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A. Prat
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P1.01 - Advanced NSCLC (ID 757)
- Event: WCLC 2017
- Type: Poster Session with Presenters Present
- Track: Advanced NSCLC
- Presentations: 1
- Moderators:
- Coordinates: 10/16/2017, 09:30 - 16:00, Exhibit Hall (Hall B + C)
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P1.01-075 - Simultaneous Multiplex Profiling of Gene Fusions, METe14 Mutations and Immune Genes in Advanced NSCLC by NCounter Technology (ID 9481)
09:30 - 09:30 | Author(s): A. Prat
- Abstract
Background:
Assessment of several immune-genes and tumor drivers is critical for individualized treatment of non-small cell lung cancer (NSCLC). We have previously demonstrated the ability of the transcript-based nCounter Technology for the detection of ALK, ROS1 and RET gene fusions, using a customized codeset (Reguart et al. Clinical Chemistry 2017). Here, we present the first results of the validation in advanced NSCLC samples of a new CodeSet designed to simultaneously characterize clinically relevant gene fusions, MET alterations and the expression of immune genes.
Method:
We have designed an in-house custom set to detect driver fusions involving 4 genes (ALK, ROS, RET, NTRK), MET exon 14 skipping mutation, MET overexpression and immune genes (PD1, PDL-1, CD4, CD8, FOXP3, GZMM, IFNG). A panel of ALK-ROS-RET-NTRK positive cell lines (H2228, H3122, SU-DHL-1, HCC78, BaF3 pBABE, LC2/ad and a NTRK-positive cell line), Hs746T (METex14), EBC-1 (overexpressing MET) and a negative cell line (PC9) were used for the initial validation of the panel. Subsequently, 58 FFPE samples from advanced NSCLC patients, previously characterized by FISH, RT-PCR and IHC, have been analyzed. Total amount of 100-150 ng RNA was used for detection. Workflow includes RNA isolation, hybridization and digital counting with for a total turnaround of 3 days. Raw counts were normalized using positive controls, negative controls and house-keeping genes.
Result:
.Results obtained with the cell lines positive for ALK, ROS1, RET and NTRK1 fusion genes were exactly coincident with their genotypes, with fusion transcripts successfully detected in all cases by 3’/5’ imbalance and direct fusion probes. In addition, METex14 splicing transcripts were detected in the Hs746T cells at significant levels, higher than those of wt MET mRNA. In contrast, METex14 mRNA counts were almost undetectable in the rest of cell lines. Regarding FFPE samples from advanced patients, 46 could be successfully analyzed by nCounter. Among 13 patients positive for ALK and ROS1 fusions, 12 were confirmed by nCounter. Regarding the METex14 splicing variant, 5 out of 6 patients previously detected by RT-PCR were also positive by nCounter.
Conclusion:
Preliminary data suggest that multiplex detection of clinically relevant drivers can be successfully achieved using nCounter Technology. The assay is simple, requires short hands-on-time, needs low input RNA and is highly efficient in detecting gene rearrangements and METex14 splicing variants. Results will be prospectively validated in a larger cohort of advanced NSCLC patients and we will determine if clusters of different inmune-phenotypes exist among oncogenic-driven NSCLC tumors.
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P2.02 - Biology/Pathology (ID 616)
- Event: WCLC 2017
- Type: Poster Session with Presenters Present
- Track: Biology/Pathology
- Presentations: 1
- Moderators:
- Coordinates: 10/17/2017, 09:30 - 16:00, Exhibit Hall (Hall B + C)
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P2.02-014 - Simultaneous Gene Profiling of Advanced NSCLC: Single-Molecule Quantification of DNA and RNA by nCounter3D™ Technology (ID 9808)
09:30 - 09:30 | Author(s): A. Prat
- Abstract
Background:
Currently, assessment of several tumor drivers is critical for individualized treatment of non-small cell lung cancer (NSCLC). Tools for molecular profiling are based on DNA, RNA and protein (PCR, NGS, FISH, IHC). However, these tests have several disadvantages including hands-on-time and tissue mass requirements. Nanostring digital barcoding technology enables simultaneous assay of different analytes, DNA and RNA from a single sample with 3D Biology Technology.
Method:
The nCounter Vantage 3D SNV:Fusions Lung Assay was used to analyze a total of 36 formalin-fixed paraffin embedded (FFPE) samples from advanced NSCLC patients. Samples were known to harbor mutations (EGFR, KRAS, NRAS, PIK3CA, BRAF, P53) or gene-fusion rearrangements (ALK, ROS1, RET, NTRK1) as verified by sequencing (Ion Torrent, Gene Reader), nCounter Elements, IHC and/or FISH. Probes were designed to target 25 genes for SNVs (104 different point and InDel mutations) as well as four genes for fusion transcripts (ALK, ROS1, RET, NTRK1) including 33 specific variants. The 3D workflow requires pre-amplification of gDNA, whereas RNA does not require any enzymatic treatment. After hybridization, the analytes (DNA/RNA) are pooled for simultaneous, single-lane, digital counting in total turnaround of 3 days. A total amount of 5ng DNA and 150ng RNA from two4-micron FFPE-sections was used for the assay without microdissection.
Result:
A total of 72 analyses (DNA/RNA) were performed with an evaluation pass of 97.2% (70/72 analyses yielded results) and 89% concordant results (64/72). Sensitivity of the technique was 92.1%. Among 41 SNVs interrogated in this study 34 were successfully detected (two not evaluable). Five new mutations were found involving NRAS, FBXW7, GNA11, FGFR2 and KRAS genes. Of those, only two were considered false positives as they were not confirmed by alternative sequencing and/or PCR. The remaining three were not assessable for test confirmation. For gene fusion analysis, 13 known positive samples were tested. All fusion transcripts were detected for ALK (n=5) RET (n=2) and NTRK1 (n=1). For ROS1 (n=5) there were 2 false negatives only detected by nCounter Elements target-specific assay.
Conclusion:
We have shown that the SNV detection chemistry can be successfully combined with fusion gene expression analysis by using the nCounter 3D™ single-workflow. The Nanostring nCounter Vantage 3D SNV:Fusions Lung Assay is highly efficient in detecting hotspot mutations as well as gene rearrangements. The assay is simple, features a brief hands-on time and requires low amounts of genomic material, supporting minimal use of precious samples.
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P3.01 - Advanced NSCLC (ID 621)
- Event: WCLC 2017
- Type: Poster Session with Presenters Present
- Track: Advanced NSCLC
- Presentations: 2
- Moderators:
- Coordinates: 10/18/2017, 09:30 - 16:00, Exhibit Hall (Hall B + C)
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P3.01-045 - Correlation of EGFR Mutation Detection in CtDNA by Two Different Platforms in Advanced NSCLC Patients from a Single Institution (ID 9475)
09:30 - 09:30 | Author(s): A. Prat
- Abstract
Background:
Circulating-free tumor DNA (ctDNA) has emerged as a sensitive and feasible non-invasive blood-based approach alternative to tissue biopsies to screen for genetic drivers in advanced non-small cell lung cancer (NSCLC) patients. Recently, the COBAS Mutation Test has been FDA-approved for the detection of EGFR mutation (EGFRm) in peripheral blood but other approaches are currently used in clinical practice. Here, we aimed to correlate two different platforms for EGFRm monitoring ctDNA in an enriched cohort of tissue-genetically phenotyped EGFRm advanced NSCLC patients.
Method:
Blood samples were prospectively obtained from an enriched cohort of EGFR+ advanced NSCLC patients. Formalin-fixed paraffin-embedded (FFPE) tumor was characterized by NGS (Ion AmpliSeq Lung Cancer Research Panel v.2) in all patients at diagnose. Plasma ctDNA derived from peripheral whole blood was evaluated by COBAS EGFR Mutation Test v2 and a Peptide Nucleic Acid (PNA) probe–based real-time polymerase chain reaction blinded to baseline tumor genotype. Diagnostic accuracy and concordance of both blood techniques was used for direct comparison with respect to the molecular status of FFPE tissue.
Result:
A total of 80 matched pairs of peripheral blood samples from 40 patients were collected. Baseline tissue NGS reported mutations at exon 19 del (n=23), exon 21 (n=10), exon 18 (n=2), exon 20 (n=2) and T790M (n=3). Four wild-type EGFR tumors were used as controls. Blood samples were obtained at diagnose (n=12) and during tyrosine-kinase inhibitor (TKi) treatment for monitoring (n=28). Overall, concordance between both blood-based techniques with respect tissue-NGS was 100% (4/4) for negative controls and 55% (20/36) for positive tissue-NGS samples. Detection based on PNA and COBAS was negative in 60% (24/40) and 32.5% (13/40) patients respectively. Among 19 samples negative by PNA at monitoring, COBAS allowed plasma EGFRm detection in 11 patients. At baseline, the only two negative samples patients by both techniques were found in patients with localized brain disease. Six patients had detectable driver T790M mutation; among three patients with T790M+ in tissue, COBAS allowed detection in plasma in one patient whereas none was identified with PNA. The other three patients had acquired T790M mutations identified only in blood, all by COBAS.
Conclusion:
In this prospective blinded validation cohort, both methods retained high specificity. However, major differences between techniques were observed for longitudinal monitoring of EGFRm in blood.
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P3.01-088i - Phase Ib Trial of Xentuzumab and Abemaciclib in Advanced or Metastatic Solid Tumors, including Advanced NSCLC (ID 8572)
09:30 - 09:30 | Author(s): A. Prat
- Abstract
Background:
There remains an unmet need for additional treatment options in patients with advanced non-small cell lung cancer (NSCLC) refractory to chemotherapy and immunotherapy. The insulin-like growth factor (IGF) axis and cyclin D-cyclin-dependent kinase (CDK) 4/6-retinoblastoma pathway have been implicated in the pathogenesis and resistance mechanisms of a variety of cancers, including NSCLC. Binding of IGF-I and -II to the IGF receptor results in upregulation of cyclin D1, and subsequent progression through the cell cycle, thus providing rationale for the simultaneous inhibition of IGF-I and -II and CDK4/6. This trial assesses the maximum-tolerated dose (MTD)/recommended phase II dose (RP2D), safety and preliminary efficacy of the IGF-ligand-neutralizing antibody, xentuzumab, in combination with abemaciclib, a selective, small-molecule inhibitor of both CDK4 and 6, in patients with solid tumors. One of two expansion cohorts (Cohort E) will further characterize the safety, tolerability, pharmacokinetics (PK) and preliminary efficacy of the combination in patients with NSCLC.
Method:
Study BI 1280.18 (NCT03099174) is a phase Ib multicenter, non-randomized, open-label, dose escalation trial with four dose-finding cohorts (Cohorts A–D) followed by two expansion cohorts (Cohorts E, F). For the NSCLC cohort (Cohort E), eligible patients include adults ≥18 years (≥20 for Japan), with measurable or evaluable disease, adequate organ function, ECOG PS≤1, and CDK4/6 inhibitor-naïve stage IV NSCLC after 1–2 lines of therapy and failure after platinum-based chemotherapy and immunotherapy. Patients with NSCLC (Cohort E) will receive the combination at the RP2D determined in patients with solid tumors (Cohort A) who will receive xentuzumab (starting dose 1,000 mg weekly i.v.) plus abemaciclib (starting dose 150 mg every 12 hours). The primary endpoint of Cohort E is the objective response (OR) in patients with pre-treated advanced NSCLC; disease control, duration of disease control, time to OR, duration of OR, and progression-free survival (PFS) are secondary endpoints. This study will be conducted in the US, Europe and Japan. Patient screening started in May 2017. Target enrolment is ~88 patients, including ~20 patients with stage IV NSCLC refractory to platinum-based chemotherapy and immunotherapy.
Result:
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Conclusion:
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