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E. Brambilla
Moderator of
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MA02 - RNA in Lung Cancer (ID 377)
- Event: WCLC 2016
- Type: Mini Oral Session
- Track: Biology/Pathology
- Presentations: 10
- Moderators:E. Brambilla, M. Noguchi
- Coordinates: 12/05/2016, 14:20 - 15:50, Stolz 2
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MA02.01 - Extracellular Vescicle miRNAs Regulate Gene Expression in Local Lung Adenocarcinoma Endothelial Cells (ID 4655)
14:20 - 15:50 | Author(s): J. Lawson, C. Dickman, R. Towle, J. Jabalee, S. Lam, C. Garnis
- Abstract
- Presentation
Background:
Extracellular vesicles are small vesicles released from all cell types which can be used as a form of cell to cell communication. Recently these extracellular vesicles have been shown to play a key role in cancer development, growth, progression and angiogenesis. These extracellular vesicles are loaded with functional mRNAs, miRNAs and proteins which can be transferred from one cell to another. Extracellular vesicles have been known to enter neighboring cells including the surrounding stroma, and even enter biofluids. Our research shows that miRNAs transferred from lung adenocarcinoma cells through extracellular vesicles influence gene expression in endothelial cells and enhance their ability to form new blood vessels.
Methods:
Using 5 lung adenocarcinoma cell lines (H1395, H1437, H2073, H2228 and H2347) we isolated extracellular vesicles using differential ultracentrifugation. RNA was extracted from the extracellular vesicles as well as the cells from which they were derived and profiled for 742 miRNAs using the miRCURY LNA[TM] Universal RT miRNA PCR system (Exiqon) to identify miRNAs that were enriched by at least 4-fold in the extracellular vesicles. Tube formation assays were conducted on a commonly used endothelial cell line HMEC-1.
Results:
We found an enrichment of a select set of miRNAs within lung adenocarcinoma extracellular vesicles. These miRNAs have previously been identified as tumor suppressors: miR-142-3p, miR-143-3p, miR-144-3p, miR-145-5p, miR-150-5p, miR-223-3p, miR-451a, miR-486-5p, miR-605-5p in various cancer types. When extracellular vesicles are isolated from miR-143 and miR-145 over expressing adenocarcinoma lines they contain an increase in their over expressed miRNAs. When these miRNA enriched exosomes were incubated with HMEC-1 cells, we observed an increase in their ability to form new blood vessels and a decrease in the expression of CAMK1D in the endothelial cells. miR-143-3p and miR-145-5p were also found to be enriched in serum samples draining directly from lung adenocarcinoma tumors compared to arterial serum.
Conclusion:
Extracellular vesicles originating from lung adenocarcinoma cells can enter into endothelial cells and increase their ability to form new blood vessels through extracellular vesicle transfer of miR-145/miR-143 suggesting that this form of communication increases angiogenesis within lung adenocarcinoma tumors.
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MA02.02 - A Novel 5-miR Signature Shows Promise as a Diagnostic Tool and as a Predictor of Cisplatin Response in NSCLC (ID 5948)
14:20 - 15:50 | Author(s): L. Mac Donagh, S.G. Gray, S. Cuffe, S.P. Finn, N. Fitzgerald, V. Young, R. Ryan, S. Nicholson, N. Leonard, K. O’byrne, M.P. Barr
- Abstract
Background:
MicroRNAs are a class of small non-coding RNAs that range in size from 19-25 nucleotides. They have been shown to regulate a number of processes within tumour biology, including metastasis, invasion and angiogenesis. More recently, miRNAs have been linked to chemoresistance in solid tumours, including lung cancer.
Methods:
MicroRNA expression within an isogenic panel of age-matched parent (PT) and cisplatin resistant (CisR) NSCLC cell lines was profiled using the 7[th] generation miRCURY LNA arrays (Exiqon). Significantly altered miRNAs within the CisR sublines were manipulated using antagomirs (Exiqon) and Pre-miRs (Ambion) and functional studies were carried out in the presence and absence of cisplatin. To examine the translational relevance of these miRNAs, their expression was examined in a cohort of chemo-naïve patient-matched normal and lung tumour tissue and serum from NSCLC patients of different histologies. To create a xenograft model of cisplatin resistance 1x10[3 ]cells H460 PT or CisR cells were injected into 5-7week old NOD/SCID mice. Tumour volume was measured over time and harvested once the tumour mass measured 500mm[3] and formalin-fixed and paraffin embedded (FFPE). Expression of the 5-miR signature was analysed within FFPE murine tumours and cisplatin resistance was investigated relative to cisplatin sensitive controls.
Results:
Profiling and subsequent validation revealed a 5-miR signature associated with our model of cisplatin resistance (miR-30a-3p, miR-30b-5p, miR-30c-5p, miR-34a-5p, miR-4286). Inhibition of the miR-30 family and miR-34a-5p reduced clonogenic survival of CisR cells when treated cisplatin. Expression of the miRNA signature was significantly altered in both adenocarcinoma (AD) and squamous cell carcinoma (SCC) relative to matched normal lung tissue and between SCC and AD tissue. miR-4286 was significantly up-regulated in SCC sera compared to normal control and AD sera. Similarly to the cell line expression of the miRNAs, the miR-30 family members and miR-34a-5p were up-regulated in the CisR xenograft FFPE tissue relative to PT.
Conclusion:
A novel miRNA signature associated with cisplatin resistance was identified in vitro, genetic manipulation of which altered clonogenic response to cisplatin. The 5-miR signature shows both diagnostic and prognostic biomarker potential across a number of diagnostically relevant biological mediums.
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MA02.03 - Expression of Oncofetal miRNAs Inactivates NFIB, a Developmental Transcription Factor Linked to Tumour Aggressiveness in Lung Adenocarcinoma (ID 5224)
14:20 - 15:50 | Author(s): D.D. Becker-Santos, B.C. Minatel, K.M. Lonergan, K.L. Thu, J.C. English, V.D. Martinez, C.E. Macaulay, W.W. Lockwood, W.P. Robinson, I. Jurisica, S. Lam, W.L. Lam
- Abstract
- Presentation
Background:
Fetal and tumour development share striking similarities, such as intense cell proliferation, angiogenesis, increased cell motility, and immune evasion. Molecular regulators, including microRNAs (miRNAs), play important roles in both fetal lung development and in the malignant transformation of adult lung cells. Consequently, investigation of lung tumour biology in the context of lung development may reveal key regulatory mechanisms that tumours hijack from normal development, which potentially play critical roles in the pathology of lung cancer.
Methods:
131 pairs of non-small cell lung cancer (NSCLC) tumour and non-malignant lung tissues and 15 human fetal lung tissue samples were profiled by miRNA-sequencing. Genes controlled by the oncofetal miRNAs identified were first investigated by miRNA-Data-Integration-Portal (mirDIP) prediction, followed by luciferase-reporter assays. Associations between patient survival and mRNA expression of oncofetal miRNA-gene targets were evaluated in independent samples (>1,400 cases) across multiple NSCLC cohorts. Immunohistochemical analysis of oncofetal miRNA targets was performed on 96 lung adenocarcinoma (LUAD) specimens.
Results:
We describe for the first time a comprehensive characterization of miRNA expression in human fetal lung tissue, and identified numerous miRNAs that recapitulate their fetal expression patterns in NSCLC. Nuclear Factor I/B (NFIB), a transcription factor essential for lung development, was identified as being frequently targeted by these oncofetal miRNAs. Overexpression of the oncofetal miRNA miR-92b-3p, significantly reduced NFIB levels in vitro. Concordantly, analysis of NFIB expression in multiple NSCLC cohorts revealed its frequent underexpression in tumours (~40-70%). This is in contrast with its recurrent oncogenic overexpression recently reported in SCLC. Low expression of NFIB was significantly associated with poorer survival in LUAD patients but not in squamous cell carcinoma patients, consistent with the functional role of NFIB in distal lung cell differentiation (i.e., precursor cells of LUAD). Furthermore, an NFIB-related gene signature was identified in LUAD tumours, comprising several well-known lung differentiation markers (e.g., TTF-1, SFTPB, ABCA3). The underexpression of NFIB protein was ultimately validated in LUAD specimens, which also revealed that tumours presenting lower levels of this transcription factor are associated with higher grade, biologically more aggressive LUAD (invasive mucinous, micropapillary and solid subtypes).
Conclusion:
This work has revealed a prominent mechanism for the downregulation of NFIB, a transcription factor essential for lung differentiation, which we found to be associated with aggressive phenotypes of LUAD and consequently, poor patient survival. Restoration of NFIB expression, specifically in LUAD, has the potential to induce lung cell differentiation and thereby reduce tumour aggressiveness.
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MA02.04 - Discussant for MA02.01, MA02.02, MA02.03 (ID 6958)
14:20 - 15:50 | Author(s): L.M. Montuenga
- Abstract
- Presentation
Abstract not provided
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MA02.05 - Distinct Angiogenic microRNA-mRNA Expression Profiles among Subtypes of Lung Adenocarcinoma (ID 5464)
14:20 - 15:50 | Author(s): M. Giordano, L. Boldrini, A. Servadio, M. Lucchi, F. Melfi, A. Mussi, G. Fontanini
- Abstract
- Presentation
Background:
Non-small cell lung cancer (NSCLC) accounts for 80% of all lung cancers and adenocarcinoma (ADC) represents the most common histological type with a heterogeneous pattern of growth classified as lepidic, acinar, papillary, solid, and micropapillary. For ADC there are restricted available therapeutic options except for patients that could benefit from target therapy. A valuable therapeutic strategy is represented by angiogenesis inhibitors such as bevacizumab that has been approved for the treatment of NSCLC patients. However, there are concerns about its treatment-related toxicity and the identification of new reliable biomarkers to stratify patients who can really benefit from antiangiogenic drugs is urgently needed. Using miRNA target prediction tools, we selected and investigated the expression level of a panel of miRNAs togheter with their mRNA target involved in the angiogenesis pathway.
Methods:
We designed a custom codeset including probes for six genes (VEGF-A, FLT1, KDR, FLT4, PDGFRa and PDGFRb) and sixteen miRNAs. The expression analysis was performed by the nCounter System® (NanoString Technologies) directly on RNA, enriched of small RNA, purified from the formalin-fixed and paraffin-embedded tumor tissues of 80 ADC patients. Of these 25 were predominatly lepidic (31.25%), 24 were predominatly solid (30%), 20 were predominatly acinar (16%), 11 were predominatly papillary (13.75%).
Results:
Comparing the expression levels of mRNAs with the different histological ADC subtypes we found a significant higher expression of VEGF-A in papillary than in other subtypes (p=0.02). In contrast PDGFRa and PDGFRb were upregulated in lepidic and downregulated in papillary subtypes (both p=0.03). Among 16 miRNAs that target the angiogenic mRNA, 6 were significantly downregulated in papillary compared to other groups.
Conclusion:
Our data suggest a distinct angiogenic miRNA-mRNA expression profile among the subtypes of ADC. The higher level of VEGF-A in papillary than in lepidic subtypes could represent a useful biomarker to stratify patients who can effectively treated with bevacizumab, which is directed against VEGF. Moreover, the regulation of angiogenic mRNA factors by miRNAs could provide a novel therapeutic approach based on their expression pattern specific for distinct ADC subtypes. Further studies are nedeed in a larger cohort of patients to confirm our results and to investigate whether different rates of response to treatment are observed among patients stratified according to the proposed biomarkers.
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MA02.06 - Discussant for MA02.05 (ID 6957)
14:20 - 15:50 | Author(s): R. Rosell
- Abstract
- Presentation
Abstract not provided
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- Abstract
- Presentation
Background:
Non-small-cell lung cancer (NSCLC) is one of the most common and high mortality rate carcinoma in China which biomarkers for diagnosis are limited. Therefore, novel biomarkers and methods with increased specificity for diagnosis are explored and required. For now, liquid biopsy for lung cancer oncogenes and next generation sequencing technique are extensive employed in NSCLC. However, increasing evidence illustrates that exosomal microRNAs in circulating fluids provide a promising way as biomarkers for noninvasive cancer diagnosis. Exosomes are 30–150 nm particles which are released from cells into the extracellular environment and stable miRNAs have been identified in plasma exosomes, which play important role in cell communication. Furthermore, exosomal miRNAs present different profiles between patients with cancer and healthy individuals. Whether exosomal miRNAs could benefit NSCLC patient diagnosis remains to be explored.
Methods:
Blood samples were collected from 40 NSCLC patients and 24 healthy volunteers matched with age, gender and blood collection time. Plasma exosomes were accessed by 110,000×g ultracentrifugation and visualized by NS300 equipment. The raw data of exosomal miRNA profiles of NSCLC patients and healthy individuals were generated by NGS around 400× read depth and its expression were measured by Taqman probe quantitive PCR
Results:
In the present study, we revealed that nearly half of exosome RNA was miRNA and NSCLS patients expressed a set of exosomal miRNAs with specificity compared with healthy volunteers. We demonstrated that miR-126-5p and miR-21-3p were down-regulated in NSCLC patients. In addition, we showed that the expression level of miR-124-3p and miR-99a-3p in NSCLC patients was higher than that of healthy individuals. Furthermore, we found miR-99a-3p was clinical stages related in NSCLC patient plasma and miR-375-3p was a potential biomarker for diagnosis and prognosis in NSCLC.
Conclusion:
Exosomal miRNAs in plasma could indicate the progress of NSCLC and a combination of the explored miRNA could serve as a promising biomarker for NSCLS diagnosis and prognosis.
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MA02.08 - Deregulation of Cis-Acting Long Non-Coding RNAs in Non-Small Cell Lung Cancer (ID 6303)
14:20 - 15:50 | Author(s): A.P. Sage, G. Stewart, D.A. Rowbotham, K.S.S. Enfield, V.D. Martinez, S. Lam, W.L. Lam
- Abstract
- Presentation
Background:
Lung cancer remains the cause of the most cancer-related deaths each year, with a 5 year survival rate of less than 17%. Targeted therapeutics have been developed against drivers of the lung adenocarcinoma (AC) subtype, but are relevant only to the proportion of patients harbouring these genetic aberrations, emphasizing the need to explore alternative mechanisms of AC development. Natural antisense transcripts (NATs) are long non-coding RNA (lncRNA) products expressed from the opposite strand of coding mRNAs. NATs can function in cis or trans to regulate the transcriptional activity of their cognate gene partner in either a positive or negative fashion. Here we take a novel approach to identify cis- NATs deregulated in lung AC, and explore the function of these genes with regards to their protein coding partner genes.
Methods:
We performed RNA-sequencing on a set of 36 lung AC and matched non-malignant lung tissues. A sign-rank test was used to identify NATs and partner genes with significantly altered expression between tumor and matched normal tissues. These findings were validated in an external dataset of 50 lung AC tumors with matched non-malignant tissue obtained from The Cancer Genome Atlas (TCGA). Survival analysis was performed using a Cox Proportional hazard model, as well as the log-rank method.
Results:
Analysis of Illumina Hi-seq data from TCGA revealed the majority (79%) of deregulated sense-antisense partnerships observed in AC displayed concordant regulation. However, several discordant cis-NAT pairs were identified including an antisense to OPA INTERACTING PROTEIN 5 (OIP5), a gene required for chromatin segregation, as well as an antisense to HIGH MOBILITY GROUP A1 (HMGA1) a gene involved in the metastatic progression of many cancer types. Both the antisense to OIP5 (OIP5-AS1) as well as the antisense to HMGA1, (HMGA1-AS1) were significantly underexpressed in AC, while we find the overlapping protein coding partner genes to be significantly overexpressed, suggesting that these genes may negatively regulate their sense counterparts. In addition both OIP5 and HMGA1 are significantly associated with 5-year survival. Patients with higher expression levels of either of these genes had a significantly shorter overall survival time than patients with low expression levels, highlighting the potential clinical importance of these genes.
Conclusion:
This study characterizes the landscape of antisense expression in AC and highlights novel mechanisms of oncogene regulation through natural antisense transcripts. Characterizing these oncogene regulatory mechanisms could uncover therapeutic intervention points and further our understanding of AC biology.
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MA02.09 - Long Non-Coding RNA Expression from Pseudogene Loci as a Novel Mechanism of Cancer Gene Regulation (ID 6287)
14:20 - 15:50 | Author(s): G. Stewart, K.S.S. Enfield, V.D. Martinez, E.A. Marshall, S. Lam, W.L. Lam
- Abstract
- Presentation
Background:
The advent of next generation sequencing has lead to the discovery of the functional importance of non-coding RNAs (ncRNAs) in a wide variety of cellular processes, and these genes can be exploited by tumours to drive the hallmarks of cancer. Pseudogenes are DNA sequences that are defunct relatives of their functional parent genes but retain high sequence homology. Long non-coding RNAs (lncRNAs) have been shown to regulate protein-coding genes; however, complex folding patterns make lncRNA function difficult to predict. Several lncRNAs expressed from pseudogene loci have been shown to regulate the protein-coding parent genes of these pseudogenes in trans due to sequence complementarity. The biological impact of this mechanism has not been investigated in lung adenocarcinoma (LUAD). We hypothesize that expression changes in lncRNAs expressed from pseudogene loci can affect the expression of corresponding protein-coding parent genes in trans, and that these events provide an alternative mechanism of cancer gene deregulation in LUAD tumourigenesis.
Methods:
We analysed RNA-seq data from 50 LUAD with matched non-malignant tissue obtained from the TCGA for both protein-coding and non-coding gene expression. Significantly differentially expressed lncRNAs located within pseudogene loci were identified by sign-rank test (p<0.001). Mann Whitney U-tests were used to identify lncRNA-parent gene pairs which significantly correlated expression, and survival analysis was performed using a Cox proportional hazard model.
Results:
Our analysis has identified 172 lncRNAs expressed from pseudogene loci that were significantly deregulated in LUAD. Remarkably, many of these lncRNAs were expressed from the loci of pseudogenes related to known cancer genes. One of these lncRNAs, CTD-2583A14.8, was expressed from a pseudogene to ubiquitin-conjugating enzyme E2C (UBE2C), which regulates tumor growth, apoptosis, and angiogenesis through phospho-ERK1/2. We find CTD-2583A14.8 as well as the UBE2C parent gene to be significantly upregulated in LUAD tumours compared to matched normal tissue. Furthermore, tumours with higher levels of CTD-2583A14.8 have significantly higher levels of UBE2C expression than tumours with low levels of CTD-2583A14.8, indicating that CTD-2583A14.8 may positively regulate UBE2C in trans.
Conclusion:
Here we show expression of lncRNAs within pseudogene loci is deregulated in LUAD, and can correlate with the expression of their protein-coding counterparts. Many of these genes associated with this putative lncRNA-pseudogene-protein-coding axis have previously been implicated in cancer. Therefore, this represents an alternative mechanism of cancer gene deregulation, and may represent novel therapeutic intervention points for the treatment of LUAD.
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MA02.10 - Discussant for MA02.07, MA02.08, MA02.09 (ID 7108)
14:20 - 15:50 | Author(s): B. Zaric
- Abstract
- Presentation
Abstract not provided
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Author of
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ED07 - Classification and Druggable Targets of Thoracic Tumors (ID 272)
- Event: WCLC 2016
- Type: Education Session
- Track: Biology/Pathology
- Presentations: 1
- Moderators:A.F. Gazdar, H. Popper
- Coordinates: 12/06/2016, 11:00 - 12:30, Hall C1
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ED07.02 - The 2015 WHO Classification of Neuroendocrine Tumors (ID 6458)
11:00 - 12:30 | Author(s): E. Brambilla
- Abstract
- Presentation
Abstract:
The 2015 WHO Classification of Neuroendocrine Tumors Elisabeth Brambilla, Professor of Pathology, Department of Pathology; CHU Grenoble INSERM U 1209; Institute of Advanced Biosciences; University Grenoble Alpes; France Neuroendocrine lung tumors were considered as separate entities in the previous WHO classification 2004: the carcinoid tumors, small cell lung carcinoma (SCLC) and large cell neuroendocrine carcinoma (LCNEC) were grouped separately. However, in the current WHO 2015 classification, they are grouped together [1]. They are listed in the order of their frequency with SCLC first as it is the most common. SCLC (15% of lung tumors) is a malignant epithelial tumor which consist of densely packed small cells with scant cytoplasm, finely dispersed granular chromatin and absent or inconspicuous nucleoli. In contrast LCNEC is made of large cells and should show both neuroendocrine morphology (rosettes, palisades) and immunohistochemical neuroendocrine markers (at least one). Both SCLC and LCNEC can be pure or combined with NSCLC components but keep their diagnostic priority (SCLC-or LCNEC- combined). Carcinoid tumors are neuroendocrine malignancies accounting for <1% of all lung cancer, divided in two categories with highly different frequencies, the typical and atypical carcinoid, the last being extremely rare. Typical carcinoids are carcinoid tumors with <2mm[2] and lacking necrosis. They measure ≥0.5 cm in size. Atypical carcinoids are carcinoid tumors with 2-10 mitoses per 2mm[2] and/or foci of necrosis. Despite the grouping of these tumors together, it is clear that the carcinoids have major clinical, epidemiologic, histologic and genetic differences compared to the high grade SCLC and LCNEC. Carcinoid patients are significantly younger, have a better prognosis and lack the strong association with smoking that applies for SCLC and LCNEC. Also compared to carcinoid tumors, SCLC and LCNEC have much higher mitotic rates (more than 11 per 2mm[2]), more necrosis and can show combinations with other lung cancer types including adenocarcinoma or squamous cell carcinoma, which testify of a common progenitor cell derivation, not shared by carcinoid which is never mixed with a non-neuroendocrine (NE) tumor type. Carcinoid tumors also have very few genetic abnormalities compared to SCLC and LCNEC which show the highest rate of mutations per megabase among all cancer[3,4,5] . While in many cases, SCLC and carcinoid tumors can be diagnosed on good quality tumor material with a high quality H&E stained section and in well preserved cytological samples, immunohistochemistry (IH)/neuroendocrine markers can be very helpful in diagnosing pulmonary NE tumors especially in small biopsies with crushed artefact. Endocrine morphology and neuroendocrine IH markers are both required for the diagnosis of LCNEC. The cases with one missing (endocrine morphology or NE markers) are considered as large cell carcinoma in the absence of other differentiation marker on resection specimens, and as non-small cell lung carcinoma on small samples (cytology or biopsy) Mitotic counts are still retained to differentiate typical carcinoids (less than 2 mitoses per mm[2]) from atypical carcinoids (2 to 10 per 2mm[2]) and from high grade NE tumors SCLC and LCNEC (more than 11 mitoses per 2 mm[2 ], for being more reproducible than KI-67 evaluation. The role of Ki-67 is mainly to separate the high grade SCLC (more than 50%) and LCNEC (more than 40%) from the carcinoid tumors (from 1 to 15%) especially in small biopsies with crushed and/or necrotic tumor cells. It is recommended to avoid the diagnosis of SCLC or LCNEC for tumors with less than 50% and 40% MIB1/KI67 index respectively. Data are conflicting regarding the use of KI-67 in separating typical from atypical carcinoid tumors, so it is not recommended in this setting. Careful counting of mitoses is essential as it is the most important histologic criteria for separating typical from atypical carcinoid and the carcinoids from the high grade SCLC and LCNEC. Due to recognition of the potential overlap in the morphology of LCNEC and basaloid squamous cell carcinoma, it can be helpful to confirm negative squamous markers (i.e. p40) in TTF-1 negative tumors that otherwise meet criteria for LCNEC. Many recent progress have been made on the comprehensive genomic profiles of SCLC [3,4 ], LCNEC [5] and carcinoids [6]. Although sharing NE features, these 3 tumors group show substantial and significant differences.Recent comprehensive genomic analyses have established the genomic profile of SCLC [3,6.] Their unique and remarkable characteristic is the universal bi-allelic alteration of both TP53 and RB1 gene (100% for P53 and 93% for RB1) by different alterations of each of the 4 alleles: non synonymous mutations, damaging mutations by complex genomic rearrangements. Locally clustered mutations, indicative of functional selection, occurred on CREBBP (15%) and EP300 (13%) genes, inactivating their histone acetylase functions. Notch family genes inactivating their protein functions occurred in 25% of SCLC [4]. Notch is considered as a master regulator of NE differentiation. LCNEC genomics share characteristic features with SCLC for a part of LCNEC (SCLC-like LCNEC) or with AD /SQC for another part (about 25%). Mutations pattern and frequency of combined cases imply a considerable plasticity of theses tumours which might represent an evolutionary trunk branching SCLC to NSCLC. Carcinoid is a unique example of a tumor driven entirely by chromatin modifiers and remodeling genes, which are not mutant in SCLC. In summary, 51% of carcinoid carried mutations in chromatin remodeling genes. In addition, the eukaryotic translation initiation factor (EIF1AX) was mutated in 9% of cases, genes of the E3 ubiquitin ligases system were mutated or rearranged in 18%. Altogether 73% of carcinoids have driver genes that are candidates for targeted therapy [6.] New evidence is provided that carcinoid is not an early progenitor of high grade NE tumors SCLC and LCNEC. References: 1. Travis WD, Brambilla E, Burke A, Marx A, Nicholson A. WHO Classification of the Tumours of the Lung, Pleura, Thymus and Heart. 4th Edition. Lyon: IARC Press; 2015. 2. Clinical Lung Cancer Genome Project (CLCGP), Network Genomic Medicine (NGM). A genomics-based classification of human lung tumors. Sci Transl Med. 2013;5(209):209ra153. doi:10.1126/scitranslmed.3006802. 3. Peifer M, Fernández-Cuesta L, Sos ML, et al. Integrative genome analyses identify key somatic driver mutations of small-cell lung cancer. Nat Genet. 2012;44(10):1104-1110. doi:10.1038/ng.2396. 4. George J, Lim JS, Jang SJ, et al. Comprehensive genomic profiles of small cell lung cancer. Nature. 2015;524(7563):47-53. doi:10.1038/nature14664. 5. Fernandez-Cuesta L, Peifer M, George J, et al. Genomic Characterization of Large-Cell Neuroendocrine Lung Tumors. J Thorac Oncol. 2015;10(9 - WCLC 2015 Abstracts: PDF Only):S185. doi:10.1097/01.JTO.0000473439.77589.a7. 6. Fernandez-Cuesta L, Peifer M, Lu X, et al. Frequent mutations in chromatin-remodelling genes in pulmonary carcinoids. Nat Commun. 2014;5:3518. doi:10.1038/ncomms4518.
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MA11 - Novel Approaches in SCLC and Neuroendocrine Tumors (ID 391)
- Event: WCLC 2016
- Type: Mini Oral Session
- Track: SCLC/Neuroendocrine Tumors
- Presentations: 1
- Moderators:P. Lara, A. Mohn-Staudner
- Coordinates: 12/06/2016, 14:20 - 15:50, Strauss 3
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MA11.05 - A Case-Control Study to Test the Use of ctDNA in the Early Detection of SCLC Reveals TP53 Mutations in Non-Cancer Controls (ID 4915)
14:20 - 15:50 | Author(s): E. Brambilla
- Abstract
- Presentation
Background:
Circulating-tumor DNA (ctDNA) is emerging as a key potential biomarker for post-diagnosis surveillance but it may also play a crucial role in the detection of pre-clinical cancer. Small-cell lung cancer (SCLC) is an excellent candidate for early detection given there are no successful therapeutic options for late-stage disease, and it displays universal inactivation of TP53 (Peifer and Fernandez-Cuesta et al., Nat Genet 2012; George et al., Nature 2015).
Methods:
We assessed the presence of TP53 mutations in the cell-free DNA (cfDNA) extracted from the plasma of 51 SCLC cases and 123 non-cancer controls. The results were further validated in an independent series of 102 non-cancer controls. We identified mutations using Needlestack (Delhomme et al., in preparation; https://github.com/IARCbioinfo/needlestack), a pipeline specifically designed to accurately detect variants at very low allelic fractions (AF).
Results:
We detected TP53 mutations in the cfDNA of 49% of the SCLC patients (35.7% of the stage I-II). While statistically significant in cases versus controls (p-value=6x10[-9]), TP53 mutations were also detected in the cfDNA of 11.4% of the non-cancer controls, and these results were confirmed in the replication series (10.8%). The presence of TP53 mutations in controls was not correlated with age, smoking, or alcohol-intake status. There was a statistically significant difference between the mutational patterns found in cases versus controls (p-value=0.008): within controls the fraction of nonsense, indel, or splicing mutations was lower than in cases. The median AF of the TP53 mutations detected in the five stage I-II SCLC (0.9%) was not significantly different from that found in controls (1.2%; p-value=0.64), while it differed from the median AF of stage III-IV SCLC tumors (8.2%; p-value=2x10[-6]). Finally, we sequenced the DNA extracted from the white-blood cells (WBC) of 39 cfDNA TP53-positive patients, from which material was available (19 cases and 20 controls). Four cfDNA TP53 mutations (1 case and 3 controls) were detected in the WBC DNA, with similar AFs to those found in the cfDNA. These AFs, below 11%, are consistent with a somatic origin in both cfDNA and WBC DNA.
Conclusion:
The detection of TP53 mutations in 11% of the 225 non-cancer controls suggests that somatic mutations in cfDNA among individuals without any cancer diagnosis is a common occurrence, and poses serious challenges for the development of ctDNA screening tests for the early diagnosis of cancer (Fernandez-Cuesta, Perdomo, and Avogbe et al., EBioMedicine 2016). We will discuss these results as well as follow-up analyses in retrospective and prospective series.
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MA12 - Miscellaneous Biology/Pathology (ID 476)
- Event: WCLC 2016
- Type: Mini Oral Session
- Track: Biology/Pathology
- Presentations: 1
- Moderators:B. Dome, W.D. Travis
- Coordinates: 12/06/2016, 14:20 - 15:50, Schubert 1
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MA12.03 - Discussant for MA12.01, MA12.02 (ID 7010)
14:20 - 15:50 | Author(s): E. Brambilla
- Abstract
- Presentation
Abstract not provided
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