- Email: [email protected]
The properties of mitochondria in glioma stem cells
abstracts 44P
Association of metastatic pattern and molecular status in metastatic lung non-small cell lung cancer adenocarcinomas
A. Dormieux1, L. Mezquita2, P-H. Cournede3, L. Lacroix2, E. Rouleau2, J. Adam4, F. Facchinetti2, F. Aboubakar4, M-V. Bluthgen2, C. Naltet2, P. Lavaud5, A. Gazzah2, C. Le Pechoux6, C. Balleyguier1, D. Planchard2, B. Besse7, C. Caramella1 1 Radiology, Institut Gustave Roussy, Villejuif, France, 2Institut Gustave Roussy, Villejuif, France, 3MICS Laboratory, CentraleSupe´lec, Gif-sur-Yvette, France, 4Pathology, Institut Gustave Roussy, Villejuif, France, 5Medical Oncology Departement, Institut Gustave Roussy, Villejuif, France, 6Radiation Oncology, Institut Gustave Roussy, Villejuif, France, 7 Oncology, Institut Gustave Roussy, Villejuif, France
45P
Circulating tumour associated cells in esophageal cancers are resistance educated per previous chemo treatments
S. Limaye1, T. Crook2, A. Ranade3, D. Patil4, D. Akolkar4, V. Datta4, S. Schuster5, R. Page6, C. Sims4, R. Patil4, A. Srinivasan4, S. Khan4, S. Patil4, V. Mhase4, S. Apurwa4, R. Datar4 1 Medical Oncology, Kokilaben Dhirubhai Ambani Hospital, Mumbai, India, 2St Luke’s Cancer Centre, Royal Surrey County Hospital, Guildford, UK, 3Avinash Cancer Clinic, Pune, India, 4Research and Innovations Department, Datar Cancer Genetics, Nashik, India, 5Datar Cancer Genetics Europe GmbH, Bayreuth, Germany, 6Biomedical Engineering, Worcester Polytechnic Institute, Worcester, MA, USA Background: Innate and acquired chemoresistance to anticancer therapies are a wellknown phenomenon in Esophageal Squamous Cell Carcinomas (ESCC). There are presently no viable approaches for real-time monitoring of resistance in ESCC. We used a novel method for chemo-interrogation (CI) by harvesting sufficient number of Circulating-Tumor Associated Cells (C-TACs) which are defined as apoptosis-resistant cells of tumorigenic origin and are positive for Epithelial Cell Adhesion Molecule (EpCAM) and pan-cytokeratins (pan-CK) irrespective of CD45 status. Methods: Peripheral blood was collected from 80 patients with confirmed diagnosis of ESCC, among whom 52 were recently diagnosed therapy-naı¨ve, while 28 were pretreated patients. Peripheral blood mononuclear cells (PBMCs) were harvested by centrifugation and treated with commercially available stabilizing agents, by a proprietary protocol, to stabilize apoptosis resistant C-TACs. Surviving C-TACs were confirmed by immunostaining for EpCAM, pan-CK and CD45. Harvested C-TACs were treated in vitro with a panel of conventional cytotoxic anticancer agents and fraction of surviving cells were estimated to determine resistance profiles. Results: Among the 52 cases of recently diagnosed therapy naı¨ve ESCC, innate chemoresistance was observed towards platinum agents in 40.8% samples (unique patientdrug combinations), taxanes in 34.6% samples, topoisomerase inhibitors in 42.9% samples and antimetabolites in 34.6% samples. Among the 28 cases of previously treated ESCC, resistance towards previously administered systemic agents was observed in 87% of all samples, which included resistance towards platinum agents in 87.5% samples, taxanes in 82.1% samples, topoisomerase inhibitors in 100% samples and antimetabolites in 85.7% samples, respectively. Conclusions: We show for the first time that sufficient C-TACs can be harvested for meaningful CI in newly diagnosed treatment naı¨ve ESCC as well as refractory ESCCs. Post-treatment chemoresistance being an order of magnitude higher than the untreated cohort, indicates that C-TACs in ESCC are resistance-educated by previous treatments and can guide treatment strategy in ESCC.
vii14 | Molecular Analysis for Personalised Therapy
Legal entity responsible for the study: The authors. Funding: Datar Cancer Genetics Limited. Disclosure: D. Patil: Full / Part-time employment: Datar Cancer Genetics Limited. D. Akolkar: Full / Part-time employment: Datar Cancer Genetics Limited. V. Datta: Full / Part-time employment: Datar Cancer Genetics Limited. S. Schuster: Full / Part-time employment: Datar Cancer Genetics Limited. C. Sims: Full / Part-time employment: Datar Cancer Genetics Limited. R. Patil: Full / Parttime employment: Datar Cancer Genetics Limited. A. Srinivasan: Full / Part-time employment: Datar Cancer Genetics Limited. S. Khan: Full / Part-time employment: Datar Cancer Genetics Limited. S. Patil: Full / Part-time employment: Datar Cancer Genetics Limited. V. Mhase: Full / Part-time employment: Datar Cancer Genetics Limited. S. Apurwa: Full / Part-time employment: Datar Cancer Genetics Limited. R. Datar: Shareholder / Stockholder / Stock options, Licensing / Royalties, Officer / Board of Directors: Datar Cancer Genetics Limited. All other authors have declared no conflicts of interest.
46P
Prevalence of homologous recombination deficiency (HRD)-related signatures indicates that a wider range of prostate cancer patients may benefit from PARP-inhibitor therapy
Z. Sztupinszki1, M. Diossy1, M. Krzystanek1, J. Borcsok1, M. Pomerantz2, V. Tisza3, S. Spisak2, O. Rusz4, I. Csabai5, M. Freedman2, Z. Szallasi3 1 Translational Cancer Genomics, The Danish Cancer Society, Copenhagen, Denmark, 2 Department of Medical Oncology, Dana-Farber Cancer Institute, Boston, MA, USA, 3 Computational Health Informatics Program, Boston Children’s Hospital, Boston, MA, USA, 4Semmelweis University, Budapest, Hungary, 5Department of Physics of Complex Systems, Department of Physics of Complex Systems, Budapest, Hungary Background: Prostate cancers with mutations in genes involved in homologous recombination (HR), most commonly BRCA2, respond favorably to PARP-inhibition and platinum-based chemotherapy. Currently targeted sequencing of key HR genes serves as an indicator of homologous recombination deficiency (HRD) in prostate cancer. It is well known, however, that other mechanisms, such as suppression of expression, rare mutations of HR-related genes can also cause HRD. Methods: HRD levels can be estimated using various mutational, genomic signatures derived from next-generation sequencing data. We used this approach to determine whether prostate cancer cases display clear signs of HRD in somatic tumor biopsies. Whole genome (n ¼ 311) and whole exome sequencing data (n ¼ 498) of both primary and metastatic prostate adenocarcinomas (PRAD) were analyzed. Results: Known BRCA-deficient samples showed robust signs of HR-deficiency associated mutational signatures. HRD-patterns were also detected in a subset of patients who did not harbor germline or somatic mutations in BRCA1/2 or other HR-related genes. The signs HRD present low intratumor heterogeneity and in most cases, the metastasis’s HRD-status is similar to the primary tumor’s. Patients with HRD signatures had a significantly worse prognosis than patients without signs of HRD. Conclusions: Based on the HRD associated mutational and genomic scar signatures, 58 % of prostate cancer cases may be good candidates for PARP-inhibitor treatment (including those with BRCA1/2 mutations). The prioritization of these patients for clinical trials, extending PARP-inhibitor therapy to all cases with HRD associated mutational signatures, even to those without BRCA1/2 mutations, will likely increase the efficacy of this treatment. Legal entity responsible for the study: The authors. Funding: Research and Technology Innovation Fund (KTIA_NAP_13-2014-0021 to Z.S.); Breast Cancer Research Foundation (BCRF-17-156 to Z.S.) and the Novo Nordisk Foundation Interdisciplinary Synergy Programme Grant (NNF15OC0016584 to Z.S.), Department of Defense through the Prostate Cancer Research Program (award number is W81XWH-18-2-0056) to Z.S. and M.F. The Danish Cancer Society grant (R90-A6213 to MK). Z.S. and J.B. was supported by Velux Foundation 00018310 grant. Disclosure: Z. Szallasi: Licensing / Royalties: Inventor on a patent used in the myChoice HRD assay: Myriad. All other authors have declared no conflicts of interest.
47P
The properties of mitochondria in glioma stem cells
N. Hong, E-J. Kim, H. Kim Department of Biotechnology, Korea University, Seoul, Republic of Korea Background: Mitochondria have a key role in cell survival, signaling, metabolism, and homeostasis. However, whether their biological properties affected by differentiation status of cells are not well known. This study aimed to compare structural and functional differences of mitochondria between differentiated cancer cells and undifferentiated cancer stem cells. Methods: Glioma stem cells (GSCs) were cultured in stem cell culture conditions to maintain their stem cell properties or serum culture conditions to induce their differentiation. Characteristics of GSCs are analyzed through sphere-forming ability and expression of stem cell/differentiation markers. Mitochondrial biogenesis and structure was examined by transmission electron microscopy analysis. Bioinformatics analysis, qRT-PCR, glucose uptake assay and lactate production assays were applied to reveal metabolic difference. Results: GSCs grown in serum culture conditions showed lower stem cell markers expression and higher differentiation markers compared to GSCs grown in stem cell culture conditions. In the serum culture conditions, biogenesis of mitochondria was
Volume 30 | Supplement 7 | November 2019
Downloaded from https://academic.oup.com/annonc/article-abstract/30/Supplement_7/mdz413.052/5613350 by guest on 31 December 2019
Background: The recent identification of molecular alterations in some lung adenocarcinomas has led to the emergence of effective targeted therapies thus drastically improving their prognosis.The aim of our study was to investigate the association between driver oncogene alterations and metastatic patterns on imaging assessment, in a large cohort of metastatic lung adenocarcinoma patients. Methods: From January 2010 to May 2017, 656 patients with stage IV lung adenocarcinoma with molecular analysis were studied retrospectively including 135 EGFR, 81 ALK, 47 BRAF, 141 KRAS, and 146 negative tumors for these 4 mutations (4N). After review of the complete imaging report by two radiologists (junior and senior) to identify metastatic sites, univariate correlation analyzes were performed. Results: We found differences in metastatic tropism depending on the molecular alteration type when compared to the non-mutated 4N group: in the EGFR group, pleural metastases were more frequent (32% versus 20%; p ¼ 0.021), and adrenal and node metastases less common (6% versus 23%; p < 0.001 and 11% versus 23%; p ¼ 0.011). In the ALK group, there were more brain and lung metastases (respectively 42% versus 29%; p ¼ 0.043 and 37% versus 24%; p ¼ 0.037). In the BRAF group, pleural and pericardial metastases were more common (respectively 47% versus 20%; p < 0.001 and 11% versus 3%; p ¼ 0.04) and bone metastases were rarer (21% versus 42%; p ¼ 0.011). Lymphangitis was more frequent in EGFR, ALK and BRAF groups (respectively 6%, 7% and 15% versus 1%); p ¼ 0,016, p ¼ 0,009 and p < 0,001). Conclusions: The application of these correlations between molecular status and metastatic tropism in clinical practice may lead to earlier and more accurate identification of patients for targeted therapy, savings in iterative biopsies, as well as improved and personalized imaging interpretation. Legal entity responsible for the study: Alison Dormieux. Funding: Has not received any funding. Disclosure: All authors have declared no conflicts of interest.
Annals of Oncology
abstracts
Annals of Oncology
48P
PLSCR1 and XKR8: New markers for low-grade glioma progression and outcome
K.V. Havrysh1, M. Bogdanov2, R. Kiyamova1 Research Laboratory "Biomarker" IFMB, Kazan Federal University, Kazan, Russian Federation, 2Department of Biochemistry and Molecular Biology, Center for Membrane Biology, University of Texas McGovern Medical School at Houston, TX, USA
1
Background: One of the most frequently diagnosed oncological diseases among young persons is Low-grade glioma (LGG). More than 80% of LGGs progress to higher-grade tumors always lethal to a patient. LGG is an extremely heterogeneous group of primary glial brain tumors with a quite variable rate of recurrence, survival, and response to treatment. Thus a study of new biomarkers associated with disease progression and patient outcome is useful to improve patient management. Genes involved in key cellular processes, e.g., apoptosis, are promising prognostic and predictive cancer markers. Phosphatidylserine (PS) plays an essential role in apoptosis signaling. Several studies suggested PS is dysregulated in the tumor microenvironment and offends the development of anti-cancer immune response. The scramblases PLSCR1 and XKR8 allegedly could be associated with the progression and outcome of LGG since they could be involved in the development of immunity via regulation of PS activity by its bidirectional translocation in the plasma membrane. Thus this study was aimed to evaluate associations of PLSCR1 and XKR8 with disease-free and overall survival of LGG patients. Methods: Gene expression values and clinical information of 530 LGG patients were downloaded from TCGA Low-Grade Glioma study using the public resource CBioPortal. According to gene expression values (RNA Seq V2 RSEM) patients were divided into two groups: with downregulated (< mean) or upregulated (> mean) PLSCR1 or XKR8 gene. The Kaplan-Meier procedure and Log-Rank test were used for survival analysis. All calculations were performed in the integrated development environment - RStudio. Results: It was shown the LGG patients with upregulated PLSCR1 (p ¼ 2.54e-14) and XKR8 (p ¼ 7.33e-15) gene had worse disease prognosis than other patients. Similar associations were seen in the study of LGG progression, where patients with upregulated PLSCR1 (p ¼ 7.75e-07) and XKR8 (p ¼ 5.59e-10) gene had a higher risk of cancer recurrence. Conclusions: Consequently, PLSCR and XKR8 genes could be considered as new prognostic biomarkers of LGG progression and outcome. Legal entity responsible for the study: Research Laboratory "Biomarker". Funding: The work was performed according to the Russian Government Program of Competitive Growth of KFU. Disclosure: All authors have declared no conflicts of interest.
49P
An exceptional response to immunotherapy doublet in combined hepatocellular carcinoma-cholangiocarcinoma
E. Tahover Oncology, Shaare Zedek Medical Center, Jerusalem, Israel Background: We present here a case of a 67-year-old male. He presented in May 2018 with severe abdominal pain and weight loss of 25%. Alpha-feto protein (AFP) and Carbohydrate antigen 19-9 (CA19-9) were within normal limits, and CA125 was 5 times upper normal limit (ULN). Computed tomography showed liver masses and enlarged retroperitoneal lymph nodes. Biopsy from a liver mass showed Combined Hepatocellular Carcinoma (HCC)-Cholangiocarcinoma (CC) (CHC). This a rare tumor, with an incidence of less than 10% of primary liver tumors. In whole exome analysis, mutations suggested the bipotent cell origin and stem cell origin. There are no guidelines or randomized trials regarding treatment. In an analysis of 36 patients who were treated with chemotherapy, the progression-free survival was 2.8 months, with an overall response rate of 5.6%. Targeted agents had minimal effect on survival.
Volume 30 | Supplement 7 | November 2019
Methods: In our case, extensive genomic, transcriptomic and proteomic testing was performed. No genomic alterations were identified, tumor mutational burden was low and microsatellite status was stable. 7 of 9 immune checkpoint genes were overexpressed. A variant in CDK12 was noted, which was shown to be associated with elevated neoantigen burden and may predict benefit from immune checkpoint therapy. Results:
Table: 49P Blood tests of case
Alk Phos GGT Ca125
Before immunotherapy
After 11 months of immunotherapy
Normal range
456 588 210
83 72 21
38-150 15-73 2-35
The patient began immunotherapy with ipilimumab and nivolumab followed by nivolumab, which he is continuing. The only side effects were hypothyroidism and Addison’s disease which are being treated. His clinical response was dramatic, he regained all the lost weight, and discontinued high dose opiate treatment. ECOG performance status improved from 3 to 0. Repeated PET-CT showed near complete response, ca125 decreased by 90% and liver function tests normalized. Conclusions: We present here an exceptional case of a rare tumor, where the patient had a clinical, laboratory and radiological response and a significant improvement in quality of life, suggesting that these tumors are sensitive to immunotherapy. No published cases of this tumor have yet been treated with immunotherapy doublet. Legal entity responsible for the study: The author. Funding: Has not received any funding. Disclosure: The author has declared no conflicts of interest.
50P
Assessing melanoma BRAF status through ddPCR of cfDNA
L. Passby1, S. Silva1, I. Brock1, G. Wells1, A. Cox1, S. Danson2 Department of Oncology and Metabolism, University of Sheffield Medical School, Sheffield, UK, 2Department of Oncology and Metabolism and Sheffield Experimental Cancer Medicine Centre, University of Sheffield Medical School, Sheffield, UK
1
Background: Targeted therapy has revolutionised the systemic treatment of melanoma. Patients with stage III/IV melanoma with a mutation in BRAF are treated with BRAF inhibitors, with BRAF mutation status being assessed by tissue analysis. Cell-free DNA (cfDNA) released from tumours provides a method for detecting BRAF mutations. This project analysed BRAF status in cfDNA through digital droplet PCR (ddPCR). We aimed to assess the relationship between cfDNA BRAF status and disease relapse and progression. Methods: Plasma from 100 patients with active or recently resected melanoma was obtained during previous work. 85 samples had cfDNA extracted. Tissue BRAF status was known for 57 samples. cfDNA was extracted from 1-2ml plasma with the QIAamp R ) following manufacturer protocol, eluting circulating nucleic acid kit (QIAGENV cfDNA into 100mL. cfDNA was quantified with SYBR green quantitative real-time PCR (Life Technologies), based on an 87bp GAPDH gene amplicon. ddPCRTM was performed using the Bio-Rad QX200 Droplet GeneratorTM and Droplet Reader as per manufacturer protocol. Analysis was performed with Bio-Rad QuantaSoft Version 1.7.4. Results: Median yield of cfDNA was 1.97ng/ml. This correlated well with other yields from this sample set (Pearson’s r ¼ 0.6687, p < 0.0005), where a greater elution volume was used. 74 samples yielded >10,000 droplets and were included for analysis. 12 samples contained BRAF mutant positive droplets. Concordance rate between tissue BRAF status and the presence/absence of cfDNA BRAF mutant droplets was 74%. 7/18 tissue BRAF mutant samples were cfDNA BRAF mutant positive, in contrast to 2/32 tissue BRAF wild-type samples. The presence of BRAF mutant positive droplets in cfDNA was significantly different between tissue BRAF mutant and wild-type groups (p ¼ 0.004). The presence of BRAF mutant droplets did not confer any significant difference in rate of relapse (p ¼ 0.758) or mortality rate (p ¼ 0.654). Conclusions: A very low volume of cfDNA can be used to detect BRAF mutations in patients with melanoma through ddPCR. Having used small yields of cfDNA, it is possible that some samples fell below the limit of detection for ddPCR. Longitudinal study is warranted to monitor cfDNA BRAF status, and whether this correlates with relapse and disease progression. Legal entity responsible for the study: The authors. Funding: Weston Park Cancer Charity. Disclosure: All authors have declared no conflicts of interest.
doi:10.1093/annonc/mdz413 | vii15
Downloaded from https://academic.oup.com/annonc/article-abstract/30/Supplement_7/mdz413.052/5613350 by guest on 31 December 2019
significantly increased. RNA sequencing analysis using GSCs grown in stem cell culture or serum culture conditions revealed that differentiated GSCs are enriched with mitochondria-related gene signature. Also, glucose metabolism was increased in GSCs grown in stem cell culture conditions compared to serum culture conditions. Conclusions: Under different culture conditions, GSCs show different mitochondrial biogenesis and cellular metabolism. Understanding mitochondria properties in cancer may lead us to discover a novel therapeutic target. Editorial acknowledgement: Cancer Growth Regulation Laboratory for technical support and helpful advice. Legal entity responsible for the study: The authors. Funding: National Research Foundation of Korea (NRF) [grant numbers: 2015R1A5A1009024 and 2017R1E1A1A01074205], the School of Life Sciences and Biotechnology for BK21 Plus, Korea University and Institute of Animal Molecular Biotechnology, Korea University. Disclosure: All authors have declared no conflicts of interest.
Association of metastatic pattern and molecular status in metastatic lung non-small cell lung cancer adenocarcinomas
A. Dormieux1, L. Mezquita2, P-H. Cournede3, L. Lacroix2, E. Rouleau2, J. Adam4, F. Facchinetti2, F. Aboubakar4, M-V. Bluthgen2, C. Naltet2, P. Lavaud5, A. Gazzah2, C. Le Pechoux6, C. Balleyguier1, D. Planchard2, B. Besse7, C. Caramella1 1 Radiology, Institut Gustave Roussy, Villejuif, France, 2Institut Gustave Roussy, Villejuif, France, 3MICS Laboratory, CentraleSupe´lec, Gif-sur-Yvette, France, 4Pathology, Institut Gustave Roussy, Villejuif, France, 5Medical Oncology Departement, Institut Gustave Roussy, Villejuif, France, 6Radiation Oncology, Institut Gustave Roussy, Villejuif, France, 7 Oncology, Institut Gustave Roussy, Villejuif, France
45P
Circulating tumour associated cells in esophageal cancers are resistance educated per previous chemo treatments
S. Limaye1, T. Crook2, A. Ranade3, D. Patil4, D. Akolkar4, V. Datta4, S. Schuster5, R. Page6, C. Sims4, R. Patil4, A. Srinivasan4, S. Khan4, S. Patil4, V. Mhase4, S. Apurwa4, R. Datar4 1 Medical Oncology, Kokilaben Dhirubhai Ambani Hospital, Mumbai, India, 2St Luke’s Cancer Centre, Royal Surrey County Hospital, Guildford, UK, 3Avinash Cancer Clinic, Pune, India, 4Research and Innovations Department, Datar Cancer Genetics, Nashik, India, 5Datar Cancer Genetics Europe GmbH, Bayreuth, Germany, 6Biomedical Engineering, Worcester Polytechnic Institute, Worcester, MA, USA Background: Innate and acquired chemoresistance to anticancer therapies are a wellknown phenomenon in Esophageal Squamous Cell Carcinomas (ESCC). There are presently no viable approaches for real-time monitoring of resistance in ESCC. We used a novel method for chemo-interrogation (CI) by harvesting sufficient number of Circulating-Tumor Associated Cells (C-TACs) which are defined as apoptosis-resistant cells of tumorigenic origin and are positive for Epithelial Cell Adhesion Molecule (EpCAM) and pan-cytokeratins (pan-CK) irrespective of CD45 status. Methods: Peripheral blood was collected from 80 patients with confirmed diagnosis of ESCC, among whom 52 were recently diagnosed therapy-naı¨ve, while 28 were pretreated patients. Peripheral blood mononuclear cells (PBMCs) were harvested by centrifugation and treated with commercially available stabilizing agents, by a proprietary protocol, to stabilize apoptosis resistant C-TACs. Surviving C-TACs were confirmed by immunostaining for EpCAM, pan-CK and CD45. Harvested C-TACs were treated in vitro with a panel of conventional cytotoxic anticancer agents and fraction of surviving cells were estimated to determine resistance profiles. Results: Among the 52 cases of recently diagnosed therapy naı¨ve ESCC, innate chemoresistance was observed towards platinum agents in 40.8% samples (unique patientdrug combinations), taxanes in 34.6% samples, topoisomerase inhibitors in 42.9% samples and antimetabolites in 34.6% samples. Among the 28 cases of previously treated ESCC, resistance towards previously administered systemic agents was observed in 87% of all samples, which included resistance towards platinum agents in 87.5% samples, taxanes in 82.1% samples, topoisomerase inhibitors in 100% samples and antimetabolites in 85.7% samples, respectively. Conclusions: We show for the first time that sufficient C-TACs can be harvested for meaningful CI in newly diagnosed treatment naı¨ve ESCC as well as refractory ESCCs. Post-treatment chemoresistance being an order of magnitude higher than the untreated cohort, indicates that C-TACs in ESCC are resistance-educated by previous treatments and can guide treatment strategy in ESCC.
vii14 | Molecular Analysis for Personalised Therapy
Legal entity responsible for the study: The authors. Funding: Datar Cancer Genetics Limited. Disclosure: D. Patil: Full / Part-time employment: Datar Cancer Genetics Limited. D. Akolkar: Full / Part-time employment: Datar Cancer Genetics Limited. V. Datta: Full / Part-time employment: Datar Cancer Genetics Limited. S. Schuster: Full / Part-time employment: Datar Cancer Genetics Limited. C. Sims: Full / Part-time employment: Datar Cancer Genetics Limited. R. Patil: Full / Parttime employment: Datar Cancer Genetics Limited. A. Srinivasan: Full / Part-time employment: Datar Cancer Genetics Limited. S. Khan: Full / Part-time employment: Datar Cancer Genetics Limited. S. Patil: Full / Part-time employment: Datar Cancer Genetics Limited. V. Mhase: Full / Part-time employment: Datar Cancer Genetics Limited. S. Apurwa: Full / Part-time employment: Datar Cancer Genetics Limited. R. Datar: Shareholder / Stockholder / Stock options, Licensing / Royalties, Officer / Board of Directors: Datar Cancer Genetics Limited. All other authors have declared no conflicts of interest.
46P
Prevalence of homologous recombination deficiency (HRD)-related signatures indicates that a wider range of prostate cancer patients may benefit from PARP-inhibitor therapy
Z. Sztupinszki1, M. Diossy1, M. Krzystanek1, J. Borcsok1, M. Pomerantz2, V. Tisza3, S. Spisak2, O. Rusz4, I. Csabai5, M. Freedman2, Z. Szallasi3 1 Translational Cancer Genomics, The Danish Cancer Society, Copenhagen, Denmark, 2 Department of Medical Oncology, Dana-Farber Cancer Institute, Boston, MA, USA, 3 Computational Health Informatics Program, Boston Children’s Hospital, Boston, MA, USA, 4Semmelweis University, Budapest, Hungary, 5Department of Physics of Complex Systems, Department of Physics of Complex Systems, Budapest, Hungary Background: Prostate cancers with mutations in genes involved in homologous recombination (HR), most commonly BRCA2, respond favorably to PARP-inhibition and platinum-based chemotherapy. Currently targeted sequencing of key HR genes serves as an indicator of homologous recombination deficiency (HRD) in prostate cancer. It is well known, however, that other mechanisms, such as suppression of expression, rare mutations of HR-related genes can also cause HRD. Methods: HRD levels can be estimated using various mutational, genomic signatures derived from next-generation sequencing data. We used this approach to determine whether prostate cancer cases display clear signs of HRD in somatic tumor biopsies. Whole genome (n ¼ 311) and whole exome sequencing data (n ¼ 498) of both primary and metastatic prostate adenocarcinomas (PRAD) were analyzed. Results: Known BRCA-deficient samples showed robust signs of HR-deficiency associated mutational signatures. HRD-patterns were also detected in a subset of patients who did not harbor germline or somatic mutations in BRCA1/2 or other HR-related genes. The signs HRD present low intratumor heterogeneity and in most cases, the metastasis’s HRD-status is similar to the primary tumor’s. Patients with HRD signatures had a significantly worse prognosis than patients without signs of HRD. Conclusions: Based on the HRD associated mutational and genomic scar signatures, 58 % of prostate cancer cases may be good candidates for PARP-inhibitor treatment (including those with BRCA1/2 mutations). The prioritization of these patients for clinical trials, extending PARP-inhibitor therapy to all cases with HRD associated mutational signatures, even to those without BRCA1/2 mutations, will likely increase the efficacy of this treatment. Legal entity responsible for the study: The authors. Funding: Research and Technology Innovation Fund (KTIA_NAP_13-2014-0021 to Z.S.); Breast Cancer Research Foundation (BCRF-17-156 to Z.S.) and the Novo Nordisk Foundation Interdisciplinary Synergy Programme Grant (NNF15OC0016584 to Z.S.), Department of Defense through the Prostate Cancer Research Program (award number is W81XWH-18-2-0056) to Z.S. and M.F. The Danish Cancer Society grant (R90-A6213 to MK). Z.S. and J.B. was supported by Velux Foundation 00018310 grant. Disclosure: Z. Szallasi: Licensing / Royalties: Inventor on a patent used in the myChoice HRD assay: Myriad. All other authors have declared no conflicts of interest.
47P
The properties of mitochondria in glioma stem cells
N. Hong, E-J. Kim, H. Kim Department of Biotechnology, Korea University, Seoul, Republic of Korea Background: Mitochondria have a key role in cell survival, signaling, metabolism, and homeostasis. However, whether their biological properties affected by differentiation status of cells are not well known. This study aimed to compare structural and functional differences of mitochondria between differentiated cancer cells and undifferentiated cancer stem cells. Methods: Glioma stem cells (GSCs) were cultured in stem cell culture conditions to maintain their stem cell properties or serum culture conditions to induce their differentiation. Characteristics of GSCs are analyzed through sphere-forming ability and expression of stem cell/differentiation markers. Mitochondrial biogenesis and structure was examined by transmission electron microscopy analysis. Bioinformatics analysis, qRT-PCR, glucose uptake assay and lactate production assays were applied to reveal metabolic difference. Results: GSCs grown in serum culture conditions showed lower stem cell markers expression and higher differentiation markers compared to GSCs grown in stem cell culture conditions. In the serum culture conditions, biogenesis of mitochondria was
Volume 30 | Supplement 7 | November 2019
Downloaded from https://academic.oup.com/annonc/article-abstract/30/Supplement_7/mdz413.052/5613350 by guest on 31 December 2019
Background: The recent identification of molecular alterations in some lung adenocarcinomas has led to the emergence of effective targeted therapies thus drastically improving their prognosis.The aim of our study was to investigate the association between driver oncogene alterations and metastatic patterns on imaging assessment, in a large cohort of metastatic lung adenocarcinoma patients. Methods: From January 2010 to May 2017, 656 patients with stage IV lung adenocarcinoma with molecular analysis were studied retrospectively including 135 EGFR, 81 ALK, 47 BRAF, 141 KRAS, and 146 negative tumors for these 4 mutations (4N). After review of the complete imaging report by two radiologists (junior and senior) to identify metastatic sites, univariate correlation analyzes were performed. Results: We found differences in metastatic tropism depending on the molecular alteration type when compared to the non-mutated 4N group: in the EGFR group, pleural metastases were more frequent (32% versus 20%; p ¼ 0.021), and adrenal and node metastases less common (6% versus 23%; p < 0.001 and 11% versus 23%; p ¼ 0.011). In the ALK group, there were more brain and lung metastases (respectively 42% versus 29%; p ¼ 0.043 and 37% versus 24%; p ¼ 0.037). In the BRAF group, pleural and pericardial metastases were more common (respectively 47% versus 20%; p < 0.001 and 11% versus 3%; p ¼ 0.04) and bone metastases were rarer (21% versus 42%; p ¼ 0.011). Lymphangitis was more frequent in EGFR, ALK and BRAF groups (respectively 6%, 7% and 15% versus 1%); p ¼ 0,016, p ¼ 0,009 and p < 0,001). Conclusions: The application of these correlations between molecular status and metastatic tropism in clinical practice may lead to earlier and more accurate identification of patients for targeted therapy, savings in iterative biopsies, as well as improved and personalized imaging interpretation. Legal entity responsible for the study: Alison Dormieux. Funding: Has not received any funding. Disclosure: All authors have declared no conflicts of interest.
Annals of Oncology
abstracts
Annals of Oncology
48P
PLSCR1 and XKR8: New markers for low-grade glioma progression and outcome
K.V. Havrysh1, M. Bogdanov2, R. Kiyamova1 Research Laboratory "Biomarker" IFMB, Kazan Federal University, Kazan, Russian Federation, 2Department of Biochemistry and Molecular Biology, Center for Membrane Biology, University of Texas McGovern Medical School at Houston, TX, USA
1
Background: One of the most frequently diagnosed oncological diseases among young persons is Low-grade glioma (LGG). More than 80% of LGGs progress to higher-grade tumors always lethal to a patient. LGG is an extremely heterogeneous group of primary glial brain tumors with a quite variable rate of recurrence, survival, and response to treatment. Thus a study of new biomarkers associated with disease progression and patient outcome is useful to improve patient management. Genes involved in key cellular processes, e.g., apoptosis, are promising prognostic and predictive cancer markers. Phosphatidylserine (PS) plays an essential role in apoptosis signaling. Several studies suggested PS is dysregulated in the tumor microenvironment and offends the development of anti-cancer immune response. The scramblases PLSCR1 and XKR8 allegedly could be associated with the progression and outcome of LGG since they could be involved in the development of immunity via regulation of PS activity by its bidirectional translocation in the plasma membrane. Thus this study was aimed to evaluate associations of PLSCR1 and XKR8 with disease-free and overall survival of LGG patients. Methods: Gene expression values and clinical information of 530 LGG patients were downloaded from TCGA Low-Grade Glioma study using the public resource CBioPortal. According to gene expression values (RNA Seq V2 RSEM) patients were divided into two groups: with downregulated (< mean) or upregulated (> mean) PLSCR1 or XKR8 gene. The Kaplan-Meier procedure and Log-Rank test were used for survival analysis. All calculations were performed in the integrated development environment - RStudio. Results: It was shown the LGG patients with upregulated PLSCR1 (p ¼ 2.54e-14) and XKR8 (p ¼ 7.33e-15) gene had worse disease prognosis than other patients. Similar associations were seen in the study of LGG progression, where patients with upregulated PLSCR1 (p ¼ 7.75e-07) and XKR8 (p ¼ 5.59e-10) gene had a higher risk of cancer recurrence. Conclusions: Consequently, PLSCR and XKR8 genes could be considered as new prognostic biomarkers of LGG progression and outcome. Legal entity responsible for the study: Research Laboratory "Biomarker". Funding: The work was performed according to the Russian Government Program of Competitive Growth of KFU. Disclosure: All authors have declared no conflicts of interest.
49P
An exceptional response to immunotherapy doublet in combined hepatocellular carcinoma-cholangiocarcinoma
E. Tahover Oncology, Shaare Zedek Medical Center, Jerusalem, Israel Background: We present here a case of a 67-year-old male. He presented in May 2018 with severe abdominal pain and weight loss of 25%. Alpha-feto protein (AFP) and Carbohydrate antigen 19-9 (CA19-9) were within normal limits, and CA125 was 5 times upper normal limit (ULN). Computed tomography showed liver masses and enlarged retroperitoneal lymph nodes. Biopsy from a liver mass showed Combined Hepatocellular Carcinoma (HCC)-Cholangiocarcinoma (CC) (CHC). This a rare tumor, with an incidence of less than 10% of primary liver tumors. In whole exome analysis, mutations suggested the bipotent cell origin and stem cell origin. There are no guidelines or randomized trials regarding treatment. In an analysis of 36 patients who were treated with chemotherapy, the progression-free survival was 2.8 months, with an overall response rate of 5.6%. Targeted agents had minimal effect on survival.
Volume 30 | Supplement 7 | November 2019
Methods: In our case, extensive genomic, transcriptomic and proteomic testing was performed. No genomic alterations were identified, tumor mutational burden was low and microsatellite status was stable. 7 of 9 immune checkpoint genes were overexpressed. A variant in CDK12 was noted, which was shown to be associated with elevated neoantigen burden and may predict benefit from immune checkpoint therapy. Results:
Table: 49P Blood tests of case
Alk Phos GGT Ca125
Before immunotherapy
After 11 months of immunotherapy
Normal range
456 588 210
83 72 21
38-150 15-73 2-35
The patient began immunotherapy with ipilimumab and nivolumab followed by nivolumab, which he is continuing. The only side effects were hypothyroidism and Addison’s disease which are being treated. His clinical response was dramatic, he regained all the lost weight, and discontinued high dose opiate treatment. ECOG performance status improved from 3 to 0. Repeated PET-CT showed near complete response, ca125 decreased by 90% and liver function tests normalized. Conclusions: We present here an exceptional case of a rare tumor, where the patient had a clinical, laboratory and radiological response and a significant improvement in quality of life, suggesting that these tumors are sensitive to immunotherapy. No published cases of this tumor have yet been treated with immunotherapy doublet. Legal entity responsible for the study: The author. Funding: Has not received any funding. Disclosure: The author has declared no conflicts of interest.
50P
Assessing melanoma BRAF status through ddPCR of cfDNA
L. Passby1, S. Silva1, I. Brock1, G. Wells1, A. Cox1, S. Danson2 Department of Oncology and Metabolism, University of Sheffield Medical School, Sheffield, UK, 2Department of Oncology and Metabolism and Sheffield Experimental Cancer Medicine Centre, University of Sheffield Medical School, Sheffield, UK
1
Background: Targeted therapy has revolutionised the systemic treatment of melanoma. Patients with stage III/IV melanoma with a mutation in BRAF are treated with BRAF inhibitors, with BRAF mutation status being assessed by tissue analysis. Cell-free DNA (cfDNA) released from tumours provides a method for detecting BRAF mutations. This project analysed BRAF status in cfDNA through digital droplet PCR (ddPCR). We aimed to assess the relationship between cfDNA BRAF status and disease relapse and progression. Methods: Plasma from 100 patients with active or recently resected melanoma was obtained during previous work. 85 samples had cfDNA extracted. Tissue BRAF status was known for 57 samples. cfDNA was extracted from 1-2ml plasma with the QIAamp R ) following manufacturer protocol, eluting circulating nucleic acid kit (QIAGENV cfDNA into 100mL. cfDNA was quantified with SYBR green quantitative real-time PCR (Life Technologies), based on an 87bp GAPDH gene amplicon. ddPCRTM was performed using the Bio-Rad QX200 Droplet GeneratorTM and Droplet Reader as per manufacturer protocol. Analysis was performed with Bio-Rad QuantaSoft Version 1.7.4. Results: Median yield of cfDNA was 1.97ng/ml. This correlated well with other yields from this sample set (Pearson’s r ¼ 0.6687, p < 0.0005), where a greater elution volume was used. 74 samples yielded >10,000 droplets and were included for analysis. 12 samples contained BRAF mutant positive droplets. Concordance rate between tissue BRAF status and the presence/absence of cfDNA BRAF mutant droplets was 74%. 7/18 tissue BRAF mutant samples were cfDNA BRAF mutant positive, in contrast to 2/32 tissue BRAF wild-type samples. The presence of BRAF mutant positive droplets in cfDNA was significantly different between tissue BRAF mutant and wild-type groups (p ¼ 0.004). The presence of BRAF mutant droplets did not confer any significant difference in rate of relapse (p ¼ 0.758) or mortality rate (p ¼ 0.654). Conclusions: A very low volume of cfDNA can be used to detect BRAF mutations in patients with melanoma through ddPCR. Having used small yields of cfDNA, it is possible that some samples fell below the limit of detection for ddPCR. Longitudinal study is warranted to monitor cfDNA BRAF status, and whether this correlates with relapse and disease progression. Legal entity responsible for the study: The authors. Funding: Weston Park Cancer Charity. Disclosure: All authors have declared no conflicts of interest.
doi:10.1093/annonc/mdz413 | vii15
Downloaded from https://academic.oup.com/annonc/article-abstract/30/Supplement_7/mdz413.052/5613350 by guest on 31 December 2019
significantly increased. RNA sequencing analysis using GSCs grown in stem cell culture or serum culture conditions revealed that differentiated GSCs are enriched with mitochondria-related gene signature. Also, glucose metabolism was increased in GSCs grown in stem cell culture conditions compared to serum culture conditions. Conclusions: Under different culture conditions, GSCs show different mitochondrial biogenesis and cellular metabolism. Understanding mitochondria properties in cancer may lead us to discover a novel therapeutic target. Editorial acknowledgement: Cancer Growth Regulation Laboratory for technical support and helpful advice. Legal entity responsible for the study: The authors. Funding: National Research Foundation of Korea (NRF) [grant numbers: 2015R1A5A1009024 and 2017R1E1A1A01074205], the School of Life Sciences and Biotechnology for BK21 Plus, Korea University and Institute of Animal Molecular Biotechnology, Korea University. Disclosure: All authors have declared no conflicts of interest.