspects of clinical testing for both tissue testing and liquid biopsy circulating tumour dna testing. The recommendations aim to improve EGFR T790M testing in Canada and to thereby improve patient care.

Unique genetic profiles from cerebrospinal fluid cell-free DNA in leptomeningeal metastases of EGFR-mutant non-small-cell lung cancer: a new medium of liquid biopsy

BackgroundLeptomeningeal metastases (LM) are more frequent in non-small-cell lung cancer (NSCLC) with epidermal growth factor receptor (EGFR) mutations. Due to limited access to leptomeningeal lesions, the purpose of this study was to explore the potential role of cerebrospinal fluid (CSF) as a source of liquid biopsy in patients with LM. Patients and methodsPrimary tumor, CSF, and plasma in NSCLC with LM were tested by next-generation sequencing. In total, 45 patients with suspected LM underwent lumbar puncture, and those with EGFR mutations diagnosed with LM were enrolled. ResultsA total of 28 patients were enrolled in this cohort; CSF and plasma were available in 26 patients, respectively. Driver genes were detected in 100% (26/26), 84.6% (22/26), and 73.1% (19/26) of samples comprising CSF cell-free DNA (cfDNA), CSF precipitates, and plasma, respectively; 92.3% (24/26) of patients had much higher allele fractions in CSF cfDNA than the other two media. Unique genetic profiles were captured in CSF cfDNA compared with those in plasma and primary tissue. Multiple copy number variations (CNVs) were mainly identified in CSF cfDNA, and MET copy number gain identified in 47.8% (11/23) of patients was the most frequent one, while other CNVs included ERBB2, KRAS, ALK, and MYC. Moreover, loss of heterozygosity (LOH) of TP53 was identified in 73.1% (19/26) CSF cfDNA, which was much higher than that in plasma (2/26, 7.7%; P<0.001). There was a trend towards a higher frequency of concomitant resistance mutations in patients with TP53 LOH than those without (70.6% versus 33.3%; P=0.162). EGFR T790M was identified in CSF cfDNA of 30.4% (7/23) of patients who experienced TKI progression. ConclusionCSF cfDNA could reveal the unique genetic profiles of LM and should be considered as the most representative liquid biopsy medium for LM in EGFR-mutant NSCLC.

142PD Detection of EGFR mutations in cerebrospinal fluid of EGFR-mutant lung adenocarcinoma with brain metastases

Background: A large proportion of lung adenocarcinoma (LADC) with epidermal growth factor receptor (EGFR) activating mutation will develop brain metastases (BMs) throughout the course of their disease. BM lesions were hard to obtain, which resulted in poor understanding of resistance mechanisms of EGFR tyrosine kinase inhibitors (EGFR-TKIs) therapy in these patients. Liquid biopsy of cerebrospinal fluid (CSF) may provide potential information of intracranial lesions. Herein, we aimed to investigate the alternative detection of EGFR mutation status in CSF. Methods: Between July 2014 and June 2017 in Beijing Chest Hospital, thirty LADC patients with BMs carrying activating EGFR mutation in their primary tumors determined by amplification-refractory mutation system (ARMS) assay were enrolled. Their matched CSF and plasma samples were collected. Droplet digital PCR assays (ddPCR) for EGFR mutations, including 19del, L858R, and T790M were developed using two millilitres CSF or plasma. The clinical response of intracranial objective response rate (iRR), intracranial progression-free survival (iPFS), and intracranial overall survival (iOS) from the time of diagnosis of BM were evaluated. Results: All of the included patients were Chinese and had been diagnosed with stage IV lung adenocarcinoma. Out of 30 patients, 21 were females and 9 males aged from 34 to 75 years old (median age of 58 years old). The majority (24/30) classified as good Eastern Cooperative Oncology Goup performance status (ECOG PS <2). In all of cases, CSF cytology were negative and examinations of cranial imaging showed no leptomeningeal metastases (LM). In ddPCR assays, EGFR mutations were detected in CSF of 10 patients (33.3%), including three cases with EGFR T790M mutations, and in plasma of 14 patients (46.7%), including six cases with EGFR T790M mutations. All EGFR T790M mutations were found during or after EGFR-TKIs treatments. Five patients with activating EGFR mutations in CSF achieved intracranial partial response (iPR) after treated with combination of first-generation EGFR-TKIs and whole brain radiotherapy (WBRT)/stereotactic radiosurgery (SRS). Three patients with EGFR T790M mutations in CSF achieved iPR after treated with second-line osimertinib. The median iOS and iPFS from the time of diagnosis of BM were 15.0 months and 11.0 months, respectively. The following Table shows the EGFR testing result in detail.Table(Abstract 142PD) EGFR testing resultPatientPrimary Tissue EGFR Mutation (ARMS)CSF EGFR MutationPlasma EGFR MutationStatusFrequencyStatusFrequency119delWTT790M0.3%219delWT19del3L858RL858R37.9%L858R2.0%4L858RWTWT519delWTWT6L858RWTL858R/T790M10.8%/26.5%7L858RWTWT819del19del69.7%19del/T790M11.0%/7.0%9L858RWTWT1019delWTWT1119delWTWT12L858RWTL858R/T790M2.4%/0.2%13L858RL858R32.8%WT1419delWTWT1519del19del/T790M13.2%/0.5%19del/T790M14.4%/3.5%16L858RWTWT17L858RL858R/T790M16.2%/2.0%L858R/T790M8.1%/2.2%18L858RWTWT1919del19del43.3%19del14.9%2019delWTWT2119delWT19del3.7%2219del19del21.8%19del7.4%23L858RWTL858R20.5%2419delWT19del11.4%2519delWTWT2619delWT19del0.8%27L858RL858R7.2%L858R5.2%2819del19del6.9%WT2919del19del/T790M35.7%/12.1%WT3019delWT19del15.3% Open table in a new tab Conclusions: It was feasible to test EGFR mutation in CSF and plasma. CSF may serve as liquid biopsy of advanced LADC with BMs by detecting cfDNA within CSF to characterize EGFR mutations. For those advanced LADC with BMs harboring EGFR mutation, dynamic monitoring EGFR mutation status of CSF would be an appropriate choice. Legal entity responsible for the study: Beijing Chest Hospital, Capital Medical University, Beijing Tuberculosis and Thoracic Tumor Research Institute, Beijing 101149, China. Funding: Beijing Municipal Administration of Hospitals Youth Talent Training Project (Grant No. QML20151502) Disclosure: All authors have declared no conflicts of interest.

The alteration of T790M between 19 del and L858R in NSCLC in the course of EGFR-TKIs therapy: a literature-based pooled analysis.

Treatment-naive epidermal growth factor receptor (EGFR) T790M mutation is more inclined to coexist with L858R than with 19 del in non-small cell lung cancer (NSCLC) patients. However, EGFR-tyrosine kinase inhibitors (EGFR-TKIs) might alter this status. We sought to compare the prevalence of T790M upon acquired resistance to EGFR-TKIs between 19 del and L858R by assembling all existing data. Electronic databases were comprehensively searched for eligible studies. The primary endpoint was the odds ratio (OR) of T790M mutation in NSCLC co-existing with L858R mutation and 19 del upon resistance to first-generation EGFR-TKIs. A random effects model was used. Stratified analysis was performed based on study type (retrospective and prospective), race (Asians and Caucasians) and sample type (tissue and plasma). A total of 25 studies involving 1,770 patients were included. The overall T790M existent rate was 45.25%. Post-resistance T790M was more frequent in 19 del than in L858R mutated patients (53% vs. 36%; OR 1.87; P<0.001). All outcomes of subgroup and overall analyses were similar. In contrast, we re-analyzed the previous meta-analysis, finding that the pooled rate of pretreatment T790M was 14% and 22% in 19 del and L858R respectively (OR 0.59; P<0.001). The increase of T790M rate was 2.79-fold in 19 del and only 0.63-fold in L858R in the course of EGFR-TKIs therapy. Opposite to the situation of de novo T790M, it was observed that T790M was more frequent in exon 19 deletion than in L858R among patients with acquired resistance to EGFR-TKIs. The difference in T790M alteration between 19 del and L858R encourages development of detection or treatment strategies for the specific resistance mechanism.

148P Method comparison for detection of epidermal growth factor receptor (EGFR) T790M mutation in matched biopsied tumour sections and plasma samples

148P Method comparison for detection of epidermal growth factor receptor (EGFR) T790M mutation in matched biopsied tumour sections and plasma samples

In-silico evidences for binding of Glucokinase activators to EGFR C797S to overcome EGFR resistance obstacle with mutant-selective allosteric inhibition

The tyrosine kinase inhibitors (TKI) against epidermal growth factor receptor (EGFR) are generally utilized as a part of patients with non-small cell lung carcinoma (NSCLC). However, EGFR T790M mutation results in resistance to most clinically available EGFR TKIs. Third-generation EGFR TKIs against the T790M mutation has been in active clinical development to triumph the resistance problem; they covalently bind with conserved Cys797 inside the EGFR active site, offering both potency and kinase-selectivity. Third generation drugs target C797, which makes the C797S resistance mutation more subtle. EGFR C797S mutation was accounted to be a main mechanism of resistance to the third-generation inhibitors. The C797S mutation gives off an impression of being an ideal target for conquering the acquired resistance to the third generation inhibitors. We have performed structure based-virtual screening strategies for binding of glucokinase activator to EGFR C797S, which can overcome EGFR resistance impediment with mutant-selective allosteric inhibition towards all kinds of mutant EGFR (T790M, L858R, TMLR) and WT EGFR. The final filter of Lipinski’s Rule of Five, Jargan’s Rule of Three and in silico ADME predictions gave 23 hits, which conform to Lipinski’s rule and Jorgensen’s rule and all their pharmacokinetic parameters are inside the appropriate range characterized for human use, in this manner demonstrating their potential as a drug-like molecule.

Selective gene amplification to detect the T790M mutation in plasma from patients with advanced non-small cell lung cancer (NSCLC) who have developed epidermal growth factor receptor tyrosine kinase inhibitor (EGFR-TKI) resistance.

The epidermal growth factor receptor (EGFR) T790M mutation is associated with resistance to EGFR tyrosine kinase inhibitors (EGFR-TKIs) in non-small cell lung cancer (NSCLC). However, tissues for the genotyping of the EGFR T790M mutation can be difficult to obtain in a clinical setting. The aims of this study were to evaluate a blood-based, non-invasive approach to detecting the EGFR T790M mutation in advanced NSCLC patients using the PointMan™ EGFR DNA enrichment kit, which is a novel method for the selective amplification of specific genotype sequences. Blood samples were collected from NSCLC patients who had activating EGFR mutations and who were resistant to EGFR-TKI treatment. Using cell-free DNA (cfDNA) from plasma, EGFR T790M mutations were amplified using the PointMan™ enrichment kit, and all the reaction products were confirmed using direct sequencing. The concentrations of plasma DNA were then determined using quantitative real-time PCR. Nineteen patients were enrolled, and 12 patients (63.2%) were found to contain EGFR T790M mutations in their cfDNA, as detected by the kit. T790M mutations were detected in tumor tissues in 12 cases, and 11 of these cases (91.7%) also exhibited the T790M mutation in cfDNA samples. The concentrations of cfDNA were similar between patients with the T790M mutation and those without the mutation. The PointMan™ kit provides a useful method for determining the EGFR T790M mutation status in cfDNA.

Standardizing biomarker testing for Canadian patients with advanced lung cancer.

The development and approval of both targeted and immune therapies for patients with advanced non-small cell lung cancer (nsclc) has significantly improved patient survival rates and quality of life. Biomarker testing for patients newly diagnosed with nsclc, as well as for patients progressing after treatment with epidermal growth factor receptor (EGFR) inhibitors, is the standard of care in Canada and many parts of the world. A group of thoracic oncology experts in the field of thoracic oncology met to describe the standard for biomarker testing for lung cancer in the Canadian context, focusing on evidence-based recommendations for standard-of-care testing for EGFR, anaplastic lymphoma kinase (ALK), ROS1, BRAF V600 and programmed death-ligand (PD-L1) at the time of diagnosis of advanced disease and EGFR T790M upon progression. As well, additional exploratory molecules and targets are likely to impact future patient care, including MET exon 14 skipping mutations and whole gene amplification, RET translocations, HER2 (ERBB2) mutations, NTRK, RAS (KRAS and NRAS), as well as TP53. The standard of care must include the incorporation of testing for novel biomarkers as they become available, as it will be difficult for national guidelines to keep pace with technological advances in this area. Canadian patients with nsclc should be treated equally; the minimum standard of care is defined in this paper.

MET or NRAS amplification is an acquired resistance mechanism to the third-generation EGFR inhibitor naquotinib

As a third-generation epidermal growth factor receptor (EGFR) tyrosine kinase inhibitor (TKI), osimeritnib is the standard treatment for patients with non-small cell lung cancer harboring the EGFR T790M mutation; however, acquired resistance inevitably develops. Therefore, a next-generation treatment strategy is warranted in the osimertinib era. We investigated the mechanism of resistance to a novel EGFR-TKI, naquotinib, with the goal of developing a novel treatment strategy. We established multiple naquotinib-resistant cell lines or osimertinib-resistant cells, two of which were derived from EGFR-TKI-naïve cells; the others were derived from gefitinib- or afatinib-resistant cells harboring EGFR T790M. We comprehensively analyzed the RNA kinome sequence, but no universal gene alterations were detected in naquotinib-resistant cells. Neuroblastoma RAS viral oncogene homolog (NRAS) amplification was detected in naquotinib-resistant cells derived from gefitinib-resistant cells. The combination therapy of MEK inhibitors and naquotinib exhibited a highly beneficial effect in resistant cells with NRAS amplification, but the combination of MEK inhibitors and osimertinib had limited effects on naquotinib-resistant cells. Moreover, the combination of MEK inhibitors and naquotinib inhibited the growth of osimertinib-resistant cells, while the combination of MEK inhibitors and osimertinib had little effect on osimertinib-resistant cells. Clinical assessment of this novel combination (MEK inhibitors and naquotinib) is worth considering in osimertinib-resistant lung tumors.

AZD9291 Increases Sensitivity to Radiation in PC-9-IR Cells by Delaying DNA Damage Repair after Irradiation and Inducing Apoptosis.

AZD9291 is a novel, irreversible epidermal growth factor receptor (EGFR) tyrosine kinase inhibitor (TKI), which is administered orally. It has been proven effective in non-small cell lung cancer (NSCLC) patients, with both EGFR-sensitizing and EGFR T790M mutations in preclinical models. However, the potential therapeutic effects of AZD9291 combined with other modalities, including ionizing radiation, are not well understood. The presence of AZD9291 significantly increases the cell-killing effects of radiation in PC-9-IR cells with a secondary EGFR mutation (T790M), which was developed from NSCLC PC-9 cells (human lung adenocarcinoma cell with EGFR 19 exon 15 bp deletion) after chronic exposure to increasing doses of gefitinib, and in H1975 cells (human lung adenocarcinoma cell with EGFR exon 20 T790M mutation de novo), but not in PC-9 cells or in H460 cells (human lung adenocarcinoma cell with wild-type EGFR). In PC-9-IR cells, AZD9291 remarkably decreases phosphorylation levels of EGFR, extracellular regulated protein kinase (ERK), and protein kinase B (AKT). AZD9291 increases sensitivity to radiation in PC-9-IR cells by delaying deoxyribonucleic acid (DNA) damage repair after irradiation and inducing apoptosis, and enhances tumor growth inhibition when combined with radiation in PC-9-IR xenografts. Our findings suggest a potential therapeutic effect of AZD9291 as a radiation sensitizer in lung cancer cells with an acquired EGFR T790M mutation, providing a rationale for a clinical trial using the combination of AZD9291 and radiation in NSCLCs harboring acquired T790M mutation.

Abstract LB-B03: Exosome-based Detection of EGFR T790M in Plasma from Non-Small Cell Lung Cancer Patients

Abstract About 60% of non-small cell lung cancer (NSCLC) patients develop resistance to targeted epidermal growth factor receptor (EGFR) inhibitor therapy through the EGFR T790M mutation. Although these patients respond well to third generation tyrosine kinase inhibitors like osimertinib, obtaining tissue biopsies for mutation profiling poses risks and is often not feasible. Detecting the mutation in a liquid biopsy from plasma has emerged as a viable alternative to biopsy, however existing liquid biopsy tests using cell-free DNA (cfDNA) often suffer from low sensitivity due to the low copy number of mutant molecules in circulation. As an example, the Food and Drug Adminstration (FDA) approved companion diagnostic for T790M mutation status in plasma (cobas® EGFR mutation Test version 2) only has a 58% sensitivity at a specificity of 80%. In this study we aimed to achieve a higher performance by combining the mutated alleles on cfDNA as well as exosomal RNA and DNA (exoNA) . ExoNA and cfDNA were extracted from the plasma of NSCLC patients with biopsy-confirmed T790M-positive (N = 102) or T790M-negative (N = 108) samples. The T790M mutation status in plasma exoNA and cfDNA was determined using an analytically validated allele-specific qPCR assay in a CLIA laboratory. The training cohort consisted of 105 samples (51 positives, 54 negatives). The validation cohort consisted of 105 samples (51 positives, 54 negatives), where ~40% of the patients had intrathoracic disease(M0/M1a). Detection of the T790M mutation in exoNA and cfDNA achieved 92% sensitivity and 89% specificity when compared to tumor biopsy results in the validation cohort. We also observed a very high sensitivity (88%) in patients with intrathoracic disease (M0/M1a), patients for whom detection by liquid biopsy has been proven particularly challenging. The combination of exoNA and cfDNA for EGFR T790M detection has a higher sensitivity and specificity compared to historical cohorts using ctDNA alone. This is especially important as it could aid giving the right treatment to patients where biopsy is not available or avoid unnecessary tumor biopsies for T790M mutation testing. Citation Format: Elena Castellanos, Dominik G. Grimm, Vasisht Tadigotla, Stefan Bentink, James Hurley, John Healy, Patricia L. Neal, Mia Sher, Dan Baughman, Chris Karlovich, Mitch Raponi, Anne Krug, Mikkel Noerholm, Luis E. Raez, Johan Skog. Exosome-based Detection of EGFR T790M in Plasma from Non-Small Cell Lung Cancer Patients [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2017 Oct 26-30; Philadelphia, PA. Philadelphia (PA): AACR; Mol Cancer Ther 2018;17(1 Suppl):Abstract nr LB-B03.

Olmutinib Induced Lichen Planus Like Eruption

Drug induced lichen planus like eruption is an uncommon cutaneous adverse effect of several drugs. This appears symmetric eruption of erythematous or violaceous plaques resembling lichen planus on the trunk and extremities. A 50-year-old male presented with scaly, violaceous plaques and dusky brown macules on whole body. For four months, the patient was treated with olmutinib, an oral, third-generation epidermal growth factor receptor-tyrosine kinase inhibitor. In May 2016, olmutinib received its first global approval in South Korea for the treatment of patients with locally advanced or metastatic epidermal growth factor receptor T790M mutation-positive non-small cell lung cancer. The biopsy specimen from the patient showed features of lichen planus. We diagnosed him with olmutinib-induced lichen planus like eruption. He was treated with oral methylprednisolone and topical desoxymethasone 0.25% ointment. At the same time, olmutinib dose was decreased to three-fourths of this patient's starting dose. After that, the cutaneous lesions improved.

Abstract B02: Liquid biopsy for screening of driver mutations in EGFR gene: Personalized medicine for use tyrosine kinase inhibitors in non-small cell lung cancer patients

Abstract Non-small cell lung cancer (NSCLC) accounts for approximately 85% of all lung cancers worldwide. In Brazil there were an estimated 28.220 new cases in 2016. The 5-year survival rate in advanced NSCLC patients is less than 5%, making necessary the development of diagnostic strategies to improve the treatment and survival of these patients. Activating mutations of epidermal growth factor receptor (EGFR) gene occur in approximately 10–15 % in Caucasian and approximately 30% in Brazilian NSCLC cases. The first- and second-generation of EGFR tyrosine kinase inhibitors (TKIs) have been widely used for advanced NSCLC patients who are eligible for the treatment. Liquid biopsies using plasma-derived cell-free DNA (cfDNA) is an emerging noninvasive test allowing a better selection and monitoring of NSCLC. The aim of this study was to evaluate the frequency of EGFR mutations using liquid biopsy samples before and after TKIs treatment in a cohort of NSCLC Brazilian patients. All the patients were investigated for EGFR mutations by real-time qPCR Cobas© EGFR Mutation Test v2. Clinical data were obtained prior to blood collection from physician and/or pathologists involved in the original diagnoses. Blood samples were collected at Progenetica Laboratory or sent from other laboratories, centrifuged immediately for the plasma separation, and frozen. Fisher´s exact test was calculated to compare frequency distribution between two groups (GraphPad Prism version 6.00). This study enrolled 535 NSCLC Brazilian patients from Southeast+South (74.9%), North+Northeast (8.8%), and Midwest+Federal District (4.7%). Regional information was missing for 11.3% of cases. As expected, NSCLC was more frequent among women (56.4%) compared to men (43.6%). Four hundred cases (74.8%) did not show any EGFR mutations; however, 135 cases presented 19Del (57.8%), L858R (16.3%), double 19Del; T790M (11.1%), double T790M; L858R (8.15%), G719X (2.9%), L861Q (1.48%), 20Ins, triple L858R; T790M; 20Ins and S768I; G719X (0.74% each one) mutations spanning 18, 19, 20, and 21 EGFR exons. Previous results from liquid biopsy for EGFR mutation were obtained for 75/535 cases (14%). Among these patients, 10/75 cases (13.3%) presented nondetected mutations. Deletions at exon 19 (19Del) and mutations at exon 21 (L858R) were the most frequent, accounting for 61.5% and 23%, respectively. Since 19Del and L858R are drug-sensitive mutations, we compared the frequency of EGFR mutations in patients before and after TKIs treatment. Interestingly, patients harboring drug-sensitive mutations before the treatment were negative for the presence of 19Del (37.5%) and L858R (66.7%) after the treatment, respectively (p&amp;lt;0.0001). In addition, EGFR T790M mutation already associated with drug resistance was identified in 3/65 (4.6%) of cases in double with 19Del before the TKIs treatment. After treatment, one patient was negative and two patients remained positive for both mutations, respectively. These results showed that liquid biopsy based on cfDNA is a powerful tool for personalized medicine for early detection, stratification, and monitoring of EGFR druggable mutations in NSCLC patients. The presence of cfDNA has been related to aggressiveness and poor prognosis of NSCLC patients, and it is important to note that not all cases have the tumor cfDNA. To our knowledge this is the first report showing the application of liquid biopsies for management of NSCLC patients in Brazil. Clinical data collection is under way for better reliability results. High index of nondetected results was observed in this study; however, new protocols using more sensitive methodologies such as next-generation sequencing and digital PCR are under development in our institution to improve the sensitivity of this test. Citation Format: Priscila D. R. Cirillo, Natália P. Lopes, Carolina Bustamante, Agatha A. Morais, Natália L. Viana, Camila S. Portella, Elvis C. C. Mateo, Maíra C. M. Freire, Mariano G. Zalis. Liquid biopsy for screening of driver mutations in EGFR gene: Personalized medicine for use tyrosine kinase inhibitors in non-small cell lung cancer patients [abstract]. In: Proceedings of the AACR International Conference held in cooperation with the Latin American Cooperative Oncology Group (LACOG) on Translational Cancer Medicine; May 4-6, 2017; São Paulo, Brazil. Philadelphia (PA): AACR; Clin Cancer Res 2018;24(1_Suppl):Abstract nr B02.

Intercalated Treatment Following Rebiopsy Is Associated with a Shorter Progression-Free Survival of Osimertinib Treatment.

PurposeEpidermal growth factor receptor (EGFR) T790M mutation serves as an important predictor of osimertinib efficacy. However, little is known about how it works among patients with various timings of T790M emergence and treatment.Materials and MethodsAdvanced EGFR-mutant lung adenocarcinoma patients with positive T790M mutation in tumor were retrospectively enrolled and observed to determine the outcomes of osimertinib treatment. We evaluated the association between patients’ characteristics and the efficacy of osimertinib treatment, particularly with respect to the timing of T790M emergence and osimertinib prescription.ResultsA total of 91 patients were enrolled, including 14 (15.4%) with primary and 77 (84.6%) with acquired T790M mutation. The objective response rate and disease controlratewere 60.9% and 85.1%, respectively. The median progression-free survival (PFS) and overall survival were 11.5 months (95% confidence interval [CI], 9.0 to 14.0) and 30.4 months (95% CI, 11.3 to 49.5), respectively. There was no significant difference in response rate and PFS between primary and acquired T790M populations. In the acquired T790M subgroup, patientswho received osimertinib after T790M had been confirmed by rebiopsy had a longer PFS than those with intercalated treatments between rebiopsy and osimertinib prescription (14.0 months [95% CI, 9.0 to 18.9] vs. 7.2 months [95% CI, 3.7 to 10.8]; adjusted hazard ratio, 0.48 [95% CI, 0.24 to 0.98; p=0.043]). Rebiopsy timing did not influence the outcome.ConclusionOsimertinib prescription with intercalated treatment following rebiopsy but not the timing of T790M emergence influenced the treatment outcome. We suggest that it is better to start osimertinib treatment once T790M mutation has been confirmed by biopsy.