- Email: [email protected]
Investigation of the mechanisms of resistance to osimertinib in patients with T790M-associated NSCLC
abstracts 51P
LIMCH1-related genes demonstrate different invasive potential of morphological structures of breast cancer
L. Tashireva1, T. Gerashchenko2, V. Alifanov1, V. Perelmuter1, N. Cherdyntseva2 General and Molecular Pathology Department, Cancer Research Institute - Tomsk National Research Medical Center, Tomsk, Russian Federation, 2Molecular Oncology and Immunology, Cancer Research Institute - Tomsk National Research Medical Center, Tomsk, Russian Federation
1
53P
ZNF518B as a transcriptional factor involved in colorectal cancer progression through the epithelial to mesenchymal transition
Riffo-Campos2, F. Papaccio1, F.G. Valiente1, N. Tarazona1, V. Gambardella1, A. opez-Rodas3, A. Cervantes1, L. Franco3, J. Castillo3 M. Cabeza-Segura1, G. L 1 Oncology, INCLIVA Instituto de Investigaci on Sanitaria, Valencia, Spain, 2Computer Science, University of La Frontera, Temuco, Chile, 3Bioquımica, Universidad de Valencia, Valencia, Spain Background: Colorectal cancer (CRC) represents a relevant public health problem. The identification of new markers involved in the mechanisms of invasiveness represents a priority in order to better understand cancer development and generate new therapeutic targets. Recently, our group demonstrated overexpression of ZNF518B gene, which encodes an unknown zinc finger transcription factor, in CRC. A transcriptome-wide gene expression profile revealed its implication in different biological processes related to the progression of CRC, especially in the epithelial to mesenchymal transition (EMT). Methods: To study the biological processes regulated by ZNF518B, we performed a ClariomS Array (Affimetrix) by silencing this gene with a specific siRNA in DLD1 and HCT116 CRC cell lines. All data were analysed using the TAC (Transcriptomic Analysis ConsoleTM) software. The array validation was performed by RT-qPCR. To explore the in vitro mechanisms related to EMT, an overexpression of ZNF518B was performed in RKO, cell line that does not express the gene. Invasion and migration assays were done using transwells and we also test adhesion capacity to type I collagen plates. Results: The analysis of ClariomS showed, after the silencing of the transcription factor ZNF518B, an alteration of 64 genes commonly modified in the DLD1 and HCT116 cell lines. These differentially expressed genes are involved in different biological processes involved in cellular focal adhesion, in the EMT process and in cell cycle. On the other hand, some proliferation pathways are also altered like MAPK, RAS, WNT and VEGFR2 signalling pathways. Through in vitro analysis, we showed that the increased expression of ZNF518B enhanced cell migration, invasion and adhesion to type I collagen. These results in vitro support the altered gene expression profile after ZNF518B silencing observed in the ClariomS array, suggesting that this gene has an important role in the progression of CRC throughout EMT. Conclusions: The transcription factor ZNF518B could be involved in the progression of tumorigenesis by inducing EMT in CRC.The transcription factor ZNF518B could be involved in the progression of tumorigenesis by inducing EMT in CRC. Legal entity responsible for the study: INCLIVA, Biomedical Research Institute.
vii16 | Molecular Analysis for Personalised Therapy
Funding: Instituto de Salud Carlos III, Rio Hortega Contract, ESMO Translational Research Fellowship Programme. Disclosure: A. Cervantes: Advisory / Consultancy, Speaker Bureau / Expert testimony, Research grant / Funding (institution): Merck Serono; Advisory / Consultancy, Speaker Bureau / Expert testimony, Research grant / Funding (institution): Roche; Advisory / Consultancy, Research grant / Funding (institution): Beigene; Advisory / Consultancy, Research grant / Funding (institution): Bayer; Advisory / Consultancy, Speaker Bureau / Expert testimony, Research grant / Funding (institution): Servier; Advisory / Consultancy, Research grant / Funding (institution): Lilly; Advisory / Consultancy, Research grant / Funding (institution): Novartis; Advisory / Consultancy, Research grant / Funding (institution): Takeda; Advisory / Consultancy, Research grant / Funding (institution): Astellas; Advisory / Consultancy: Pierre Fabre; Research grant / Funding (self): Genentech; Research grant / Funding (institution): Fibrogene; Research grant / Funding (institution): Amcure; Research grant / Funding (institution): Sierra Oncology; Research grant / Funding (institution): AstraZeneca; Research grant / Funding (institution): MedImmune; Research grant / Funding (institution): BMS; Research grant / Funding (institution): MSD; Speaker Bureau / Expert testimony: Amgen; Speaker Bureau / Expert testimony: Fundation Medicine. All other authors have declared no conflicts of interest.
54P
Investigation of the mechanisms of resistance to osimertinib in patients with T790M-associated NSCLC
M. Stepanova, F. Moiseenko, A. Zhabina, V. Klimenko, N. Rysev, A. Myslik, E. Artemieva, K. Shelekhova, A. Bogdanov, N. Volkov, V. Moiseenko St. Petersburg Clinical Research Center of Specialized Types of Care (Oncology), St. Petersburg, Russian Federation Background: Osimertinib (Osi) is a 3rd generation TKI that crosses the blood-brain barrier (BBB), which has been shown to be effective in both the second and first line of treatment in patients with NSCLC with an EGFR mutation. In use of Osi in patients with T790M associated progression during therapy with earlier TKIs, the median PFS was 10.1 months. The duration of treatment may vary significantly for each patient. The study of the mechanisms of resistance to 3rd generation TKIs is the subject of clinical studies, among the most common variants of resistance - the appearance of the gate-keeper mutation C797S - in 7% of cases. Methods: It is prospective, single-center study we examined the appearance of resistance in patients receiving Osi treatment.The search for the C797S mutation was carried out using an original diagnostic platform based on real-time PCR, using the BioRad Real-Time CFX96 amplifier. Primers and fluorescent probes were synthesized by Applied Biosystems compony. The study included patients with progression during therapy with Osi. Before treatment, and then every 2 months until progression of the disease appease, blood sampling was performed to conduct a qualitative assessment of ctDNA (ex19del, L858R, T790M, C797S). Results: From August 2016 to August 2019, in 12/22 patients, progression of the disease during the Osi. Emong them, 66.7% (7/12) are women, 33.3% (4/12) are men. The average age is 64.6 years (55-80). 1/12 smoked for over 30 years. The molecular genetic profile in 58.3% (7/12) of the patients is ex19del, 33.3% (4/12) of L858R, 8.4% (1/12) is a combination of rare mutations G719S þ S768I. The median PFS during the Osi treatment was 9.6 months. (0.7 - 18.4).The analysis of ctDNA was performed in 11/12 patients: an activated mutation was detected in all cases, 1/12 recorded the mutation C796S (NGS), T790M and C797S (cis/ trans) were not detected.Histological material was taken from 4/12 patients, 2 of which showed the transformation of the tumor into small cell lung cancer. Conclusions: The investigation of appearence the resistance mechanisms in the treatment with osimertinib did not reveal the EGFR C797S mutation, which may be associated with a small number of patients included in the study.Grant supported RUSSCO / RakFond 2018-01-YS-ECI. Legal entity responsible for the study: St. Petersburg Clinical Research Center of Specialized Types of Care (Oncology). Funding: RakFond (Russia). Disclosure: All authors have declared no conflicts of interest.
55P
NGS-based multi-gene panel analysis in early-onset colorectal cancer patients
G. Zhunussova1, G. Afonin2, S. Abdikerim1, A. Jumanov2, A. Perfilyeva1, D. Kaidarova2, L. Djansugurova1 1 Institute of General Genetics and Cytology, Almaty, Kazakhstan, 2Kazakh Institute of Oncology and Radiology, Asfendiyarov Kazakh National Medical University, Almaty, Kazakhstan Background: Colorectal cancer (CRC) keeps an upward incidence and rejuvenation trend in Kazakhstan, as well as worldwide, which is the motive for revising traditional risk factors and searching for cause-effective relationships at the genome level. Genetic nature of early-onset CRC is still unclear. The aim of this study was to determine the spectrum of cancer-related gene mutations among early-onset CRC patients. Methods: The study included 125 CRC patients aged between 17 and 50 from Kazakhstan. DNAs were extracted from blood using the GeneJET Purification Kit. Next-generation sequencing (NGS) was done on the MiSeq using the TruSightCancer Kit. NGS data were analyzed by the MiSeq Reporter and Variant Studio. SIFT and
Volume 30 | Supplement 7 | November 2019
Downloaded from https://academic.oup.com/annonc/article-abstract/30/Supplement_7/mdz413.058/5613296 by guest on 23 December 2019
Background: Tumor cell invasion is an integral component of breast cancer progression. In cell motility, including tumor cells, involves the actin cytoskeleton, which is a complex system with many regulatory points. The new tumor supressor protein LIMCH1, which has an actin binding domain, is a promising molecular target to suppress invasion. The presented work is aimed at revealing features of LIMCH1 expression in cells that demonstrate various forms of invasion in breast cancer. Methods: The study included 37 patients with invasive breast carcinoma of a non-specific type (all molecular types). A list of genes was formed and molecular networks associated with LIMCH1 signaling were constructed. Using the previously obtained unique data of transcriptomic microarray profiling of various morphological structures of invasive breast carcinoma of a non-specific type (GSE80754), differentially expressed genes associated with LIMCH1 in the morphological structures were annotated. Results: Tumor cells can move collectively or individually. The structural diversity of the infiltrative component of invasive breast carcinima represent an attractive model for investigation of tumor cell invasion. The solid and trabecular structures, may be considered as a morphological manifestation of collective cell invasion. Discrete groups and single tumor cells, are an example of individual cell invasion. Analyze of expression LIMCH1-related genes in different morphological structures showed that proinvasive XPO1 gene downregulated in trabecular structures. In single tumor cells expression of ODF2 gene that had stimulation activity for prolifiration were supressed comparing with trabecular structures. SH3KBP1 gene in solid structures is upregulated comparing other structures. Well known that SH3KBP1 revents epidermal growth factor receptor degradation. Also, HuR gene expression were upregulated in solid and trabecular structures comparing single tumor cells. Conclusions: The obtained data showed that the morphological structures of invasive breast carcinoma have significant functional heterogeneity associated with the manifestations of invasion regulated by the LIMCH1 protein. The study was supported by RFBR (#18-515-16001). Legal entity responsible for the study: Tomsk National Research Medical Center. Funding: Russian Foundation for Basic Research (project #18-515-16001). Disclosure: All authors have declared no conflicts of interest.
Annals of Oncology
LIMCH1-related genes demonstrate different invasive potential of morphological structures of breast cancer
L. Tashireva1, T. Gerashchenko2, V. Alifanov1, V. Perelmuter1, N. Cherdyntseva2 General and Molecular Pathology Department, Cancer Research Institute - Tomsk National Research Medical Center, Tomsk, Russian Federation, 2Molecular Oncology and Immunology, Cancer Research Institute - Tomsk National Research Medical Center, Tomsk, Russian Federation
1
53P
ZNF518B as a transcriptional factor involved in colorectal cancer progression through the epithelial to mesenchymal transition
Riffo-Campos2, F. Papaccio1, F.G. Valiente1, N. Tarazona1, V. Gambardella1, A. opez-Rodas3, A. Cervantes1, L. Franco3, J. Castillo3 M. Cabeza-Segura1, G. L 1 Oncology, INCLIVA Instituto de Investigaci on Sanitaria, Valencia, Spain, 2Computer Science, University of La Frontera, Temuco, Chile, 3Bioquımica, Universidad de Valencia, Valencia, Spain Background: Colorectal cancer (CRC) represents a relevant public health problem. The identification of new markers involved in the mechanisms of invasiveness represents a priority in order to better understand cancer development and generate new therapeutic targets. Recently, our group demonstrated overexpression of ZNF518B gene, which encodes an unknown zinc finger transcription factor, in CRC. A transcriptome-wide gene expression profile revealed its implication in different biological processes related to the progression of CRC, especially in the epithelial to mesenchymal transition (EMT). Methods: To study the biological processes regulated by ZNF518B, we performed a ClariomS Array (Affimetrix) by silencing this gene with a specific siRNA in DLD1 and HCT116 CRC cell lines. All data were analysed using the TAC (Transcriptomic Analysis ConsoleTM) software. The array validation was performed by RT-qPCR. To explore the in vitro mechanisms related to EMT, an overexpression of ZNF518B was performed in RKO, cell line that does not express the gene. Invasion and migration assays were done using transwells and we also test adhesion capacity to type I collagen plates. Results: The analysis of ClariomS showed, after the silencing of the transcription factor ZNF518B, an alteration of 64 genes commonly modified in the DLD1 and HCT116 cell lines. These differentially expressed genes are involved in different biological processes involved in cellular focal adhesion, in the EMT process and in cell cycle. On the other hand, some proliferation pathways are also altered like MAPK, RAS, WNT and VEGFR2 signalling pathways. Through in vitro analysis, we showed that the increased expression of ZNF518B enhanced cell migration, invasion and adhesion to type I collagen. These results in vitro support the altered gene expression profile after ZNF518B silencing observed in the ClariomS array, suggesting that this gene has an important role in the progression of CRC throughout EMT. Conclusions: The transcription factor ZNF518B could be involved in the progression of tumorigenesis by inducing EMT in CRC.The transcription factor ZNF518B could be involved in the progression of tumorigenesis by inducing EMT in CRC. Legal entity responsible for the study: INCLIVA, Biomedical Research Institute.
vii16 | Molecular Analysis for Personalised Therapy
Funding: Instituto de Salud Carlos III, Rio Hortega Contract, ESMO Translational Research Fellowship Programme. Disclosure: A. Cervantes: Advisory / Consultancy, Speaker Bureau / Expert testimony, Research grant / Funding (institution): Merck Serono; Advisory / Consultancy, Speaker Bureau / Expert testimony, Research grant / Funding (institution): Roche; Advisory / Consultancy, Research grant / Funding (institution): Beigene; Advisory / Consultancy, Research grant / Funding (institution): Bayer; Advisory / Consultancy, Speaker Bureau / Expert testimony, Research grant / Funding (institution): Servier; Advisory / Consultancy, Research grant / Funding (institution): Lilly; Advisory / Consultancy, Research grant / Funding (institution): Novartis; Advisory / Consultancy, Research grant / Funding (institution): Takeda; Advisory / Consultancy, Research grant / Funding (institution): Astellas; Advisory / Consultancy: Pierre Fabre; Research grant / Funding (self): Genentech; Research grant / Funding (institution): Fibrogene; Research grant / Funding (institution): Amcure; Research grant / Funding (institution): Sierra Oncology; Research grant / Funding (institution): AstraZeneca; Research grant / Funding (institution): MedImmune; Research grant / Funding (institution): BMS; Research grant / Funding (institution): MSD; Speaker Bureau / Expert testimony: Amgen; Speaker Bureau / Expert testimony: Fundation Medicine. All other authors have declared no conflicts of interest.
54P
Investigation of the mechanisms of resistance to osimertinib in patients with T790M-associated NSCLC
M. Stepanova, F. Moiseenko, A. Zhabina, V. Klimenko, N. Rysev, A. Myslik, E. Artemieva, K. Shelekhova, A. Bogdanov, N. Volkov, V. Moiseenko St. Petersburg Clinical Research Center of Specialized Types of Care (Oncology), St. Petersburg, Russian Federation Background: Osimertinib (Osi) is a 3rd generation TKI that crosses the blood-brain barrier (BBB), which has been shown to be effective in both the second and first line of treatment in patients with NSCLC with an EGFR mutation. In use of Osi in patients with T790M associated progression during therapy with earlier TKIs, the median PFS was 10.1 months. The duration of treatment may vary significantly for each patient. The study of the mechanisms of resistance to 3rd generation TKIs is the subject of clinical studies, among the most common variants of resistance - the appearance of the gate-keeper mutation C797S - in 7% of cases. Methods: It is prospective, single-center study we examined the appearance of resistance in patients receiving Osi treatment.The search for the C797S mutation was carried out using an original diagnostic platform based on real-time PCR, using the BioRad Real-Time CFX96 amplifier. Primers and fluorescent probes were synthesized by Applied Biosystems compony. The study included patients with progression during therapy with Osi. Before treatment, and then every 2 months until progression of the disease appease, blood sampling was performed to conduct a qualitative assessment of ctDNA (ex19del, L858R, T790M, C797S). Results: From August 2016 to August 2019, in 12/22 patients, progression of the disease during the Osi. Emong them, 66.7% (7/12) are women, 33.3% (4/12) are men. The average age is 64.6 years (55-80). 1/12 smoked for over 30 years. The molecular genetic profile in 58.3% (7/12) of the patients is ex19del, 33.3% (4/12) of L858R, 8.4% (1/12) is a combination of rare mutations G719S þ S768I. The median PFS during the Osi treatment was 9.6 months. (0.7 - 18.4).The analysis of ctDNA was performed in 11/12 patients: an activated mutation was detected in all cases, 1/12 recorded the mutation C796S (NGS), T790M and C797S (cis/ trans) were not detected.Histological material was taken from 4/12 patients, 2 of which showed the transformation of the tumor into small cell lung cancer. Conclusions: The investigation of appearence the resistance mechanisms in the treatment with osimertinib did not reveal the EGFR C797S mutation, which may be associated with a small number of patients included in the study.Grant supported RUSSCO / RakFond 2018-01-YS-ECI. Legal entity responsible for the study: St. Petersburg Clinical Research Center of Specialized Types of Care (Oncology). Funding: RakFond (Russia). Disclosure: All authors have declared no conflicts of interest.
55P
NGS-based multi-gene panel analysis in early-onset colorectal cancer patients
G. Zhunussova1, G. Afonin2, S. Abdikerim1, A. Jumanov2, A. Perfilyeva1, D. Kaidarova2, L. Djansugurova1 1 Institute of General Genetics and Cytology, Almaty, Kazakhstan, 2Kazakh Institute of Oncology and Radiology, Asfendiyarov Kazakh National Medical University, Almaty, Kazakhstan Background: Colorectal cancer (CRC) keeps an upward incidence and rejuvenation trend in Kazakhstan, as well as worldwide, which is the motive for revising traditional risk factors and searching for cause-effective relationships at the genome level. Genetic nature of early-onset CRC is still unclear. The aim of this study was to determine the spectrum of cancer-related gene mutations among early-onset CRC patients. Methods: The study included 125 CRC patients aged between 17 and 50 from Kazakhstan. DNAs were extracted from blood using the GeneJET Purification Kit. Next-generation sequencing (NGS) was done on the MiSeq using the TruSightCancer Kit. NGS data were analyzed by the MiSeq Reporter and Variant Studio. SIFT and
Volume 30 | Supplement 7 | November 2019
Downloaded from https://academic.oup.com/annonc/article-abstract/30/Supplement_7/mdz413.058/5613296 by guest on 23 December 2019
Background: Tumor cell invasion is an integral component of breast cancer progression. In cell motility, including tumor cells, involves the actin cytoskeleton, which is a complex system with many regulatory points. The new tumor supressor protein LIMCH1, which has an actin binding domain, is a promising molecular target to suppress invasion. The presented work is aimed at revealing features of LIMCH1 expression in cells that demonstrate various forms of invasion in breast cancer. Methods: The study included 37 patients with invasive breast carcinoma of a non-specific type (all molecular types). A list of genes was formed and molecular networks associated with LIMCH1 signaling were constructed. Using the previously obtained unique data of transcriptomic microarray profiling of various morphological structures of invasive breast carcinoma of a non-specific type (GSE80754), differentially expressed genes associated with LIMCH1 in the morphological structures were annotated. Results: Tumor cells can move collectively or individually. The structural diversity of the infiltrative component of invasive breast carcinima represent an attractive model for investigation of tumor cell invasion. The solid and trabecular structures, may be considered as a morphological manifestation of collective cell invasion. Discrete groups and single tumor cells, are an example of individual cell invasion. Analyze of expression LIMCH1-related genes in different morphological structures showed that proinvasive XPO1 gene downregulated in trabecular structures. In single tumor cells expression of ODF2 gene that had stimulation activity for prolifiration were supressed comparing with trabecular structures. SH3KBP1 gene in solid structures is upregulated comparing other structures. Well known that SH3KBP1 revents epidermal growth factor receptor degradation. Also, HuR gene expression were upregulated in solid and trabecular structures comparing single tumor cells. Conclusions: The obtained data showed that the morphological structures of invasive breast carcinoma have significant functional heterogeneity associated with the manifestations of invasion regulated by the LIMCH1 protein. The study was supported by RFBR (#18-515-16001). Legal entity responsible for the study: Tomsk National Research Medical Center. Funding: Russian Foundation for Basic Research (project #18-515-16001). Disclosure: All authors have declared no conflicts of interest.
Annals of Oncology