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Noemi Reguart
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YI04 - Basics of Immunology (ID 110)
- Event: WCLC 2019
- Type: Young Investigator Session
- Track: Young Investigators
- Presentations: 4
- Now Available
- Moderators:Noemi Reguart, Julie R Brahmer
- Coordinates: 9/08/2019, 13:30 - 15:00, Dublin (1997)
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YI04.01 - PD-L1: Basics of Biology (Now Available) (ID 3708)
13:30 - 15:00 | Presenting Author(s): Ignacio Gil-Bazo
- Abstract
- Presentation
Abstract not provided
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YI04.02 - Immuno-Oncology Trials: Other Endpoints Needed? (Now Available) (ID 3709)
13:30 - 15:00 | Presenting Author(s): Martin Reck
- Abstract
- Presentation
Abstract
The implementation of immunotherapies has completely changed and improved management of patients with non-small-cell lung cancer (NSCLC). In contrast to cytotoxic agents and targeted therapies immunotherapies do not directly affect the tumor but lead to an indirect reactivation of T-cell mediated immune reaction. Therefore different endpoints may be required for an appropriate assessment of efficacy of these novel compounds.
In various trials it could be demonstrated that in contrast to conventional anti tumor therapies, which show an impact on early overall survival (OS), immunotherapies seem to impact the long term survival rates, leading to a phenomenon, which is characterized as „plateau of long term survival“.
In contrast other conventional markers of efficacy like response rate (RR) or progression free survival (PFS) do not appear as the best endpoints to reflect the complete efficacy of immunotherapies at least not in biomarker unselected populations. In a number of randomized phase 3 trials in untreated as well as pretreated patients no significant differences in PFS were shown despite the fact that these trials demonstrated a clear survival benefit in favour of the immunotherapies or immunotherapy combinations. In addition exploratory analyses revealed a survival benefit even in patients, who received immunotherapies beyond progression suggesting that indeed response and PFS may be suboptimal endpoints.
Reflecting these experiences clearly new statistical approaches are needed to focus on OS as probably the most relevant endpoint and to provide the opportunity to capture the late impact of immunotherapies on OS with the prolongation of long-term survival in an adequate way. Currently the first modifications of conventional statistical models are in development evaluating for example the model of disproportional hazard ratios and others in upcoming trials.
Another significant problem is the appearance of „crossing“ Kaplan Meier curves, which has frequently been observed in trials with immunotherapy combinations. These curves suggest, that there are different subpopulations with different sensitivities toward the novel treatments represented. Reflecting the lack of valid predictive biomarkers, which are able to separate the different subgroups, innovative statistical models are needed to define and to describe the benefitting populations.
Lastly the „explosion“ of numerous clinical trials investigating novel immunotherapy combinations will require a modification of our traditional endpoints. Given the number of novel agents and the speed of development it won´t be feasible to conduct individual signal generating phase II trials for each of these agents. New models of platform trials with a new model of statistical assessment are required to provide the opportunity to investigate multiple combinations in one trial and to provide the flexibility to add novel combinations, which just may have been developed to ongoing protocols.
In summary immunotherapies have substantially contributed to therapeutic improvements in NSCLC, but they are requesting a change of our conventional consideration of efficacy. Besides all endpoints of objective efficacy it will be of paramount importance that future endpoints also cover patient relevant endpoints like tolerability, improvement of quality of life or duration of symptom control.
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YI04.03 - The Place of Immunotherapy in NSCLC Therapeutic Algorythm (Now Available) (ID 3710)
13:30 - 15:00 | Presenting Author(s): Rolf Stahel
- Abstract
- Presentation
Abstract
Place of immunotherapy in the NSCLC treatment algorithm
Therapy with immune checkpoint inhibitors has entered standard of care for patients with metastatic NSCLC. Data from extended phase I studies are available on the long-term effect of single agent immune checkpoint inhibition in later line. These studies demonstrate a 16% 5-year survival for nivolumab and pembrolizumab in pretreated patients (1), the impact of PD-L1 expression on long term outcome (2) and the importance of obtaining an objective response (3). Nivolumab and pembrolizumab, both PD-1 directed antibodies and atezolizumab, a PD-L1 directed antibody all have been approved for second or later line therapy based on comparative studies with docetaxel monotherapy demonstrating superior survival and an improved toxicity profile. However, results of recent studies in first line treatment have led to a rapid adoption of immune checkpoint inhibition upfront. This started with the results of the KEYNOTE-024 study demonstrating survival benefit of single agent pembrolizumab over platin-based combination therapy for patients with advanced NSCLC with a tumor proportion score of 50% or higher (4). With these results, PD-L1 testing has entered clinical routine. A series of clinical trials examined the survival impact of adding immune checkpoint inhibition to standard platin-based chemotherapy. Based on these results pembrolizumab added to platin-based chemotherapy, carboplatin and pemetrexed in non-squamous NSCLC (KEYNOTE,-189), carboplatin-paclitaxel or nab-paclitaxel in squamous NSCLC (KEYNOTE-407) (5,6), or alternatively atezolizumab combined with bevacizumab, carboplatin and paclitaxel (IMpower150) or with carboplatin and nab-paclitaxel (Impower130) for non-squamous NSCLC (7,8) have become standard of care, independent of the expression of PD-L1 in tumor tissues. The results of these t and additional trials in first line metastatic NSCLC have been summarized recently (9). Obviously, important questions remain to be answered. One open question regards the upfront treatment of patients with tumors having a strong PD-L1 expression. Lacking randomized data the debate remains open whether pembrolizumab alone or its combination with platin-based chemotherapy would ultimately be the best option. The KEYNOTE-042 trial compared pembrolizumab with platin-based chemotherapy in patients with tumors with at least 1% PD-L1 expression. The trial was positive for pembrolizumab, however the analysis of patient subgroups according to PD-L1 expression suggests the benefit to be mostly derived from patients with strongly expressing tumors and thus these results might only marginally impact clinical practice. A second open question regards the optimal selection of patients which could be spared from upfront treatment with chemotherapy and could benefit from single agent immune checkpoint or combined PD-+/PD-L1 and CTLA4 inhibition. Two trials examining the role of nivolumab in first line treatment (CHECKMATE-026 and CHECKMATE-227) demonstrated that a high tumor mutation burden is predictive for a better response to immune checkpoint inhibition. The later trial (10) and the analysis subgroups of the MYSTIC trial are in support of the hypothesis of potential superior effects of combining PD-1/PD-L1 inhibition with CTLA4 inhibition for patients with a high tumor mutation burden, measured directly in tumor tissue or indirectly on circulating tumor DNA, however, formal prove from prospective trials is pending. It is important to note that with the exception of IMpower150 and Impower130, all other trials excluded patients with known EGFR and ALK mutations, the main reason being the relative lack of efficacy of second line immune checkpoint inhibition in these patients. The benefit in progression-free survival and overall survival seen in patients with EGFR mutated tumors when treated with the atezolizumab bevacizumab quadruplicate, but not with the atezolizumab triplicate lends support for the use of the quadruplicate regimen for patients with EGFR mutated tumors failing tyrosine kinases.
Gettinger, et al. Five-Year Follow-Up of Nivolumab in Previously Treated Advanced Non-Small-Cell Lung Cancer: Results From the CA209-003 Study. J Clin Oncol 2018 Jun 10;36(17):1675-1684
Garon E, et al. Five-Year Overall Survival for Patients With Advanced Non‒Small-Cell Lung Cancer Treated With Pembrolizumab: Results From the Phase I KEYNOTE-001 Study. J Clin Oncol 2019 Jun 2:JCO1900934
Horn, L, et al. Safety and clinical activity of atezolizumab monotherapy in metastatic non-small-cell lung cancer: final results from a phase I study. Eur J Cancer 2018 Sep;101:201-209.
Reck M, et al. Pembrolizumab versus Chemotherapy for PD-L1-Positive Non-Small-Cell Lung Cancer. NEJM 2016 Nov 10;375(19):1823-1833
Ghandi L, et al. Pembrolizumab plus Chemotherapy in Metastatic Non-Small-Cell Lung Cancer. NEJM 2018 May 31;378(22):2078-2092.
Pas-Arez L, et al. Pembrolizumab plus Chemotherapy for Squamous Non-Small-Cell Lung Cancer. 2018 Nov 22;379(21):2040-2051.
Sozinski MA, et al. Atezolizumab for First-Line Treatment of Metastatic Nonsquamous NSCLC. NEJM 2018 Jun 14;378(24):2288-2301.
West H, et al. Atezolizumab in combination with carboplatin plus nab-paclitaxel chemotherapy compared with chemotherapy alone as first-line treatment for metastatic non-squamous non-small-cell lung cancer (IMpower130): a multicentre, randomised, open-label, phase 3 trial. Lancet Oncol 2019 May 20. pii: S1470-2045(19)30167-6.
Peters S, et al. How to Make the Best Use of Immunotherapy as First-Line Treatment for Advanced/Metastatic Non-Small-Cell Lung Cancer. Ann Oncol 2019 Mar 26. pii: mdz109. doi: 10.1093/annonc/mdz109
Hellman MD, et al. Nivolumab plus Ipilimumab in Lung Cancer with a High Tumor Mutational Burden. NEJM 2018 May 31;378(22):2093-2104.
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YI04.04 - How to Select Patients for Immunotherapy (Now Available) (ID 3711)
13:30 - 15:00 | Presenting Author(s): Naiyer A Rizvi
- Abstract
- Presentation
Abstract
With the wide application of immune checkpoint blockade in the treatment of cancer, biomarkers may determine the line of therapy immunotherapy may be administered or when to combine with other agents. Patient selection on clinical characteristics and co-morbidities also need to be incorporated into treatment decision making. An overview of patient characteristics and external influencing factors will be provided in this review including more complicated scenarios such as patients with co-existing autoimmune disease or solid tumor transplant populations.
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Author of
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EP1.01 - Advanced NSCLC (ID 150)
- Event: WCLC 2019
- Type: E-Poster Viewing in the Exhibit Hall
- Track: Advanced NSCLC
- Presentations: 1
- Now Available
- Moderators:
- Coordinates: 9/08/2019, 08:00 - 18:00, Exhibit Hall
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EP1.01-41 - Feasibility of EBUS-TBNA Cytologies for an Extensive Assessment of Predictive Biomarkers in Lung Cancer (Now Available) (ID 1640)
08:00 - 18:00 | Author(s): Noemi Reguart
- Abstract
Background
Clinical guidelines support the determination of several driver genes as well as PD-L1 to drive treatment decisions in non-small cell lung cancer (NSCLC). Endobronchial-ultrasound transbronchial needle aspiration (EBUS-TBNA) cytology specimens are useful for the initial diagnosis of NSCLC, although its capacity to provide enough material for a complete genotyping remains controversial. The aim of this study is to determine the yield of EBUS for a comprehensive multiplex genotyping in patients (pts) with suspected NSCLC.
Method
In this single-center, ongoing, prospective study, samples from mediastinal lymph nodes were obtained from pts undergoing EBUS-TBNA for lung cancer diagnosis/staging. Following malignant confirmation and appropriate cell content by rapid on-site evaluation, the study sample was obtained and formalin-fixed paraffin-embedded (FFPE). Three analytes were evaluated (DNA/RNA/protein). DNA and RNA were extracted and analyzed by Oncomine Solid Tumour panel (22 genes) and a customized nCounter panel (ALK, ROS; RET, NTRK, METDe14). Tumor Proportion Score (TPS) for PD-L1 protein expression was evaluated by an expert pathologist and scored into <1% (negative), 1-49% (weakly positive) and 50% (high).
Result
Twenty-five pts with NSCLC have been included and cytology samples of 20 of them molecularly characterized (5 still in progress). Overall, cytological analysis of EBUS-TBNA yield a complete characterization for the three analytes (DNA/RNA/protein) in 15 pts (75%). EBUS-TBNA sampling was sufficient for both, Nanostring and Oncomine evaluation, in a total of 18 pts (90%): 15 patients (83%) had any alteration detected by oncomine (TP53 61% [11/18],KRAS 44% [8/18], EGFRe 195.5% [1/18], BRAF V600E 5,5% [1/18], DDR2 5.5% [1/18], STK11 11% [2/18]) and 1 pt (5.5 %) by nanostring (METDex14). A total of 19 samples were sufficient for PD-L1 expression scoring (95%). TPS for PD-L1 expression was negative in 8 pts (42%), week in 4 (21%) and high in 7 pts (37%). Overall, half of the pts evaluated (10/20) would be potential candidates for an upfront personalized treatment strategy using targeted agents or immunotherapy.
Conclusion
EBUS-TBNA is a promising alternative source of material for NSCLC genotyping and allows the identification of pts candidates for personalized therapies.
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IBS02 - Making Sense of Treatment with so Many Options: My Algoritm (Ticketed Session) (ID 33)
- Event: WCLC 2019
- Type: Interactive Breakfast Session
- Track: Advanced NSCLC
- Presentations: 1
- Now Available
- Moderators:
- Coordinates: 9/08/2019, 07:00 - 08:00, Toronto (1985)
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IBS02.01 - Making Sense of Treatment with so Many Options: My Algoritm - A European View (Now Available) (ID 3318)
07:00 - 08:00 | Presenting Author(s): Noemi Reguart
- Abstract
- Presentation
Abstract
The treatment paradigm for metastatic non-small cell lung cancer (NSCLC) continues to evolve quickly due to the introduction of immunotherapy and new targeted therapies as well as our increasing knowledge of molecular signaling pathways. As a result of treatments becoming more efficacious and more personalized, outcomes for selected patients with NSCLC are increasing. Algorithms change rapidly and those published one year ago are already outdated today. In this interactive session, we will discuss current treatment algorithm / recommendations for advanced NSCLC patients, those whose tumors have driver oncogenes as well as wild-type lung tumors. We will describe the decision-making process to arrive at a treatment recommendation for a particular patient from the perspective of three different areas of the world: Europe, North-America and Australia. Recommendations for the best standards of care, based on evidence-based medicine and local clinical practice guidelines will be covered and applied in this case to the European local-practice context.
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MA15 - Usage of Computer and Molecular Analysis in Treatment Selection and Disease Prognostication (ID 141)
- Event: WCLC 2019
- Type: Mini Oral Session
- Track: Pathology
- Presentations: 1
- Now Available
- Moderators:John Le Quesne, Noriko Motoi
- Coordinates: 9/09/2019, 15:45 - 17:15, Tokyo (1982)
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MA15.10 - Stromal Markers of Activated Tumor Associated Fibroblasts Predict Poor Survival and Are Associated with Necrosis in Non-Small Cell Lung Cancer (Now Available) (ID 2212)
15:45 - 17:15 | Author(s): Noemi Reguart
- Abstract
- Presentation
Background
Tumor associated fibroblasts (TAFs) are essential contributors of the progression of non-small cell lung cancer (NSCLC). Most lung TAFs exhibit an activated phenotype characterized by the expression of α-SMA and fibrillar collagens. However, the prognostic value of these activation markers in NSCLC remains unclear.
Method
We conducted a retrospective multicentric study of the prognostic value of the standard markers of activated fibroblasts. For this purpose, we conducted a quantitative image analysis of α-SMA immunostaining and picrosirius red staining of fibrillar collagens imaged by bright-field and polarized microscopy, respectively, using tissue microarrays with samples from 220 surgical patients, which elicited a percentage of positive staining area for each marker and patient.
Result
Kaplan-Meier curves showed that all TAF activation markers were significantly associated with poor survival, and their prognostic value was independent of TNM staging as revealed by multivariate analysis, which elicited an adjusted increased risk of death after 3 years of 129% and 94% for fibrillar collagens imaged with bright-field (p = 0.004) and polarized light (p = 0.003), respectively, and of 89% for α-SMA (p = 0.009). We also found a significant association between all TAF activation markers and tumor necrosis, which is often indicative of hypoxia, supporting a pathologic link between tumor desmoplasia and necrosis/hypoxia.
Conclusion
Our findings identify patients with large histologic coverage of fibrillar collagens and α-SMA+ TAFs to be at higher risk of recurrence and death, supporting that they could be considered for adjuvant therapy. Moreover it supports that antifibrotic drugs aiming to target tumor fibrosis may be an effective therapeutic approach to improve survival in NSCLC.
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OA08 - Advanced Models and "Omics" for Therapeutic Development (ID 133)
- Event: WCLC 2019
- Type: Oral Session
- Track: Biology
- Presentations: 1
- Now Available
- Moderators:Luis M Montuenga, Julie George
- Coordinates: 9/09/2019, 11:00 - 12:30, Interlaken (1988)
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OA08.07 - Aberrant Epigenetic SMAD3 Signaling in Tumor-Associated Fibroblasts Modulates Fibrosis and Response to Nintedanib in NSCLC (Now Available) (ID 1972)
11:00 - 12:30 | Author(s): Noemi Reguart
- Abstract
- Presentation
Background
Tumor-associated fibroblasts (TAFs) exhibit a fibrotic phenotype in non-small cell lung cancer (NSCLC) that has beeen associated with critical steps of cancer progression. Paradoxically, we reported that the profibrotic TGF-β transcription factor SMAD3 was epigenetically downregulated through promoter hypermethylation in TAFs from NSCLC patients compared to patient-matched control fibroblasts. In addition, we reported that the antifibrotic drug nintedanib elicited a stronger inhibition of the fibrotic phenotype and its tumor-promoting effects in TAFs from adenocarcinoma (ADC) patients compared to squamous cell carcinoma (SCC) patients upon TGF-β1 stimulation in vitro, which was consistent with the selective therapeutic response to nintedanib observed in a clinical trial in ADC (but not SCC) patients. These previous results support the hypothesis that TGF-β1 signaling may be altered in lung TAFs according to their histologic subtype.
Method
In this study we tested our working hypothesis by determining the expression and activity of SMAD3 and its closely related homologue SMAD2 in patient-derived TAFs and paired control fibroblasts, and by dissecting their potential contribution to the differential therapeutic responses to nintedanib observed in ADC and SCC using in vitro and in vivo preclinical models.
Result
In vitro studies revealed a marked SMAD3 epigenetic repression through promoter hypermethylation, a low pSMAD3/pSMAD2 ratio and a limited fibrotic phenotype selectively in SCC-TAFs. In contrast, ADC-TAFs overexpressed a panel of fibrotic markers upon TGF-β1 stimulation concomitantly with a high pSMAD3/pSMAD2 ratio and a limited SMAD3 promoter methylation. Histologic analysis of a large patient cohort (112 ADC, 96 SCC) confirmed that the extent of fibrosis is larger in ADC than SCC patients. In addition, knocking-down SMAD3 in ADC-TAFs was sufficient to reduce the antifibrotic and antigrowth effects of nintedanib in vitro and in tumor xenografts in vivo. On the other hand, long-term exposure of pulmonary fibroblasts to cigarette smoke condensate was sufficient to hypermethylate the SMAD3 promoter. Since SCC and ADC tumors typically arise in the upper airways and distal pulmonary sites, respectively, it is conceivable that fibroblasts might be more exposed to the smoking epigenetic effects on SMAD3 in SCC.
Conclusion
We report for the first time that tumor fibrosis is higher in ADC than SCC patients, in association with a selective therapeutic response to the antifibrotic drug nintedanib in the former, and identify the subtype-specific extent of SMAD3 epigenetic repression in TAFs and the subsequent aberrant SMAD3/SMAD2 imbalance as major regulatory mechanisms of tumor fibrosis and response to nintedanib in NSCLC.
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P1.01 - Advanced NSCLC (ID 158)
- Event: WCLC 2019
- Type: Poster Viewing in the Exhibit Hall
- Track: Advanced NSCLC
- Presentations: 2
- Moderators:
- Coordinates: 9/08/2019, 09:45 - 18:00, Exhibit Hall
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P1.01-43 - Programmed-Death Ligand 1 Spectrum in a Large Cohort of Genetically Characterized Non-Small Cell Lung Cancer Patients (ID 1760)
09:45 - 18:00 | Author(s): Noemi Reguart
- Abstract
Background
Programmed-death ligand 1 (PD-L1) expression, assessed by immunohistochemistry (IHC), is used to select patients (pts) for checkpoint inhibitors. However, the pattern of PD-L1 expression across rare oncogenic alterations remains poorly characterized. We aimed to evaluate PD-L1 expression in a large dataset of pts with activating molecular alterations to define the potential responsiveness to immunotherapy.
Method
From 2016 to 2018, we prospectively characterized advanced NSCLC pts from a single institution. Tumor Proportion Score (TPS) for PD-L1 protein expression was evaluated by IHC (22C3 clone) using formalin-fixed paraffin-embedded tumor samples and scored into <1% (negative), 1-49% (weakly positive) and >=50% (high). DNA and RNA were extracted and analysed by Oncomine Solid Tumour panel (22 genes) and a customized nCounter-based panel (ALK, ROS1, RET, NTRK1, MET∆14). Associations between PD-L1 expression and the most prevalent driver alterations were assessed. Smoking habit and its possible relationship with PD-L1 expression was also appraised. Statistical analyses were performed using Kruskal-Wallis and Mann-Whitney tests.
Result
TPS for PD-L1 expression was available for a total of 140 patients (pts) fully genotyped. The cohort included: 36% women; 89% adenocarcinoma; 87% smokers. TPS for PD-L1 expression was negative (<1%), weak (1-49%) and high (>=50%) in 40%, 35% and 25% of pts, respectively. Actionable drivers were found in 84 pts (60%) being KRAS (n=43.31%) the most commonly detected, followed by EGFR (n=22.16%), BRAF (n=7.5%), MET∆14 (n=8.6%), ALK (n=3.2%) and ERBB2 (n=1, 1%). Thirty (21.4%) [ME1] pts harboured TP53 co-mutations. By comparing all genotyped cohorts, MET∆14 alteration was associated with higher PD-L1 expression levels compared with other subgroups (median TPS 58.62 vs 25.26, p=0.017[ME2] ). In addition, PD-L1 expression was also higher in pts harbouring any TP53 co-mutation than those with any alteration but TP53-WT (median TPS 41.53 vs 21.45, p=0.035). When KRAS-mut, BRAF-mut and EGFR-mut were evaluated separately, PD-L1 expression was higher in TP53 mutated tumors compared to TP53 WT only in BRAF-mut (p=0.028). Finally, no significant differences were found regarding patients’ smoking status (p=0.527).
Conclusion
Our results suggest differential expression of PD-L1 based on the presence of MET alterations and TP53 mutations and highlight the need of further characterizing PD-L1 expression across oncogenic alterations.
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P1.01-56 - Increased ROS1 and RET Transcripts in Fusion-Negative NSCLC Patients (ID 2477)
09:45 - 18:00 | Presenting Author(s): Noemi Reguart
- Abstract
Background
Fusion involving anaplastic lymphoma kinase (ALK), RET proto-oncogene (RET) or v-ros UR2 sarcoma virus oncogene homolog 1 (ROS1) occur in non-small cell lung cancers (NSCLC) and are important biomarkers for targeted therapies. However, little is known about the RNA expression levels of these genes regardless of fusions.
Method
We used a custom nCounter panel (NanoString Technologies) designed to detect several genetic alterations, including fusions and mRNA expression levels of ALK, ROS1 and RET in formalin-fixed paraffin embedded (FFPE) samples. RNA was purified from NSCLC tumor samples and analyzed with the custom panel. The counts corresponding to the 3’ probes were normalized using the geometrical mean of the housekeeping genes and then added to evaluate total mRNA expression levels. Cut-off values for overexpression were established as the average counts for each gene plus two times the standard deviation.
Result
A total of 400 stage III-IV NSCLC patients (p) from two different institutions were retrospectively analyzed. Overexpression of ALK was found in 55 p (13.8%). Of them, 48 (87%) were also positive for EML4-ALK fusions. One ALK-translocated patient with low levels of ALK mRNA expression did not respond to therapy. Fifteen p (3.8%) showed ROS1 overexpression. In contrast with ALK, only three of them (15%) had a concomitant ROS1 fusion. Among the remaining 12 patients overexpressing ROS1, four were ALK positive, five harbored mutations in EGFR and three were non-smoker females with no known drivers. Regarding RET, high expression levels were found in 14 p (3.5%) and only one of them showed a RET fusion (7%). Among the remaining 13 p, three presented neuroendocrine features and seven were smoker or ex-smoker without other known drivers.
Conclusion
Overexpression of ALK mRNA in NSCLC is associated with EML4-ALK translocations. In contrast, a significant number of fusion negative patients show high ROS1 or RET mRNA levels. Further research is warranted to determine the clinical relevance of this finding.
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P1.03 - Biology (ID 161)
- Event: WCLC 2019
- Type: Poster Viewing in the Exhibit Hall
- Track: Biology
- Presentations: 3
- Moderators:
- Coordinates: 9/08/2019, 09:45 - 18:00, Exhibit Hall
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P1.03-02 - Nintedanib Selectively Inhibits Angiogenesis Induced by the Conditioned Medium of Lung Adenocarcinoma TAFs in Vitro (ID 2193)
09:45 - 18:00 | Author(s): Noemi Reguart
- Abstract
Background
Nintedanib is an antifibrotic and antiangiogenic drug that was clinically approved to treat EGFR-wild-type lung adenocarcinoma (ADC) patients based on the positive therapeutic results elicited in combination with docetaxel in the LUME-Lung1 clinical trial in ADC but not in squamous cell carcinoma (SCC) patients. However, the mechanisms underlying the selective therapeutic effects of nintedanib in ADC remain poorly understood. Tumor-associated fibroblasts (TAFs) are the most abundant stromal cell type and have been implicated in all steps of tumor progression, including growth, angiogenesis, invasion, metastasis and resistance to therapies. Of note, we recently reported that nintedanib elicits larger antifibrotic effects in ADC-TAFs compared to SCC-TAFs in vitro. However, the antiangiogenic effects of nintedanib in TAFs remain unknown.
Method
ADC-TAFs and SCC-TAFs were activated with TGF-β1 in the presence or absence of nintedanib, and the corresponding conditioned medium was used to stimulate endothelial cells (HUVECs and HMVECs) to either migrate or form vascular networks on Matrigel.
Result
Our results showed that the conditioned medium from ADC-TAFs and SCC-TAFs induced an increase in human endothelial cell migration. In contrast, the conditioned medium from ADC-TAFs promoted an increase in angiogenesis that was selectively inhibited by nintedanib, whereas such inhibition was not observed in SCC-TAFs.
Conclusion
Our results reveal that nintedanib inhibits the pro-angiogenic effects elicited by factors secreted by ADC-TAFs but not SCC-TAFs, thereby supporting that angiogenesis may be regulated by TAFs through distinct mechanisms in ADC and SCC.
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P1.03-15 - Non-Invasive Detection of Secondary Resistance Mutations in ALK-Positive NSCLC Patients by Next-Generation Sequencing (ID 1658)
09:45 - 18:00 | Author(s): Noemi Reguart
- Abstract
Background
ALK inhibitors have led to important improvements in ALK-positive non-small cell lung cancer (NSCLC) patient’s survival and quality of life. However, despite the good responses, resistance mutations inevitably emerge. Several resistance mutations in ALK domain have been describe. Remarkably different mutations can confer different sensitivities to different ALK inhibitors. However, 2nd and 3rd line treatment is often prescribe empirically without knowing the molecular mechanism underlying treatment failure.
Method
21 samples from ALK-positive NSCLC patients were collected at disease progression. Circulating Nucleic Acids were isolated from platelets, exosomes and plasma. Libraries were prepared using 20ng of template and Oncomine™ Pan-Cancer Cell-Free Assay. Samples were sequenced on an Ion GeneStudio S5 Plus System. Sequencing data was first analyzed using Torrent Suite software. Subsequently variant calling, annotation and filtering was performed on the Ion Reporter (v5.10) platform using the Oncomine TagSeq Pan-Cancer Liquid Biopsy w2.1 workflow.
Result
In 14 (67%) patients a somatic mutation was identified in the plasma sample collected at disease progression. The average number of mutations detected per sample was 2.6. Noteworthy, 14 mutations were found in oncogenes that have been previously associated with ALK inhibitors resistance (5 mutations in ALK locus, 4 mutations in PIK3CA, 1 mutation in EGFR, 1 mutation in KIT, 1 mutation in KRAS, 1 mutation in MTOR and 1 mutation in MYC). The rest of mutations (N=21) were found in TP53 gene. Secondary resistance mutation in ALK locus occurred in 24% of the cases. Specifically, p.G1269A (N=2), p.G1202E (N=1), p.R1275Q (N=1) mutations were found in ALK-positive NSCLC who had progressed on crizotinib and p.G1202R mutation was found in 1 ALK-positive NSCLC who had progressed on ceritinib.
Conclusion
Secondary ALK-TKI resistance mutations could be detected using liquid biopsies in a high proportion of patients. Non-invasive molecular profiling of samples collected at disease progression is feasible being useful for further treatment selection in ALK-positive NSCLC patients.
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P1.03-26 - Genetic and Molecular Profiling of Non-Smoking Related Lung Adenocarcinomas (ID 1701)
09:45 - 18:00 | Author(s): Noemi Reguart
- Abstract
Background
The etiology and many details of the genomic profile and molecular basis of lung adenocarcinomas (LuADs) in nonsmoker patients remain elusive. Further, the scarcity of primary cultures available from non-smoking related lung adenocarcinomas (NSK-LuADs) contributes to hamper our biological understanding of these tumors.
Method
We established patient-derived cancer cell (PDCs) cultures from NSK-LuADs, and performed whole exome sequencing (WES) and RNA sequencing (RNA-seq) analysis to delineate their genomic architecture. For validations, we analyzed independent cohorts of LuADs.
Result
The analysis revealed non-smoker related alterations such as those at the growth factor receptors RET, ALK, EGFR and ERBB2. There were also mutations affecting signal transduction molecules such as AKT1, BRAF and KRAS, and mutations in tumor suppressor genes, including TP53, CDKN2A, RB1, ARID1A, ATM and STK11. We also identified new fusions and recurrent mutations in some genes, one of them, a possible regulator of gene expression, affecting ten percent of the LuADs, thus constituting a potentially relevant tumor suppressor gene. We also report a predominance of RB1-inactivation, mostly complex intragenic rearrangements (homozygous deletions or duplications) in EGFR-mutant tumors. Three EGFR-/RB1-mutant tumors, treated with EGFR-TKIs, and one EGFR-wild type tumor, treated with standard chemotherapy, developed small cell lung cancer and/or squamous cell carcinoma transformation, evident in the re-biopsies and/or PDCs. Finally, we found pathogenic germ-line mutations at genes associated to familiar-cancer syndromes, especially the TP53-associated Li Fraumeni syndrome, affecting ten percent of EGFR-mutant LuADs patients, underscoring a genetic predisposition origin for a subset of NSK-LuADs.
Conclusion
The recurrent gene inactivation found in candidate gene in LuADs heralds a tumor suppressor role which deserves further exploration. The pre-existent inactivation of RB1 predominates in EGFR-mutant tumors and may underlie an extremely ductile nature, albeit additional gene alterations are required to overcome sensitivity to the TKIs. Given their potential clinical and therapeutic implications, testing for RB1-alterations and for the Li-Fraumeni syndrome in EGFR-mutant LuADs patients may need to be incorporated in the clinical settings.
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P2.03 - Biology (ID 162)
- Event: WCLC 2019
- Type: Poster Viewing in the Exhibit Hall
- Track: Biology
- Presentations: 3
- Moderators:
- Coordinates: 9/09/2019, 10:15 - 18:15, Exhibit Hall
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P2.03-01 - MMP1 Secreted by Cancer Cells Induces a Pro-Tumorigenic Senescent Phenotype in Fibroblasts in Large Cell Carcinoma of the Lung (ID 2195)
10:15 - 18:15 | Author(s): Noemi Reguart
- Abstract
Background
Tumor associated fibroblasts (TAFs) are key effector cells of cancer progression. Senescent TAFs have been reported in a growing list of aggressive cancer subtypes including the aggressive subtype large cell carcinoma (LCC) of the lung. We previously reported that LCC cells induce fibroblast senescence in normal fibroblasts after co-culture, revealing that paracrine signaling must be involved. Moreover, we found that senescent fibroblasts secrete factors that stimulate the growth and invasion of LCC cells beyond the stimulation elicited by non-senescent fibroblasts, supporting that fibroblast senescence may contribute to the aggressive nature of LCC. Whole-genome transcriptional profiling on a panel of non-small cell lung cancer (NSCLC) cell lines, including adenocarcinoma (ADC), squamous cell carcinoma (SCC) and LCC, identified MMP1 as highly overexpressed in LCC cells compared to non-LCC cells. Here we examined the role of MMP1 in LCC cells in the paracrine induction of fibroblast senescence.
Method
We silenced MMP-1 expression in LCC cancer cell lines by shRNA and analyzed common senescence markers after co-culture with normal fibroblasts, including β-galactosidase staining as well as the expression of common factors of the senescence-associated secretory phenotype (SASP) by qRT-PCR. In addition, the growth and invasion pro-tumorigenic effects elicited by the conditioned medium of fibroblasts co-cultured with shMMP1 or shscramble LCC cells was analyzed.
Result
Knocking-down MMP1 in LCC cells was sufficient to abrogate fibroblast senescence induction in co-cultures with LCC cells, as well as the growth and invasion enhancement of LCC cells elicited by the conditioned medium of fibroblasts. The addition of active recombinant MMP1 partially rescued the fibroblast senescent phenotype in co-culture, yet it was not sufficient to induce senescence when added to fibroblasts cultured alone.
Conclusion
Our results unveil a process of “niche construction” by LCC cells that is driven by the overexpression of MMP1, which induces senescence in adjacent fibroblasts. Moreover, our results unveil a novel biological function of MMP1 (i.e. paracrine senescence induction in fibroblasts), that is strikingly different from its well-known collagenolytic function. Our results also show that MMP1 is necessary but not sufficient to induce fibroblast senescence. Moreover, our findings support that the aberrant carcinoma cell-fibroblast crosstalk mediated by MMP1 may be a suitable therapeutic target in LCC.
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P2.03-17 - Optimization of an Ex-Vivo Preclinical Model for Drug Testing (ID 2385)
10:15 - 18:15 | Author(s): Noemi Reguart
- Abstract
Background
Predicting drug response in advanced lung cancer patients remains a major challenge. A promising approach to tackle this challenge is based on testing drugs in an ex-vivo preclinical model using cultured precision-cut fresh tumour slices, which maintain not only the tumour itself but also its microenvironment. However, even though different protocols have been reported for ex-vivo models with different time-windows in a growing list of cancer types, no standard protocols are currently defined in lung cancer. To address this limitation, we have begun to optimize a protocol for an ex-vivo preclinical model for drug testing in lung cancer, using samples from the standard in vivo model of bleomycin-induced lung fibrosis, which resembles the fibrotic stroma of non-small cell lung cancers.
Method
Bleomycin-induced fibrotic and control rat lungs were precision-cut with a tissue-chopper and cultured on cell culture inserts up to 12 days in a sterile setting. Two main variables were tested: the coating of the cell culture insert (bare or collagen-I coated), and the interaction with the culture medium (submerged or floating). Small lung pieces were collected at days 0, 2, 6, 9 and 12 and examined by hematoxylin-eosin staining to assess their integrity and viability.
Result
Tissue architecture was maintained equally well in all culture conditions up to 12 days. In contrast, cellular content appeared to decline after day 6; since only subsets of cells in the periphery of the explants remained viable at day 9-12. No contamination was detected at any culture time.
Conclusion
Our preliminary results suggest that our protocols for an ex-vivo preclinical model maintain the integrity and viability of lung cells and their microenvironment up to one week, which is comparable to time-windows previously reported in other cancer types. This time-window is expected to be sufficient to test drug responses, thereby underlying the potential of this preclinical model for drug testing as well as for the study of drug resistance mechanisms.
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P2.03-35 - Stromal TIMP-1 Drives Tumor Progression in Lung Adenocarcinoma Through CD63 Interaction (ID 2196)
10:15 - 18:15 | Author(s): Noemi Reguart
- Abstract
Background
Tumor associated fibroblasts (TAFs) are important regulators of tumor growth and resistance to therapies. We have recently shown that TAFs in vitro from lung adenocarcinoma (ADC) and squamous cell carcinoma (SCC) respond positively to the antifibrotic drug nintedanib in the former only, thereby mimicking the selective therapeutic effects of nintedanib in ADC reported in the LUME-Lung1 clinical trial. We also showed that the tumor-promoting effects of TAFs are driven by different mechanisms in ADC and SCC. However, it remains to be elucidated the key signaling molecules involved in the aberrant fibroblast-carcinoma crosstalk in ADC. Tissue inhibitor of metalloproteinases 1 (TIMP-1) is a multifunctional protein that has been associated with poor prognosis in lung cancer and other cancer types, and is downregulated by nintedanib in a bleomycin model of pulmonary fibrosis. Moreover, our preliminary analysis revealed that the TIMP-1 cell surface binding protein CD63, is overexpressed in ADC compared to SCC. Therefore, our objective was to study whether the selective tumor-promoting effects of ADC-TAFs are mediated by the interaction of stromal TIMP-1 with epithelial CD63.
Method
ADC-TAFs and SCC-TAFs were stimulated with TGF-β1 in the presence or absence of nintedanib, and the TIMP-1 content in their conditioned medium was determined by ELISA. TIMP-1 was reduced in ADC-TAFs by siRNA, and the corresponding conditioned medium was used to assess the impact of TIMP-1 on the growth, invasion and survival of the CD63-high ADC cell line H1437. Likewise, CD63 expression in H1437 cells was reduced, by siRNA. In addition we performed immunohistochemical analyses of CD63 in tissue sections from lung cancer patients.
Result
Our results showed that TIMP-1 secretion induced by TGF-β1 is significantly larger in ADC-TAFs compared to SCC-TAFs. Likewise, nintedanib elicited a larger downregulation of TIMP-1 secretion in ADC-TAFs compared to SCC-TAFs. We also confirmed that CD63 expression is higher in ADC patients than SCC, and revealed that knocking-down CD63 in H1437 ADC cells is sufficient to reduce the growth and invasion elicited by the conditioned medium of activated ADC-TAFs. Likewise, knocking-down TIMP-1 in ADC-TAFs was sufficient to downregulate the growth, invasion and survival of H1437 elicited by the conditioned medium of these TAFs.
Conclusion
Collectively, our results unveil a novel stroma-carcinoma interaction driven by TIMP-1 and CD63 selectively in lung ADC, and support that such crosstalk is a major regulator of the aberrant tumor-promoting effects of ADC-TAFs that are downregulated selectively by nintedanib.
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P2.04 - Immuno-oncology (ID 167)
- Event: WCLC 2019
- Type: Poster Viewing in the Exhibit Hall
- Track: Immuno-oncology
- Presentations: 2
- Moderators:
- Coordinates: 9/09/2019, 10:15 - 18:15, Exhibit Hall
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P2.04-22 - Programmed Death 1-mRNA Expression Predicts Benefit to Anti-PD1 Monotherapy in a Prospective Cohort of Advanced NSCLC (ID 1092)
10:15 - 18:15 | Author(s): Noemi Reguart
- Abstract
Background
Immunotherapy (IO) targeting PD1 or PD-L1 represents a new treatment option for patients (pts) with advanced non-small cell lung cancer (NSCLC). Besides PD-L1 IHC, other predictive biomarkers are being explored as potential predictors of outcomes. Our group has recently described an association between PD1-mRNA expression and response to IO in a retrospective multi-tumor dataset (Annals Oncol 2018). We aimed to corroborate these results in a prospective cohort of advanced NSCLC.
Method
We prospectively evaluated the expression of 7 immune-related genes (CD4, CD8, PD1, PDL1, IFNG, GZMM andFOXP3) and 5 housekeeping genes in formalin-fixed paraffin-embedded tumor samples obtained before anti-PD1 therapy using the nCounter platform (Nanostring Technologies). The study cohort included consecutive pts with advanced NSCLC, ECOG/PS 0-1, no targetable oncogenes, treated in the first or second-line setting with anti-PD1 monotherapy from June 2017 to January 2019. Associations between the expression of PD1 mRNA (as a continuous variable and using a previously defined pre-specified cutpoint) and response (complete and partial response) were assessed using logistic regression analysis. Kaplan-Meier method was used for survival analysis. PD-L1 IHC tumor cell expression was assessed using the 22C3 clone. Pearson correlation between PD-L1 IHC and PD1 mRNA was explored.
Result
A total of 43 pts were included (men 79%; adenocarcinoma 53%; nivolumab 55%; pembrolizumab 45%; first-line 30%; second-line 70%). Response occurred in 23% of pts and was significantly associated with PD1 (p=0.029) and FOXP3 (p=0.035) expression. Using the pre-established PD1 cutoff (Annals Oncol 2018), 37% and 63% samples were PD1-high and PD1-low, respectively. PD1-high was significantly associated with increased overall response rate (ORR) (43% vs 11%, OR=6.22 [CI=1.31-29.44], p=0.021) and progression-free survival (HR 0.36 [CI= 0.14-0.90], p=0.028) but not with overall survival (p=0.117). PD-L1 IHC expression was available in 35 cases, of which 46% had high (>=50%) expression levels. A moderate concordance (0.49) was observed between PD-L1 IHC and PD1-mRNA. In this subset analysis, high PD-L1 IHC was significantly associated with response (50% vs 11%, OR=8.50 [CI= 1.45-49.53], p=0.043). Importantly, when combining predefined high/low-sets for both biomarkers (PD-L1 IHC/PD1-mRNA), response was significantly increased in PD-L1-high/PD1-high compared to PD-L1-low/PD1-low (ORR 58% vs 0%, p=0.019).
Conclusion
PD1-mRNA expression is associated with response to anti-PD1 monotherapy and can increase the predictive ability of PD-L1 IHC. Further validation of these findings in pivotal clinical trials evaluating IO in advanced NSCLC pts seems warranted.
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P2.04-61 - Preliminary Report of a Multidisciplinary Task Group for the Study of Immune-Mediated Pulmonary Toxicity (ID 1647)
10:15 - 18:15 | Author(s): Noemi Reguart
- Abstract
Background
Immunotherapy (IO) is now the standard of care for many tumor types. However, it is not free of risks, being pulmonary toxicity one of the most relevant immune-related adverse event due to its severity. Differential diagnosis with other pulmonary complications such as infections or tumor dissemination further complicates its management.
Method
In order to raise awareness, gather information, and to discuss early management strategies in patients (pts) with immune-related interstitial lung disease (irILD), in 2017 we created a multidisciplinary task group comprised of pneumologists, pathologists, oncologists and radiologists. We herein report the main features of the first series of pts treated with IO who subsequently developed ILD, prospectively identified from a tertiary University Hospital over a period of two years (2017-2019), focusing on clinical presentation, radiological patterns, outcomes and therapeutic intervention.
Result
We identified a total of 23 pts with suspicion of irILD. Patients mainly received programmed cell death-1 (PD-L1) inhibitors (61%). Main characteristics are summarized in Table 1. ILD occurred more often in males, and former or current smokers (91%), with a median age of 62 years. The most common radiological pattern was the presence of ground-glass opacities (87%), followed by consolidations (61%). Forty-eight percent of the cases had grade 3 severity according to the Common Terminology Criteria for Adverse Events (CTCAE). Thirteen of the patients (57%) underwent a fibrobronchoscopy during the diagnostic period and a specific microorganism was isolated from BAL in three cases (13%) (Aspergillus fumigatus, cytomegalovirus and herpes type 1 virus). Ten pts (43%) underwent transbronchial biopsies being focal organizing pneumonia and desquamative changes the most common pathological patterns observed. Twenty patients (87%) received prednisone (1mg/kg/day) and thirteen of them (57%) also received antibiotic treatment.
ConclusionPatients (%)
Mean age (years)
61.63 ± 12.35
Gender
Male
19 (83%)
Female
4 (17%)
Smoking history
Current
2 (9%)
Never
7 (30%)
Former
14 (61%)
Type of cancer
Lung
8 (35%)
Kidney
5 (22%)
Skin
4 (17%)
Others*
6 (26%)
* Others: haemathologic, bladder, liver, sigma, urothelial, timic.
Immunotherapy
Nivolumab
8 (35%)
Pembrolizumab
6 (26%)
Durvalumab
2 (9%)
Others*
7 (30%)
* Others: Atezolizumab, Nivolumab+Ipilimumab, Atezolizumab+Daratumumab, Atezolumab+Bevacizumab, CX-072, Nivolumab/Nivolumab+Ipilipumab, Avelumab.
Immuno-mediated pulmonary toxicity is a rare but severe complication that carries a significant mortality. Due to their complexity, multidisciplinary approach is required to provide an adequate treatment and to guarantee early intervention.
