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V. Ludovini
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MA04 - HER2, P53, KRAS and Other Targets in Advanced NSCLC (ID 380)
- Event: WCLC 2016
- Type: Mini Oral Session
- Track: Advanced NSCLC
- Presentations: 1
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MA04.06 - Signaling Networks in KRAS-Mutant Advanced NSCLC: A Complex Landscape Involving Immunoresponse, Inflammation and DNA Repair (ID 5768)
16:00 - 17:30 | Author(s): V. Ludovini
- Abstract
- Presentation
Background:
KRAS is the most frequently mutated oncogene in Non-Small Cell Lung Cancer (NSCLC) and its role as prognostic and predictive biomarker remains widely debated. Unfortunately, KRAS direct targeting strategies have been unsuccessful and no approved target therapy exists for KRAS-mutant-NSCLC. This pilot study evaluated the activated signaling architecture of advanced NSCLC harbouring a KRAS mutation to better characterize the signaling network driving this subgroup of pulmonary malignancies.
Methods:
Twenty Stage IV Formalin-fixed, paraffin-embedded (FFPE) NSCLCs were collected from chemo-naïve patients at S. Maria della Misericordia Hospital (Perugia, Italy). Ten tumors were KRAS-wild-type (KRAS-WT) and ten were KRAS-Mutant (KRAS-MUT). Whole-tissue lysates were obtained for all samples. Signaling network analysis was performed using the Reverse Phase Protein Array (RPPA) platform to quantitatively evaluate the expression/activation of 148 key proteins and phosphoproteins involved in cellular growth, survival, proliferation, apoptosis, autophagy, inflammation, invasion and cell motility. Wilcoxon Rank-Sum Test was used to compare the signaling architecture of KRAS-MUT and KRAS-WT tumours. All p-values <0.05 were considered significant. Non-parametric correlation analysis was performed to explore the signaling interconnection within each group of patients. Only correlations with p<0.0001 were considered significant.
Results:
This preliminary analysis revealed a statistically significant different activation level of 20 proteins between the KRAS-MUT and KRAS-WT samples. Five of the proteins that were statistically different in the KRAS-MUT group are involved in the inflammatory immunoresponse (ASK1 S83 p<0.01, Axl Y702 p=0.01, Stat2 Y690 p<0.01, Tyk2 Y1054/Y1055 p=0.01 and Twist p<0.01) and six in cell cycle control and DNA repair (ATM S1981 p=0.01; Bcl-xL p=0.03; Cleaved Caspase 3 D175 p=0.02; Histone H3 S10 p<0.01; p53 S15 p<0.01; p27 T187 p=0.04). The analytes that were statistically significant were all lower in the KRAS-MUT group compared to the WT (except for p27 T187 which decreased in the KRAS-MUT group compared to KRAS-WT). Pair-wise correlation analysis of the signaling proteins showed an overall more complex protein-protein interaction network and pathway activation (included AKT/mTOR signaling pathway) in the KRAS-MUT population with high number of statistically significant correlations compared to the KRAS-WT group.
Conclusion:
This pilot study indicated that the effect of KRAS mutation status on protein signaling in NSCLC was an alteration of the immunoresponse axis and DNA repair network. If validated in a larger cohort of patients, these results could have important clinical implications for stratification KRAS-MUT advanced NSCLC patients towards more efficacious targeted treatment and to identify new therapeutic targets based on multi-targets/multi-pathways KRAS inhibitory approach. (AIRC-supported study).
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P3.02b - Poster Session with Presenters Present (ID 494)
- Event: WCLC 2016
- Type: Poster Presenters Present
- Track: Advanced NSCLC
- Presentations: 3
- Moderators:
- Coordinates: 12/07/2016, 14:30 - 15:45, Hall B (Poster Area)
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P3.02b-006 - Role of TP53 Mutations in Determining Primary Resistance to First-Line Tyrosine Kinase Inhibitors in EGFR-Mutated NSCLC Patients (ID 3861)
14:30 - 15:45 | Author(s): V. Ludovini
- Abstract
Background:
Patients with non-small-cell lung cancer (NSCLC) carrying specific mutations at epidermal growth factor receptor (EGFR) gene are usually sensitive to treatments with tyrosine kinase inhibitors (TKIs). However, not all EGFR-mutated patients respond equally to TKI treatments, and approximately 20-30% show primary resistance. Although the mechanisms responsible for acquired resistance are known, those responsible for primary resistance are not completely understood. In this study we aimed to assess the role of TP53 mutations in a cohort of advanced EGFR-mutated NSCLC patients receiving first-line TKIs. We analyzed TP53 gene status in relation to outcome in terms of overall response rate, disease control rate (DCR), response duration, progression free survival (PFS) and overall survival (OS).
Methods:
We retrospectively analyzed 136 patients with advanced EGFR-mutated NSCLC treated with first-line TKIs from January 2012 to April 2015. Exons 5-8 of TP53 gene were amplified by PCR and sequenced by direct sequencing on 123 patients. DCR was defined as the sum of complete response, partial response and stable disease. The survival endpoints examined were PFS and OS. PFS was defined as the time from start of first-line treatment to disease progression or death, whichever occurred first. OS was defined as the time from start of first-line treatment to death.
Results:
TP53 mutations were observed in 37 (30.1%) patients:10 (27.0%), 6 (16.2%), 9 (24.3%) and 12 (32.4%) in exons 5, 6, 7 and 8, respectively. DCR was 70% in TP53-mutated patients compared to 88% in TP53-wt patients (relative risk, RR: 3.17 [95% CI 1.21-8.48], p=0.019). In particular, a 42% DCR was observed in patients with TP53 exon 8 mutation compared to 87% in exon 8 wt patients (RR 9.6 [2.71-36.63], p<0.001). Shorter median PFS and OS were observed in patients with TP53 exon 8 mutations compared to other patients (4.2 months vs 12.5 months [p=0.058] and 16.2 months vs 32.3 months [p=0.114], respectively); these differences became significant in the subgroup of patients with EGFR exon 19 deletion (4.2 months vs 16.8 months [p<0.001] and 7.6 months vs not reached [p=0.006], respectively), hazard ratio (HR) 6.99 (95% CI 2.34-20.87, p<0.001) and HR 4.75 (95% CI 1.38-16.29, p=0.013), respectively.
Conclusion:
TP53 mutations, in particular exon 8 mutations and those defined as nondisruptive, reduce responsiveness to TKI treatment and induce a worse prognosis in EGFR-mutated NSCLC patients, especially in those carrying exon 19 deletions.
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P3.02b-008 - Quantification and Monitoring of Treatment Response in EGFR Mutant NSCLC Patients by Digital-PCR in Plasma cftDNA (ID 5351)
14:30 - 15:45 | Author(s): V. Ludovini
- Abstract
Background:
The identification of activating epidermal growth factor receptor (EGFR) mutations is essential for deciding therapy of non-small cell lung cancer (NSCLC) patients. Circulating cell-free tumor DNA (cftDNA) holds promise as a non-invasive methodology for tumor monitoring in solid malignancies. Among advanced NSCLC patients with an acquired resistance to EGFR-tyrosine kinase inhibitors (TKIs), about 50% carry T790M mutation, but its frequency in EGFR-TKI-naıve patients and dynamic change during therapy remains unclear. We hypothesized that EGFRmutation analysis detection in cftDNA for NSCLC may be feasible for monitoring treatment response to EGFR-TKIs and also predict drug resistance.
Methods:
EGFR sensitive mutations and T790M were analyzed using digital PCR (d-PCR) (Quant studio 3D, life technologies) in longitudinally (at baseline, at 4, 8, 20, 60, 120, 180, 270, 360 days) collected plasma samples (n=50) from 8 tissue-confirmed EGFR-mutant NSCLC patients treated with an EGFR-TKI (Gefitinib N = 4; Erlotinib N = 1; Afatinib N = 3). DNA extracted from plasma of 8 healty blood donors were used to detect the specificity of EGFR mutant assay. Tumor assessment was performed according to RECIST criteria 1.1 every two months.
Results:
The sensitivity of d-PCR in plasma versus tissue was 71.4%. No EGFR mutation was present in the 8 control cases (specificity of 100%). Of four patients who developed progression disease (PD), in the samples of progression, T790M was detected in 75% of cases. The frequency of T790M in pre-TKI plasma samples was of 37.5%. EGFR sensitive mutations decreased at PD while T790M mutation increased in 75% of patients. Patients with concomitant pre-TKI EGFR 19 deletion and T790M showed a PD before of 12 months compared to those with L858R. T790M was frequently detected when new lesions were developed. Four patients had T790M level decreased to undetectable level with longer PFS than those with detectable T790M in blood.
Conclusion:
Our results indicated that d-PCR was a highly sensitive and useful method for detecting the T790M mutation. Moreover, dynamically monitoring T790M change might help determining EGFR-TKI resistance. We thank Italian Association for Cancer Research (AIRC) for supporting the study.
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P3.02b-035 - Cell Free Tumor DNA to Monitor Response to Tyrosine Kinase Inhibitors in Patients with EGFR-Mutant Non-Small Cell Lung Cancer (ID 4038)
14:30 - 15:45 | Author(s): V. Ludovini
- Abstract
Background:
Treatment with epidermal growth factor receptor (EGFR) tyrosine kinase inhibitors (TKIs) has improved outcome of EGFR mutant non-small-cell lung cancer (mEGFR-NSCLC) patients. Monitoring the presence of EGFR sensitizing and resistance (such as T790M) mutations in response to treatment may have a clinical impact on the therapeutic strategy. Detecting these alterations in circulating free tumor DNA (cftDNA) can be an easier and more safe way to obtain information about the EGFR mutational status.
Methods:
Analyses have been conducted in NSCLC patients with a tissue-confirmed EGFR mutation, treated in first-line setting with TKIs. EGFR-sensitive and EGFR exon 20 mutations were analyzed in cftDNA extracted from plasma collected at baseline, after 8 and 20 days’ treatment, and every 4 months of therapy until progression. EGFR analyses were performed using PANAmutyper kit (PANAGENE).
Results:
Of the 16 mEGFR-NSCLC patients treated with first-line TKIs to date (4 with gefitinib, 2 with erlotinib and 10 with afatinib), 9 (56%) showed EGFR-sensitivity mutation at baseline in cftDNA: 6 had an exon 19 deletion, 1 an exon 21 L861Q mutation and 2 an exon 21 L858R mutation synchronous to exon 20 mutations (one insertion and one T790M point mutation). In these 2 last patients, exon 20 mutations were not identified in tumor tissue. The baseline mutation became undetectable in cftDNA in all the 6 patients with EGFR exon 19 deletion at different time point from the beginning of TKIs: in 5 patients after 21 days and in 1 after 8 days. All these patients had partial response at the first radiological evaluation. The subject harboring EGFR L858R mutation synchronous to exon 20 insertion was responsive to TKI and showed the disappearance of exon 20 insertion in cftDNA a the first clinical evaluation, whereas EGFR L858R disappeared after 4 cycles of treatment. Patient with EGFR L858R and T790M didn’t respond to TKI and progressed after 2 months of treatment. At present 3 out of 9 patients progressed but only one showed appearance of T790M in cftDNA during TKI. In the 7 mEGFR-NSCLC with undetectable cftDNA mutation at baseline no changes were seen during treatment.
Conclusion:
EGFR mutation analysis in cftDNA could give important information concerning the activity of TKIs. In particular the disappearance of mutation in cftDNA may be an early parameter of response that has to be validated in prospective trials. Moreover, cftDNA may give integrative information with respect to that obtained from tissue analysis, bypassing the problem of tumor heterogeneity.
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P3.02c - Poster Session with Presenters Present (ID 472)
- Event: WCLC 2016
- Type: Poster Presenters Present
- Track: Advanced NSCLC
- Presentations: 1
- Moderators:
- Coordinates: 12/07/2016, 14:30 - 15:45, Hall B (Poster Area)
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P3.02c-068 - Immunotherapy against Non Small Cell Lung Cancer (NSCLC): Looking for Predictive Factors to Avoid an Untargeted Shooting (ID 5207)
14:30 - 15:45 | Author(s): V. Ludovini
- Abstract
Background:
The use of immunotherapy for the whole Non Small Cell Lung Cancer (NSCLC) population, is like an untargeted shooting. So trying to discover predicitve factors to response still represents the key to the problem. We retrospectively analyzed a cohort of patients (pts) treated with Nivolumab, in the attempt to correlate clinical and molecular features with response.
Methods:
69 heavily pretreated advanced NSCLCs (16 squamous/ 53 adenocarcinomas) were retrospectively evaluated for response to Nivolumab. Pts’ samples from a subgroup of responders (14/17 pts, 82%), were further analyzed for PD-L1/PD-1 expression by immunoistochemistry (IHC), and for TILs density. We used rabbit monoclonal antibodies anti PD-L1 [clone E1L3N] for tumor cell expression (0-3, negative-intense) and mouse monoclonal antibody anti PD-1 [clone EH33] for TILs.
Results:
Clinico-pathologic characteristics: mostly smoker males (81%), PS 0-1 (85%), EGFR+ 7%, K-RAS+ 23%. Overall response rate was 25% (2% complete response and 23% partial response), stable disease 30%, progressive disease 41%. Median progression free survival (PFS) and overall survival (OS) for the entire cohort were 2.9 and 8.3 months (mo) respectively. 1 and 2-y OS rates were both 44% (95% CI, 29-58). Pts with EGFR + NSCLC showed a significantly lower median OS with respect to the wild type cohort (4.5 vs NR; p < 0.005) as well as pts with brain metastases (4.1 vs NR), while a trend toward improvement in PFS for K-RAS+ was seen. A subgroup analysis according to the time to progression to prior chemotherapy regimen (< 3 mo versus > 6 mo), confirmed a poorer survival for those with rapid spread of disease. Among laboratory tests, a better outcome for those who developed G2 leucopenia was demostrated (OS 8.3 vs 5.0 mo). Severe drug-related adverse events occurred in only 5.7% of pts. PD-L1, PD-1, TIL expression for 14/17 pts with OR, were as follows: PD-L1 > 5% 6/14 pts (43%); PD-1 2/14 (14%); focal TILs presence 7/14 (50%).
Conclusion:
Nivolumab confirms activity in NSCLC with durable responses and accettable safety profile. Of note, 44% of our patients were alive at 2 years. No predictive role emerged in our small cohort, according PD-L1, PD-1 and TILs expression, for those obtaining a tumor response. Interactions among alternative factors such as smoking habit, mutational status, time to progression, bone marrow toxicities (ie leucopenia), may have more powerful association with response and clinical outcome. Updated clinical activity and biomarker analysis will be presented.
