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Sedova, Marina, Alexy Ongpin, Jennifer Burke, et al. "Abstract 764: Fully automated sample-to-report NGS workflow for comprehensive genomic profiling for myeloid neoplasms." Cancer Research 82, no. 12_Supplement (2022): 764. http://dx.doi.org/10.1158/1538-7445.am2022-764.
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Abstract Introduction: Myeloid malignancies are associated with a broad and diverse set of genomic alterations, including SNVs, insertions, deletions and gene fusions. Comprehensive characterization of genetic mutations in hematological disorders currently requires a variety of diagnostic tests and takes multiple days to complete. We developed a fully automated NGS Myeloid Assay that offers an easy to use sample-to-report workflow and the capability for processing up to 8 samples per day. Methods: The Genexus System is comprised of two software linked instruments, the Genexus Purification System and the Genexus Integrated Sequencer. The Genexus Purification System was used to isolate the DNA and RNA from blood or bone marrow samples from precharacterized myeloid samples representative of Acute Myeloid Leukemia (AML) and Myelodysplastic Syndrome (MDS) and from the blood of healthy donors. The Genexus Integrated Sequencer was used to dilute the nucleic acids to optimal concentration and to sequence the samples in replicates with Oncomine Myeloid Genexus v2 Assay. Six DNA and RNA samples were sequenced per run per day along with commercially available analytical controls and a No Template Control. The report was generated by the Genexus Software analysis pipeline optimized to detect different variant types with high sensitivity and specificity. Results: The purification workflows were tested with blood input of 50-400uL for DNA and 50-150uL for RNA. Genexus Purification System extracted and quantified nucleic acids showed input dependent yields. DNA and RNA yields obtained with 50uL sample inputs consistently met the 27.5ng DNA and 15ng RNA minimum requirements for Genexus Integrated Sequencer workflows. DNA libraries had >97% Uniformity of Amplicon Coverage and >95% Target Base Coverage at 350x. The percentage ratio of Mapped Reads for DNA and RNA libraries was approximately 80:20. Detected genetic variations included key hotspots in CEBPA, FLT3, IDH1/2, NPM1, NRAS, RUNX1, and U2AF1 genes that are prevalent in AML and MDS. Genexus Variant Calling results showed high reproducibility and high concordance to the Ion GeneStudio S5 sequencing platform (>95%). The analytical controls, AcroMetrix Oncology Hotspot Control, Seraseq Myeloid Mutation DNA Mix and Seraseq Myeloid Fusion RNA Mix, were sequenced with Sensitivity and PPV >95%. Conclusion: The Genexus System offers an automated sample-to-report workflow with minimal hands-on-time and run results in 30 hours which allows an easy to use solution for next day turnaround time. When used with the Oncomine Myeloid GX v2 Assay, it provides accurate and comprehensive information on diverse mutations including fusions that are relevant to the study of myeloid cancers. For research use only. Not for use in diagnostic procedures. Citation Format: Marina Sedova, Alexy Ongpin, Jennifer Burke, Collyn Seeger, Sarah Brozio, Janice Au-Young, Jiajie Huang, Thilanka Jayaweera, Iris Casuga, Milton Huynh, Fiona Hyland. Fully automated sample-to-report NGS workflow for comprehensive genomic profiling for myeloid neoplasms [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 764.
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Oliveira, Douglas Hamilton, and Vandré Felipe de Oliveira Nicolau. "Comunicação SMTP em aplicações Genexus – um comparativo entre a linguagem nativa e a não nativa." Revista Processando o Saber 03, no. 01 (2011): 82–99. https://doi.org/10.5281/zenodo.15571842.
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O artigo visa demonstrar a comunicação de aplicações GENEXUS com servidores de e-mails utilizando o protocolo SMTP que é responsável pelo envio de mensagens de correio eletrônico, para isso serão comentados duas formas de envio: a forma nativa do GENEXUS com a utilização de variáveis que, na verdade, são objetos instanciados de classes especificas do GENEXUS que tratam do protocolo SMTP e a outra forma usando linguagem externa não nativa que, no caso, será a linguagem da Microsoft C#.NET no framework 3.5. Para tanto, o artigo desenvolve um estudo de caso onde uma solução Genexus com recursos de envio de mensagens aplicado em um ERP acadêmico sofreu uma atualização sendo necessária a introdução de um web service que passou a gerenciar exclusivamente os processos de envio de mensagens.
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Reibán-Lucero, Franklin I., and Diego M. Cordero-Guzmán. "Catálogo de refactorización de código fuente para la herramienta CASE GeneXus." Polo del Conocimiento 4, no. 6 (2019): 173. http://dx.doi.org/10.23857/pc.v4i6.1005.
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<p style="text-align: justify;">La herramienta de ingeniería de software asistido por computador (CASE) GeneXus se utiliza para desarrollar software en base a la abstracción del conocimiento, permite modelar entidades del mundo real y definir sus características y comportamiento de forma declarativa pero también permite crear objetos basados en lógica procedural cuya construcción depende exclusivamente del desarrollador pues la herramienta no brinda asistencia de ningún tipo para el desarrollo, esto implica que no se verifique exhaustivamente la congruencia con el conocimiento que se captura en las entidades modeladas y que tampoco se optimicen los procesos implementados en los mencionados objetos basados en lógica procedural.</p>
 <p style="text-align: justify;">El objetivo de esta investigación fue crear un catálogo de refactorización de código fuente adaptando las mejores prácticas, de este ámbito, al desarrollo de software con la herramienta CASE GeneXus, de forma que se evite el deterioro causado por los ajustes de mantenimiento y la adición de nuevas funciones en una aplicación desarrollada con GeneXus. Esto se cumplió estableciendo un conjunto de reglas para optimizar las aplicaciones desarrolladas con GeneXus como un catálogo de refactorización de código fuente.</p>
 <p style="text-align: justify;">Los resultados obtenidos benefician especialmente a la comunidad de desarrolladores GeneXus, una comunidad basada en una cultura «Open Source» y en constante crecimiento. La relación entre la reducción del costo global de mantenimiento y la mejora de la inteligibilidad del código fuente de una aplicación desarrollada con GeneXus, es el beneficio más notorio, y, es también, el aspecto que se analiza mediante la discusión y conclusiones de este documento.</p>
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Zochowski, Kayla, Dinesh Cyanam, Geoffrey Lowman, et al. "Abstract 42: High-throughput next-generation sequencing research solutions for detection of oncology variants, gene fusion events, and key oncology endpoints." Cancer Research 82, no. 12_Supplement (2022): 42. http://dx.doi.org/10.1158/1538-7445.am2022-42.
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Abstract Introduction: The Ion Torrent Genexus System has redefined the genomic profiling paradigm as the first fully-integrated, next-generation sequencing (NGS) research platform to provide an automated sample-to-report workflow with results in a single day. With a purification instrument, an enhanced chip architecture, and downstream reporting, the Genexus System provides a convenient solution to enable oncology research. Here we highlight the oncology research applications and high-throughput NGS capabilities of the Genexus System with Oncomine Comprehensive Assay Plus (OCA Plus), an oncology research panel that can detect variants, fusions, and evaluate key oncology research endpoints. We demonstrate the ability of OCA Plus on Genexus to evaluate tumor mutational burden (TMB), microsatellite instability (MSI), loss of heterozygosity (LOH), and homologous recombination repair deficiency (HRD). Methods: The high-throughput capabilities of the Genexus System enable it to support large oncology research panels such as OCA Plus, which comprises over 13,000 amplicons. The extensive per sample coverage allows for comprehensive DNA and RNA genomic profiling of relevant cancer biomarkers in over 500 genes including detection of over 1,300 fusion isoforms. We utilized high-molecular weight and FFPE samples, reference controls, and orthogonally tested research samples to evaluate DNA variant calling, RNA fusion calling, and key oncology research endpoints, including MSI, LOH, TMB, and HRD. Results: Commercially sourced reference control and research samples were sequenced using OCA Plus on the Genexus System. Sequencing data metrics showed ≥24 million reads per sample, with four samples supported per run. The high-throughput capacity of the Genexus chip architecture results in &gt;95% of amplicons achieving a minimum of 500X coverage with an average coverage uniformity of ≥95% when evaluated across all &gt;13,000 amplicons. Variant calling was assessed using the AcroMetrix Oncology Hotspot Control which has 328 hotspot variants covered by OCA Plus. Variant calling performance showed a hotspot and de novo variant sensitivity and PPV &gt;95%. MSI score and status were assessed using known MSI-High and microsatellite stable (MSS) research samples and FFPE samples of interest. Results show high concordance with data from OCA Plus on GeneStudio. Conclusions: The increased throughput of the Genexus System combined with minimal touchpoints and a rapid turnaround time enables comprehensive genomic profiling for research assays such as OCA Plus where an increased number of sequencing reads leads to greater sensitivity for detecting rare variants and low-level fusion transcripts. Further, accurate characterization of key oncology research endpoints, such as TMB, MSI, LOH, and HRD, allow the Genexus System to accelerate research in the field of oncology. Citation Format: Kayla Zochowski, Dinesh Cyanam, Geoffrey Lowman, Jennifer Kilzer, Sameh El-Difrawy, Yan Zhu, Tanaya Puranik, Alex Phan, Derek Wong, Edith Kwong, Coleen Nemes, Iris Casuga, Frances Chan, Eun Ji Kim, Jianjun Guo, Vinay Mittal, Emily Norris, Shrutii Sarda, Mohit Gupta, Fan Shen, Steven Roman, Gabriel Vargas, Ying Jin, Annie Kraltcheva, Paul Williams, Amneet Gulati, Justin Rigby, Christopher Hoff, Richard Meade, Elaine Wong-Ho, Andrew Wong, Jamsheed Ghadiri, David Garcia-Viramontes, Scott Myrand, Seth Sadis. High-throughput next-generation sequencing research solutions for detection of oncology variants, gene fusion events, and key oncology endpoints [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 42.
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Casuga, Iris, Frances Chan, Milton Huynh, et al. "Abstract 2944: Rapid and accurate variant calling of FFPE samples with the Genexus System." Cancer Research 82, no. 12_Supplement (2022): 2944. http://dx.doi.org/10.1158/1538-7445.am2022-2944.
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Abstract Next-Generation Sequencing technology has enhanced oncology research by enabling the detection of all cancer related variants into one assay for research and drug discovery programs. The Oncomine࣪ Comprehensive Assay v3, a pan-cancer panel, used with the Ion Torrent࣪ Genexus࣪ System allows for formalin-fixed paraffin embedded (FFPE) samples to be examined across 161 unique genes in an automated sample to result workflow in 30hrs. This study demonstrates ≥ 95% Sensitivity and PPV for detecting SNV, Indel, and Copy Number variants of clinical FFPE samples along with fusions. When tested using analytical controls, the Genexus System achieves ≥ 98% Sensitivity for hotspot variants, ≥ 95% for de novo variants, and 100% Sensitivity for fusion variants. The Ion Torrent࣪ Genexus࣪ System is fully automated and consists of two software linked instruments, the Ion Torrent࣪ Genexus࣪ Purification System and the Ion Torrent࣪ Genexus࣪ Integrated Sequencer. For this study, the purification system was used to sequentially extract and quantify DNA & RNA samples from human colon and lung FFPE tumor tissue. The purification instrument provides minimal hands-on-time, ease of use and compatibility with the integrated sequencer. It extracts and quantifies the nucleic acids, records the quantitation values and transfers the nucleic acids to an output plate. The plate is transferred from the purification instrument directly to the sequencer for sample dilution, library preparation, and sequencing using the Oncomine࣪ Comprehensive Assay v3 panel. Variant calling analysis of the DNA & RNA pairs is completed immediately following sequencing. Results show that the variants are detected reproducibly with ≥ 95% Sensitivity and PPV when compared to the Ion GeneStudio࣪ S5 System as an orthogonal method. All variants in the report were correct with p-values ≤ 10-5. Additionally, a subset of the extracted samples was evaluated using Sanger Sequencing to verify hotspot mutations found in samples sequenced on both the Genexus࣪ System and GeneStudio࣪ S5 System. Results show that all expected hotspot variants were detected. In summary, the Ion Torrent࣪ Genexus࣪ System is a reliable and fast turnaround solution for sample-to-variant calling results. When used with the Oncomine࣪ Comprehensive Assay v3 panel, the system provides accurate identification of tumor markers for oncology research. Citation Format: Iris Casuga, Frances Chan, Milton Huynh, Gregory R. Govoni, Kayla Zochowski, Thilanka Jayaweera, Janice Au-Young. Rapid and accurate variant calling of FFPE samples with the Genexus System [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 2944.
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Kreston, Sarah, Claire Gould, Kylie Blair, Leisa Jackson, Janice Riley, and Gary A. Pestano. "Abstract 5547: Evaluation of two clinically focused targeted NGS systems for liquid biopsy testing shows a high level of concordance in resulting actionable mutations." Cancer Research 83, no. 7_Supplement (2023): 5547. http://dx.doi.org/10.1158/1538-7445.am2023-5547.
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Abstract Introduction: The use of blood-based molecular diagnostics is increasingly becoming routine in clinical oncology practice. The purpose of this study was to evaluate variant resulting concordance for the four major somatic variant classes using two clinical next-generation sequencing (NGS) systems for conducting highly sensitive blood-based analyses. All donor specimens were de-identified remnants from patients previously diagnosed with advanced NSCLC. We utilized two independent panels, systems and bioinformatic pipelines that are focused on clinical testing for key actionable variants. The fifty-gene panel Oncomine Precision Assay (OPA) GX was performed on the Genexus Integrated Sequencer and the 52 gene GeneStrat NGS (GSNGS) test was run as a reference on the Ion GeneStudio S5 PRIME system. The four major classes of mutations evaluated included Single Nucleotide Variants (SNV), Insertions and Deletions (INDEL), copy number amplifications (CNAs) and gene fusions. Methods: For this study, de-identified reference GSNGS cell-free nucleic acid (cfNA) remnant specimens (n=33 variants) that passed all validated QC bioinformatics thresholds were blinded and tested on the Genexus. DNA concentration was measured by fluorometry, and specimens were diluted to an input of 6.6 ng to 53.4 ng for library preparation. The Genexus system is pre-programmed to process specimens to result generation. Variant calls for all mutation classes were conducted using the on-board bioinformatics pipelines, and results were compared to those generated by the GeneStrat NGS Test. We included specimens harboring somatic variant mutations in KRAS, NRAS, BRAF, EGFR, ERBB2, and FGFR; CNAs; EML4-ALK fusions; and MET exon 14 skipping. Results: All samples passed the OPA (Genexus) bioinformatic quality control criteria and mapped reads were highly consistent, demonstrating accuracy of calls between both the Ion Torrent platforms, informatic pipelines and panels. We observed 100% concordance in variant calls for SNVs, INDELs and fusions between the two platforms. There was one CNA variant observed on the S5 that was not called by the Genexus workflow, yielding overall concordance of 97% (32 of 33). Of note, 44 additional somatic variants (SNV and INDELs) and 1 additional fusion were detected using the Genexus workflow. This was due to the lower threshold for variant calling on this system. Conclusion: The high concordance of the independent workflows for the detection of nucleic acid variants in circulation demonstrates the capability of both systems to be used for testing of clinical specimens. Specifically, actionable variants in the four major mutation classes were successfully detected in the reference and test specimen set. As a part of clinical validation, orthogonal testing of variants will need to be conducted. Citation Format: Sarah Kreston, Claire Gould, Kylie Blair, Leisa Jackson, Janice Riley, Gary A. Pestano. Evaluation of two clinically focused targeted NGS systems for liquid biopsy testing shows a high level of concordance in resulting actionable mutations. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 5547.
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Lowman, Geoffrey Marc, Dinesh Cyanam, Emily Norris, et al. "Abstract 232: Fully automated comprehensive genomic profiling for detection of cancer variants, gene fusions, and complex oncology endpoints." Cancer Research 83, no. 7_Supplement (2023): 232. http://dx.doi.org/10.1158/1538-7445.am2023-232.
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Abstract Introduction: We present the comprehensive genomic profiling performance of the Ion Torrent Genexus system using the Oncomine Comprehensive Assay Plus (OCA Plus), a 500+ gene targeted AmpliSeq-based oncology research panel that evaluates DNA variants (including copy number alterations), RNA fusions, and key oncology research endpoints including tumor mutational burden (TMB), microsatellite instability (MSI), and homologous recombination repair deficiency (HRD) via characterization of genomic instability by the newly introduced Genomic Instability Metric (GIM). Methods: The Ion Torrent Genexus System provides comprehensive genomic profiling via automated sample-to-report workflow with next day results. The Genexus System supports oncology research panels such as OCA Plus, which is comprised of over 13,000 amplicons, and enables low input requirements of just 20ng of FFPE DNA and RNA. This study utilized cell lines, reference controls, and orthogonally tested FFPE research samples to evaluate detection of DNA variants, copy number alterations, RNA fusions, and key research endpoints, including MSI, TMB, and HRD. The OCA Plus panel was also evaluated for the ability to detect arm-level copy number changes in orthogonally validated FFPE samples. Results: Commercial reference controls and FFPE research samples were sequenced using OCA Plus on the Genexus System to an average depth of ≥24 million reads per sample, with four DNA and RNA samples supported per run. SNV and MNV calling performance was assessed using the AcroMetrix Oncology Hotspot Control which has 377 variants covered by OCA Plus and delivered SNV sensitivity and PPV &gt;99% and MNV sensitivity of &gt;99% and PPV &gt;95%. MSI status was assessed using orthogonally tested FFPE samples from various tumor tissues (stomach, endometrial, colorectal) and returned status concordance of 99.4% with sensitivity and PPV &gt;99%. The TMB endpoint was tested using commercial controls and FFPE samples with a correlation of r2 &gt; 0.90 to orthogonal measurements. RNA Fusion reference controls showed 100% positive correlation. Copy number gain detection shows sensitivity of 99% and PPV &gt;95%, while homologous copy loss gives 100% PPV and sensitivity of &gt;90%. We also demonstrate high concordance to orthogonal methods in detection of HER2 amplifications, and the ability to detect arm-level copy number alterations such as 1p/19q co-deletions in IDH1 positive glioma samples. Conclusion: The Genexus System combines minimal touch points and a rapid turnaround time to enable comprehensive genomic profiling for research assays such as OCA Plus for detection of rare variants and low-level fusion transcripts. Further, by providing accurate characterization of key oncology research endpoints, the Genexus System can accelerate research in oncology. For research use only. Not for use in diagnostic procedures. Citation Format: Geoffrey Marc Lowman, Dinesh Cyanam, Emily Norris, Michelle Toro, Coleen Nemes, Tanaya Puranik, Yan Zhu, Alex Phan, Derek Wong, Portia Bernado, Anelia Kralcheva, Srinivas Nallandhighal, Loni Pickle, April Bigley, Mohit Gupta, Ying Jin, Sameh El-Difrawy, Amir Marcovitz, Fatima Zare, Charles Scafe, Yu-Ting Tseng, Jianjun Guo, Vinay Mittal, Scott Myrand, Santhoshi Bandla, Paul Williams, Eugene Ingerman, Elaine Wong-Ho, Seth Sadis, Mark Andersen, Rob Bennett. Fully automated comprehensive genomic profiling for detection of cancer variants, gene fusions, and complex oncology endpoints [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 232.
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Low, Siew-Kee Kee, Ken Uchibori, Rie Hayashi, et al. "Evaluation of Genexus system that automates specimen-to-report for cancer genomic profiling within a day using liquid biopsy." Journal of Clinical Oncology 38, no. 15_suppl (2020): 3538. http://dx.doi.org/10.1200/jco.2020.38.15_suppl.3538.
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3538 Background: Genomic profiling of patients’ tumors using NGS system help in facilitating molecular-guided therapy. The turnaround time from specimen to report by the NGS system is important to deliver result timely for clinical decisions. The Genexus Integrated Sequencer automates all steps of the targeted NGS workflow starting from nucleic acid of formalin-fixed paraffin-embedded tissues or plasma that significantly reduce laborious procedures. Importantly, the whole specimen-to-report workflow delivers results in a single day. In this study, we evaluated detection rate of alteration using Oncomine Precision Assay (OPA) on Genexus system with cell-free DNA (cfDNA) from non-small cell lung cancer (NSCLC). Methods: Among the cfDNA from 48 plasma samples of NSCLC were evaluated, 19 were newly-diagnosed cases with EGFR mutations in cancer tissues and 29 cases were patients who experienced progression of disease after first line of EGFR, ALK or ROS1-targeted therapy. 13-20ng of input cfDNA were subjected to automated Genexus Integrated Sequencer for library construction using OPA panel, templating and sequencing. OPA panel covers actionable hotspot mutations, copy number gains or loss, fusion drivers. The concordance of mutation profiles between the tumor tissue and cfDNA and detection of a resistance mutation(s) during molecular-targeted therapy were evaluated. Results: The sequencing resulted in median overall reads of 8,698,358, median overall depth of 30,648 (range 15,069-48,707) and median molecular coverage of 1,595 (range 859-2,550). Among 48 samples examined, 44 were detected to carry at least one somatic mutation, giving the detection rate of 92%. A total of 17 of 19 newly diagnosed EGFR-positive patients were also detected to carry EGFR mutations. Importantly, these 17 patients carry the same mutation that was found in tissue samples implying complete concordance. In addition, we found novel resistance mutations in plasma of the patients who were under EGFR, ALK or ROS1 targeted therapies. Conclusions: Genexus Integrated Sequencer is a fully automated and highly accurate NGS system with a 1-day turnaround time that could assist clinicians to make more timely decision. Novel actionable, resistance mutations were detected using OPA panel that provide potential options for molecular-guided therapy and may help the better understanding of resistance mechanism of targeted therapy.
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Schageman, Jeoffrey. "Oncomine Dx Express CE-IVD liquid biopsy assay for non-small cell lung cancer: Performance review and analytical validation." Journal of Clinical Oncology 41, no. 16_suppl (2023): e21218-e21218. http://dx.doi.org/10.1200/jco.2023.41.16_suppl.e21218.
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e21218 Background: Clinical diagnostics assays for oncology are becoming more readily available due to the advancement and democratization of Next-Generation Sequencing (NGS). Additionally, liquid biopsy can be used in NGS to detect genetic variants in circulating tumor DNA and RNA. Liquid biopsy is a less invasive option compared to traditional biopsy methods for early detection and continuous monitoring of cancer treatment outcomes. Here, we discuss screening of over 3,500 non-small cell lung cancer (NSCLC) samples, and the analytical validation of the Oncomine Dx Express Test (ODxET) and Genexus Dx Integrated Sequencer for detection of clinically significant variants in liquid biopsy samples. Methods: Our team screened over 3,500 NSCLC liquid biopsy samples in search of high priority clinical variants. Variants selected for analytical validation studies included ERBB2 exon 20 insertion, EGFR exon 20 insertion, EGFR exon 19 deletion, EGFR T790M, KRAS G12C, BRAF V600E, and RNA fusion isoforms including ALK, NTRK1/2/3, RET, and ROS1 oncogenic drivers. Over 550 screened samples were found to be positive for these variants of interest. This screening was performed on the Genexus Dx Integrated Sequencer according to the user guide. Results: The Genexus Dx Integrated Sequencer automates library preparation, sequencing, analysis, and reporting QC metrics and variant calls. Sequencing run setup is quick and straightforward, taking less than 15 minutes to start a run and just over 24 hours to go from nucleic acid to report. Limit of detection (LoD) for DNA SNVs, insertions, and deletions at 5 ng DNA input level ranged from 0.65% to 1.82% allelic frequency (AF), depending on the variant. The higher DNA input of 30 ng resulted in a lower LoD range, from 0.31% to 0.42% AF. RNA fusion and splice variant LoD at 5 ng sample input ranged from 9.9 to 19.6 molecular counts. The higher 30 ng input resulted in a lower LoD range for RNA variants as well, ranging from 6.4 to 8.0 molecular counts. In the analytical accuracy study, the false positive rate was found to be 0.2% for SNVs, 0% for insertions/deletions, and 0% for fusion targets. In the analytical reproducibility study, the average with-in run repeatability call rate (No Calls excluded) was 99.64% for DNA variants and 98.75% for RNA variants. Conclusions: Here, we demonstrated that the ODxET workflow on Genexus Dx Integrated Sequencer is a fast and efficient tool for testing clinical NSCLC liquid biopsy samples with high sensitivity and specificity.
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Burke, Jennifer, Frances Chen, Jiajie Huang, Geoffrey Lowman, Timothy Looney, and Marina Sedova. "69 Automated TRB locus haplotype analysis by long-amplicon TCRB chain sequencing for potential immune-related adverse events biomarker research." Journal for ImmunoTherapy of Cancer 8, Suppl 3 (2020): A75. http://dx.doi.org/10.1136/jitc-2020-sitc2020.0069.
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BackgroundIdentifying potential predictive biomarkers for immune related adverse events (irAEs) following checkpoint blockade inhibition (CPI) remains an outstanding goal of immune-oncology translational research. Polymorphism with the T cell receptor variable gene (TRBV) has been proposed as a potential risk factor for irAEs owing to a potential link between TRBV polymorphism and chronic autoimmune disease. Efforts to interrogate the potential biomarker utility of TRBV polymorphism have been hampered by the repetitive nature of the TRB locus. Our research has demonstrated a method for inferring TRB locus haplotypes from long-amplicon TCRB chain sequencing data, which we used to identify major haplotype groups in from nucleic acid. Here we present our research for a potential automated method for haplotype group assignment from TCRB chain sequencing data.MethodsRearranged TCRB chains from 10 blood samples were amplified and sequenced from 25ng peripheral blood total RNA via the Oncomine™ TCRB-LR assay using the Genexus™ Integrated Sequencer. 12 samples were run per chip with 4 samples run in each lane. TCRB clonotyping and repertoire feature analysis was performed using Genexus™ analysis software. Automated haplotype group assignment was performed by generation and comparison of TRBV allele profiles to those presented previously.1 For context, TCR evenness, convergence, and haplotype group assignment were compared to values obtained following analysis of the same samples via the GeneStudio™ S5 platform and Ion Reporter™ 5.12 software.ResultsTCR Evenness and convergence values were highly correlated across replicates run on the Genexus™ Integrated Sequencer (Spearman correlation >0.95 and >0.70, respectively). Evenness at equivalent clone count and convergence at equivalent sequencing depth were not significantly different across platforms (Spearman correlation >0.88). Haplotype group assignments demonstrated 100% agreement across replicates on both platforms.ConclusionsOur research has demonstrated a potential automated and reproducible method for TRB haplotype group assignment via the Oncomine™ TCR-Beta LR Assay, GX run on the Genexus™ Integrated Sequencer. Future studies will be needed to evaluate the potential biomarker utility of TRB haplotyping for the prediction of irAEs.For research use only not for diagnostic procedures.ReferenceLooney T, Duose D, Lowman G, Linch E, Hajjar J, Topacio-Hall D, Xu M, Zheng J, Alshawa A, Tapia C, Stephen B, Wang L, Meric-Bernstam F, Miller L, Glavin A, Lin L, Gong J, Conroy J, Morrison C, Hyland F, Naing A. Haplotype Analysis of the T-Cell Receptor Beta (TCRB) Locus by Long-amplicon TCRB Repertoire Sequencing. J Immunother Precis Oncol 2019;2:137–143.
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Jiwani, Shahanawaz, Jerald P. Radich, Maria Saeed, et al. "Rapid Clinical Mutation Screening for AML Using the Genexus Platform." Blood 142, Supplement 1 (2023): 2288. http://dx.doi.org/10.1182/blood-2023-188270.
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Introduction: The promise of “precision medicine” is using therapeutic agents matched to the inferred biology of a patient's particular disease. In acute myeloid leukemia this approach demands fast and accurate assessment of the mutation and cytogenetic landscape of each AML case. The NCI Myeloid Malignancies Molecular Analysis for Therapy Choice program (“myeloMATCH”) is a precision medicine clinical trial initiative across the NCI National Clinical Trials Network for patients with newly diagnosed acute myeloid leukemia and myelodysplasia. In myeloMATCH the goal is to have cytogenetic and mutation assays performed within 72 hours of central lab receipt, allowing for a rapid assignment of the appropriate clinical trial from the myeloMATCH trial portfolio. Here we report analytic performance of rapid genomic profiling laboratory developed test based on the Thermo Fisher Genexus Oncomine Myeloid Panel GXv2 (NCI Myeloid Assay version 2, or “NMAv2”) which performs rapid sequencing of SNVs, indels, and gene fusions, capturing 1,661 hotspot variants (45 genes) and 779 fusions (35 genes) with a turnaround time of &lt;48 hours from specimen receipt to clinical reporting. Methods: The validation testing was performed by two harmonized laboratories; NCI Molecular Characterization laboratory (MoCha) and Fred Hutch Molecular Oncology (MO). We conducted analytical validation and harmonization experiments for CLIA/CAP and FDA regulatory compliant use in this trial across both accredited clinical laboratories with harmonized workflows. Samples are extracted with AllPrep per manufacture protocol. Library preparation with the Oncomine Myeloid Assay GX v2 and the sequencing reactions are performed on the automated Ion Torrent Genexus System. Sequencing data generated by NMAv2 was analyzed using Ion Torrent Genexus system version 6.6.2.1. The default manufacturer workflow has been customized using a custom designed hotspot browser extensible data (BED) file, sequence variant baseline (SVB) file, fusion target file, torrent variant caller (TVC) version 5.18 and Oncomine extended filter chain (OEFC) version 5.16. In NMA, the default 2.5%, and 3% VAFs were set up for calling hotspot and non-hotspots SNV/indel, 1% VAF for FLT3-ITD and 100 read counts for all fusion, except 2000 read counts for KMT2A-PTD. Results: The validation set included 148 unique samples and comprised of 76 AML/MDS patients (49 peripheral blood, 27 bone marrow aspirate), 51 healthy donor blood, 6 healthy donor bone marrow, 7 cancer cell lines, 5 normal HapMap cell lines, and 3 contrived materials encompassing 364 known mutations (143 SNPs, 132 indels, 27 fusions). 180 runs were completed for this validation (106 at FH and 74 at MoCha) with 89.44% success rate in passing run level metrics. Results of the validation testing are detailed in Table 1. High reproducibility in detecting all reportable variants was observed, with overall &gt;99% mean positive percentage agreement (PPA) across both labs (Figure 1). The limit of detection for each variant type was &lt;0.06% for hotspot SNVs, 0.07% for non-hotspot SNVs, 0.51% for hotspot indels, ~1% for non-hotspot indels, 0.23% for FLT3-ITDs, and 50 minimum reads at 0.1% tumor content for fusions. 99.39% concordance was observed by testing 86 blinded validation sample set with analytically validated orthogonal assays. Conclusions: The NMAv2 has high specificity, sensitivity, accuracy and reproducibility (inter-lab and intra-lab) with sequencing results generated within 48 hours of assay initiation. As such, the assay is well suited for use to rapidly categorize the genomic alteration of AML to support clinical care in patients with active myeloid malignancies, including those patients potentially enrolled in the myeloMATCH program. Table 1: Results of validation experiment results for the NMAv2 Figure 1: Harmonization results for the NMAv2 between MoCha and MO labs, Right Panel = Indel; Left panel = SNV
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De Luca, Caterina, Francesco Pepe, Gianluca Russo, et al. "Technical Validation of a Fully Integrated NGS Platform in the Real-World Practice of Italian Referral Institutions." Journal of Molecular Pathology 4, no. 4 (2023): 259–74. http://dx.doi.org/10.3390/jmp4040022.
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Aims: To date, precision medicine has played a pivotal role in the clinical administration of solid-tumor patients. In this scenario, a rapidly increasing number of predictive biomarkers have been approved in diagnostic practice or are currently being investigated in clinical trials. A pitfall in molecular testing is the diagnostic routine sample available to analyze predictive biomarkers; a scant tissue sample often represents the only diagnostical source of nucleic acids with which to conduct molecular analysis. At the sight of these critical issues, next-generation sequencing (NGS) platforms emerged as referral testing strategies for the molecular analysis of predictive biomarkers in routine practice, but the need for highly skilled personnel and extensive working time drastically impacts the widespread diffusion of this technology in diagnostic settings. Here, we technically validate a fully integrated NGS platform on diagnostic routine tissue samples previously tested with an NGS-based diagnostic workflow by a referral institution. Methods: A retrospective series of n = 64 samples (n = 32 DNA, n = 32 RNA samples), previously tested using a customized NGS assay (SiRe™ and SiRe fusion), was retrieved from the internal archive of the University of Naples Federico II. Each sample was tested by adopting an Oncomine Precision Assay (OPA), which is able to detect 2769 molecular actionable alterations [hotspot mutations, copy number variations (CNV) and gene fusions] on fully integrated NGS platforms (Genexus, Thermo Fisher Scientific (Waltham, MA, USA). The concordance rate between these technical approaches was determined. Results: The Genexus system successfully carried out molecular analysis in all instances. A concordance rate of 96.9% (31 out of 32) was observed between the OPA and SiRe™ panels both for DNA- and RNA-based analysis. A negative predictive value of 100% and a positive predictive value of 96.9% (62 out of 64) were assessed. Conclusions: A fully automatized Genexus system combined with OPA (Thermo Fisher Scientific) may be considered a technically valuable, time-saving sequencing platform to test predictive biomarkers in diagnostic routine practice.
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Jasti, Madhuri, Swasti Raut, Diarra Hassell, et al. "Abstract 3671: Analytical performance of the Oncomine™ Dx Express Test, a CE-IVD NGS liquid biopsy assay for identification of clinically relevant variants." Cancer Research 84, no. 6_Supplement (2024): 3671. http://dx.doi.org/10.1158/1538-7445.am2024-3671.
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Abstract For in vitro Diagnostic use only. Not available in all regions including the United States. Introduction Minimally invasive blood-based liquid biopsy using cell-free total nucleic acid (cfTNA) and next generation sequencing (NGS) has substantially evolved in the field of clinical diagnostics for oncology, for detection of molecular therapeutic targets in non-small cell lung cancer (NSCLC). Here we describe analytical validation results for cfTNA using the CE-IVD Oncomine™ Dx Express Test (ODxET) assay, a qualitative in vitro diagnostic test that uses targeted NGS technology, the Ion Torrent Genexus™ Dx Integrated Sequencer to detect SNV, indel and copy number gain present in 42 genes and fusions in 7 genes from cfTNA isolated from NSCLC plasma samples. Methods Plasma samples from NSCLC patients were screened for variants of interest on several important gene loci. Screening involved isolation of cfTNA from K2-EDTA-derived plasma using the Genexus™ Cell-Free Total Nucleic Acid Purification kit on an automated purification system and sequencing was performed on the Genexus™ Dx integrated sequencer using the ODxET assay. Plasma samples having clinically relevant variants from the set of screened plasma samples were used to perform the validation studies. Results Analytical sensitivity and specificity of NSCLC plasma samples were characterized through determination of limit of detection (LOD) and limit of blank (LOB), respectively. The LOD level tested for DNA SNVs, and indels at 5 ng (minimum) input level ranged from 0.62% to 1.82% allelic frequency (AF), while 30 ng (maximum) input shows the LOD level ranging from 0.23% to 0.42% AF. The LOD for the tested RNA fusions ranged from 4.2 to 19.6 molecular counts for 5ng input and 4.9 to 8.0 molecular counts for 30ng input. Furthermore, LOB for the ODxET assay was determined by testing wild type (WT) cfTNA extracted from 30 healthy donor blood plasma samples, which were confirmed to be negative at all variant locations. The analytical accuracy study shows the false positive rate at 0.20% for SNVs, 0% for indel locations and fusion targets. In addition, the analytical reproducibility study shows 99.64% intra-run repeatability call rate for DNA variants and 98.75% for RNA variants. Conclusion In conclusion, we establish that the ODxET assay on Ion Torrent Genexus Dx Integrated Sequencer, is fast and efficient while demonstrating high sensitivity, specificity, and reproducibility for testing NSCLC liquid biopsy samples. Citation Format: Madhuri Jasti, Swasti Raut, Diarra Hassell, Nader Ezzedine, Daniela Garcia, Stephen Wunsch, Nick Siepert, Elliott Martinez, Emilia Ostrowska, Jeff Schagemann, Jian Gu, Thilanka Jayaweera, Luming He, Angie Cheng. Analytical performance of the Oncomine™ Dx Express Test, a CE-IVD NGS liquid biopsy assay for identification of clinically relevant variants [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 3671.
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Dhawale, Tejaswini, Valentina Nardi, Charlotte Wang, et al. "Impact of Ultra Rapid Molecular Profiling on Treatment Delays and Healthcare Utilization of Patients Hospitalized for Acute Leukemia." Blood 144, Supplement 1 (2024): 5058. https://doi.org/10.1182/blood-2024-208059.
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Background Molecular testing is fundamental to treatment decisions in patients with acute leukemia. The impact of rapid molecular profiling on the timing of treatment initiation and healthcare utilization in patients hospitalized for suspected acute leukemia has not been described. Methods We conducted a retrospective chart review of patients hospitalized and treated for suspected acute leukemia at Massachusetts General Hospital between September 2023 and April 2024. We conducted a detailed workflow analysis comparing the time to obtaining molecular profiling results from admission [“time to results” or TTR] between routine (legacy) methods (i.e., RT-PCT, karyotype, FISH, targeted DNA and RNA sequencing) and an ultra-rapid genomic profiling test (Thermo Fisher Genexus Oncomine Myeloid Panel, or “Genexus”) which was run in parallel. We used linear regression models to evaluate the impact of TTR on time to treatment initiation from admission [TTTI] and hospital length of stay [LOS]. Results Of the 111 patients who had ultra-rapid genomic profiling conducted during the study period, 57 (51.4%) were adult patients who were hospitalized for diagnostic workup and treatment of suspected acute leukemia. The most common leukemia subtypes were acute myeloid leukemia (n=39/57, 68.4%), B-lymphoblastic leukemia (n=9/57, 15.8%), and acute promyelocytic leukemia (n=4/57, 7.0%). The mean TTTI was 5.2 days (range 1-17) and mean LOS was 30.9 days (range 3-86). Genexus had a mean TTR of 3.1 days (range 1-8), whereas legacy methods had a mean TTR of 5.4 days (range 3-10). Longer TTR was associated with longer TTTI (b=0.95; SE=0.20; P=&lt;0.001) and longer LOS (b=4.08; SE=1.30; P=&lt;0.003). Conclusions Ultra-rapid molecular profiling reduces the time to obtaining actionable genetic sequencing results for patients hospitalized for suspected acute leukemia by 2 days compared to legacy methods. Delays in obtaining molecular profiling of these patients is associated with treatment initiation delays and longer hospital length of stay, underscoring the importance of expanded access to ultra-rapid molecular profiling for acute leukemia. Further work is needed to elucidate the full economic impact of ultra-rapid molecular profiling on overall healthcare utilization.
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Matsumoto, Shingo, Takaya Ikeda, Kiyotaka Yoh, et al. "Impact of rapid multigene assays with short turnaround time (TAT) on the development of precision medicine for non-small cell lung cancer (NSCLC)." Journal of Clinical Oncology 39, no. 15_suppl (2021): 9094. http://dx.doi.org/10.1200/jco.2021.39.15_suppl.9094.
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9094 Background: A variety of oncogene drivers have been identified in NSCLC and molecularly-stratified precision medicine has led to improved survival in advanced NSCLC. Next-generation sequencing (NGS)-based testing is utilized to detect actionable gene alterations; however, the TAT of NGS is often too long to translate into clinical decision making. Thus, rapid multi-gene testing alternatives are needed. Methods: A lung cancer genomic screening project (LC-SCRUM-Asia) capturing clinical outcome was established in 2013 to identify patients with oncogene drivers and to support the development of novel targeted therapies. Since February 2013 to May 2019 (LC-SCRUM-Asia 1st-phase), single gene testing and/or a targeted NGS assay, Oncomine Comprehensive Assay (OCA), were used for the genomic screening. Since June 2019 to December 2020 (2nd-phase), a multi-gene PCR assay (Amoy 9-in-1 test) and a rapid NGS assay (Genexus/Oncomine Precision Assay [OPA]) were also implemented as rapid multi-gene testing. Results: A total of 10667 Japanese NSCLC patients, including 6826 in the 1st-phase and 3841 in the 2nd-phase, were enrolled in the LC-SCRUM-Asia. Success rate for OCA: 93%, for 9-in-1 test: 98%, for Genexus/OPA: 96%. Median TAT for OCA: 21 days, for 9-in-1 test: 3 days, for Genexus/OPA: 4 days. The frequencies of genetic alterations detected in the 1st-/2nd-phase were EGFR: 17/24%, KRAS: 15/16%, HER2 ex20ins: 4/3%, ALK fusions: 3/3%, RET fusions: 3/2%, ROS1 fusions: 3/2%, MET ex14skip: 2/2%, BRAF V600E: 1/1%, NRG1 fusions: 0/0.2% and NTRK3 fusions: 0.05/0.04%. Overall percent agreement of 9-in-1 test compared with OCA for EGFR/KRAS/HER2/BRAF/MET/ALK/ROS1/RET/NTRK3 alterations was 98%, and that of OPA compared with OCA was 95%. The rate of patients who received targeted therapies as 1st-line treatment was significantly elevated in the 2nd-phase compared with the 1st-phase (510/3841 [13%] vs. 567/6826 [8%], p < 0.001). Through the genomic screening, 1410 (37%) and 1269 (18%) candidate patients for clinical trials of KRAS, HER2, BRAF, MET, ALK, ROS1, RET or TRK-targeted drugs were identified in the 2nd-phase and in the 1st-phase, respectively. The rate of patients who were actually enrolled into the genotype-matched clinical trials were also significantly higher in the 2nd-phase than in the 1st-phase (222 [6%] vs. 186 [3%], p < 0.001). In 1st-line treatments for advanced NSCLC patients, the median progression-free survival was 8.5 months (95% CI, 7.7−9.4) in the 2nd-phase (n = 1839) versus 6.1 months (95% CI, 5.9−6.3) in the 1st-phase (n = 4262) (p < 0.001). Conclusions: Both the 9-in-1 test and Genexus/OPA had short TATs (3−4 days), high success rates (96−98%) and good concordance (95−98%) compared with another NGS assay (OCA). These rapid multi-gene assays highly contributed to enabling precision medicine and the development of targeted therapies for advanced NSCLC.
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Siepert, Nicholas, Jeoffrey Schageman, Jian Gu, et al. "Abstract 5601: Oncomine™ dx express CE-IVD liquid biopsy assay for non-small cell lung cancer: Performance review and analytical validation." Cancer Research 83, no. 7_Supplement (2023): 5601. http://dx.doi.org/10.1158/1538-7445.am2023-5601.
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Abstract Introduction - Clinical diagnostics assays for oncology are becoming more readily available due to the advancement and democratization of Next-Generation Sequencing (NGS). Additionally, liquid biopsy can be used in NGS to detect genetic variants in circulating tumor DNA and RNA. Liquid biopsy is a less invasive option compared to traditional biopsy methods for early detection and continuous monitoring of cancer treatment outcomes. Here, we discuss screening of over 3,500 non-small cell lung cancer (NSCLC) samples, and the analytical validation of the Oncomine™ Dx Express Test (ODxET) and Genexus™ Dx Integrated Sequencer for detection of clinically significant variants in liquid biopsy samples. Methods - Our team screened over 3,500 NSCLC liquid biopsy samples in search of high priority clinical variants. Variants selected for analytical validation studies included ERBB2 exon 20 insertion, EGFR exon 20 insertion, EGFR exon 19 deletion, EGFR T790M, KRAS G12C, BRAF V600E, and RNA fusion isoforms including ALK, NTRK1/2/3, RET, and ROS1 oncogenic drivers. Over 550 screened samples were found to be positive for these variants of interest. This screening was performed on the Genexus™ Dx Integrated Sequencer according to the user guide. Results - The Genexus™ Dx Integrated Sequencer automates library preparation, sequencing, analysis, and reporting QC metrics and variant calls. Sequencing run setup is quick and straightforward, taking less than 15 minutes to start a run and just over 24 hours to go from nucleic acid to report. Limit of detection (LoD) for DNA SNVs, insertions, and deletions at 5 ng DNA input level ranged from 0.65% to 1.82% allelic frequency (AF), depending on the variant. The higher DNA input of 30 ng resulted in a lower LoD range, from 0.31% to 0.42% AF. RNA fusion and splice variant LoD at 5 ng sample input ranged from 9.9 to 19.6 molecular counts. The higher 30 ng input resulted in a lower LoD range for RNA variants as well, ranging from 6.4 to 8.0 molecular counts. In the analytical accuracy study, the false positive rate was found to be 0.2% for SNVs, 0% for insertions/deletions, and 0% for fusion targets. In the analytical reproducibility study, the average with-in run repeatability call rate (No Calls excluded) was 99.64% for DNA variants and 98.75% for RNA variants. Conclusion - Here, we demonstrated that the ODxET workflow on Genexus™ Dx Integrated Sequencer is a fast and efficient tool for testing clinical NSCLC liquid biopsy samples with high sensitivity and specificity. For in vitro diagnostic use. Not available in all regions including the United States. Citation Format: Nicholas Siepert, Jeoffrey Schageman, Jian Gu, Thilanka Jayaweera, Madhu Jasti, Luming He, Stephen Wunch, Emilia Ostrowska, Tasha Delacour, Diarra Hassell, Thomas Bowden, Daniela Garcia, Stephanie Tong, Swasti Raut, Nader Ezzedine, Elliott Martinez, Kelli Bramlett. Oncomine™ dx express CE-IVD liquid biopsy assay for non-small cell lung cancer: Performance review and analytical validation. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 5601.
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Normanno, Nicola, José Carlos Machado, Edoardo Pescarmona, et al. "European Real-World Assessment of the Clinical Validity of a CE-IVD Panel for Ultra-Fast Next-Generation Sequencing in Solid Tumors." International Journal of Molecular Sciences 24, no. 18 (2023): 13788. http://dx.doi.org/10.3390/ijms241813788.
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Molecular profiling of solid tumors facilitates personalized, targeted therapeutic interventions. The ability to perform next-generation sequencing (NGS), especially from small tissue samples, in a short turnaround time (TAT) is essential to providing results that enable rapid clinical decisions. This multicenter study evaluated the performance of a CE in vitro diagnostic (IVD) assay, the Oncomine Dx Express Test, on the Ion Torrent Genexus System for detecting DNA and RNA variants in solid tumors. Eighty-two archived formalin-fixed paraffin embedded (FFPE) tissue samples from lung, colorectal, central nervous system, melanoma, breast, gastric, thyroid, and soft tissue cancers were used to assess the presence of single nucleotide variants (SNVs), insertions and deletions (indels), copy number variations (CNVs), gene fusions, and splice variants. These clinical samples were previously characterized at the various academic centers using orthogonal methods. The Oncomine Dx Express Test showed high performance with 100% concordance with previous characterization for SNVs, indels, CNVs, gene fusions, and splice variants. SNVs and indels with allele frequencies as low as 5% were correctly identified. The test detected all the expected ALK, RET, NTRK1, and ROS1 fusion isoforms and MET exon 14-skipping splice variants. The average TAT from extracted nucleic acids to the final variant report was 18.3 h. The Oncomine Dx Express Test in combination with the Ion Torrent Genexus System is a CE-IVD-compliant, performant, and multicenter reproducible method for NGS detection of actionable biomarkers from a range of tumor samples, providing results in a short TAT that could support timely decision- making for targeted therapeutic interventions.
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Sheffield, Brandon S., Andrea Beharry, Joanne Diep, et al. "Point of Care Molecular Testing: Community-Based Rapid Next-Generation Sequencing to Support Cancer Care." Current Oncology 29, no. 3 (2022): 1326–34. http://dx.doi.org/10.3390/curroncol29030113.
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Purpose: Biomarker data are critical to the delivery of precision cancer care. The average turnaround of next-generation sequencing (NGS) reports is over 2 weeks, and in-house availability is typically limited to academic centers. Lengthy turnaround times for biomarkers can adversely affect outcomes. Traditional workflows involve moving specimens through multiple facilities. This study evaluates the feasibility of rapid comprehensive NGS using the Genexus integrated sequencer and a novel streamlined workflow in a community setting. Methods: A retrospective chart review was performed to assess the early experience and performance characteristics of a novel approach to biomarker testing at a large community center. This approach to NGS included an automated workflow utilizing the Genexus integrated sequencer, validated for clinical use. NGS testing was further integrated within a routine immunohistochemistry (IHC) service, utilizing histotechnologists to perform technical aspects of NGS, with results reported directly by anatomic pathologists. Results: Between October 2020 and October 2021, 578 solid tumor samples underwent genomic profiling. Median turnaround time for biomarker results was 3 business days (IQR: 2–5). Four hundred eighty-one (83%) of the cases were resulted in fewer than 5 business days, and 66 (11%) of the cases were resulted simultaneously with diagnosis. Tumor types included lung cancer (310), melanoma (97), and colorectal carcinoma (68), among others. NGS testing detected key driver alterations at expected prevalence rates: lung EGFR (16%), ALK (3%), RET (1%), melanoma BRAF (43%), colorectal RAS/RAF (67%), among others. Conclusion: This is the first study demonstrating clinical implementation of rapid NGS. This supports the feasibility of automated comprehensive NGS performed and interpreted in parallel with diagnostic histopathology and immunohistochemistry. This novel approach to biomarker testing offers considerable advantages to clinical cancer care.
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Eno, Celeste, Wenjuan Zhang, and Eric Vail. "27. Review and comparison of the Oncomine Myeloid Assay GX v2 on Genexus System." Cancer Genetics 268-269 (November 2022): 10. http://dx.doi.org/10.1016/j.cancergen.2022.10.030.
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Arjuna, Srividya, Mauli Shah, Antonio Dono, et al. "SDPS-44 RAPID DETECTION OF MUTATIONS IN CSF-CFTNA WITH THE GENEXUS INTEGRATED SEQUENCER." Neuro-Oncology Advances 5, Supplement_3 (2023): iii25. http://dx.doi.org/10.1093/noajnl/vdad070.098.
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Abstract Brain metastases commonly arise in patients with lung and breast carcinomas and pose distinctive clinical challenges. Genomic alterations are fundamental for diagnosis and therapy selection in cancer patients. However, the turn-around-time (TAT) of standard next generation sequencing (NGS) assays is a limiting factor in the timely delivery of genomic information for clinical decision making. The GenexusTM Sequencer automates the NGS workflow delivering results in less than 24hrs. The Oncomine Precision Assay (OPA) evaluates somatic mutations, copy number variations and fusions in hot-spot regions of 50 cancer-related genes. In this study, we evaluated genomic alterations in 46 cerebrospinal fluid (CSF) samples from 33 patients with metastatic lung cancer (mLC) (N=32) and metastatic breast cancer (mBC) (N=14) to the brain. Cell-free total nucleic acids, (cfTNA) were extracted using QIAamp Circulating Nucleic Acid kit with cfDNA concentrations ranging from 0.1-11.2(ng/ul) and 0.5-6.6 (ng/ul) for mLC and mBC samples, respectively. Median base coverage was 37,913X and 33,576.5X for mLC and mBC samples, respectively, with cfDNA input ranging from 2-20ng. Mutations were detected in 15/32 (46.8%) mLC and 8/14 (57.1%) mBC CSF samples. Among the samples with no mutations detected, 18/23 (78.2%) had suboptimal DNA input (&lt;20ng). The EGFR exon 19 deletion was detected in 4 samples from one patient, with increasing mutant allele fraction in CSF over time, highlighting the potential of CSF-cfTNA analysis for monitoring patients. Moreover, the EGFR T790M mutation was detected in a patient with prior EGFR inhibitor treatment. In addition, ESR1 D538G and ESR1::CCDC170 alterations, associated with endocrine therapy resistance, were detected in 2 mBC patients. The average TAT from cfTNA-to-results was ~16hrs. In summary, our results indicate that CSF-cfTNA analysis with the OPA in the Genexus instrument can provide clinically relevant information in patients with brain metastases with a short TAT.
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Echeverria, Delvis. "Tiempo de Respuestas y Experiencia de Usuario Estudio Experimental." Revista Latinoamericana de Ingenieria de Software 4, no. 5 (2016): 231. http://dx.doi.org/10.18294/relais.2016.231-234.
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La investigación está fundamentada por un estudio experimental, enfocado en la evaluación del comportamiento de los usuarios finales interactuando con tiempos de respuestas en <br />aplicaciones reales. Se tomará como base el aporte de los autores Doherty y Thadani, investigadores de IBM relacionado con los tiempos de respuestas, y las investigaciones de Jacob Nielsen. Se llevarán a cabo un test con usuarios finales, y se expondrán optimizaciones a través de del uso de herramientas que contribuirán a mejorar los tiempos de respuestas de las aplicaciones web. Todo esto engloba una primera instancia de un proyecto que se está llevando a cabo en la empresa GeneXus Consulting, pretendiéndose lograr un punto de encuentro entre el rendimiento de las aplicaciones y la experiencia de usuario.
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Siepert, Nicholas, Emilia Ostrowska, Luming Qu He, et al. "Abstract 5030: Evaluation of different blood collection tubes for liquid biopsy NGS applications." Cancer Research 84, no. 6_Supplement (2024): 5030. http://dx.doi.org/10.1158/1538-7445.am2024-5030.
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Abstract Intro: Liquid biopsy is fast becoming a standard practice in oncology research due to recent advancements in Next Generation Sequencing (NGS). To detect genetic variants, whole blood samples are collected in blood collection tubes (BCTs) and fractionated to obtain plasma which is then used to isolate cell free nucleic acid (cfNA) for analysis. While the limited availability of cfNA is a challenge in liquid biopsy, storage condition and duration between blood collection and fractionation have a significant impact on the quantity and quality of the cfNA due to degradation and genomic DNA and RNA contamination. There are several BCTs available on the market offering stabilization of cfNA profiles to overcome the above challenges. Methods: Here we compare the NGS performance of cfNA derived from three commonly used BCTs on the market, BD Vacutainer™ K2EDTA Tubes, Streck™ Cell-Free DNA, and PAXgene® Blood ccfDNA tubes. Matched whole blood samples collected in the above-mentioned tube types were obtained from commercial vendors which were derived from 38 healthy donors as well as 6 cancer research samples. Whole blood samples were fractionated as per manufacturer recommendations and cfNA was isolated and quantified using Genexus™ Cell-Free Total Nucleic Acid Purification Kit on Genexus™ purification system. Isolated cfNA was then sequenced using Ion Ampliseq™ HD target amplification assay on Genexus™ Integrated sequencer. Results: Data shows that K2EDTA collected samples contain slightly higher amounts of cfNA compared to PAXgene® and Streck™ samples. When the cfDNA profiles were analyzed by Agilent™ BioAnalyzer™, K2EDTA derived cfDNA showed varying degrees of genomic DNA contamination which can inflate the cfDNA concentration. However, both Streck™ and PAXgene® derived cfDNA showed little to no detectable genomic DNA contamination. As a result, a higher percentage of PAXgene® and Streck™ samples did not meet the minimum input requirement (5ng) for the sequencing assay. Sequencing data for DNA showed that K2EDTA samples had higher total reads and median molecular coverage compared to the other two BCT types. However, all samples had passing QC metrics, and mean read length, on target reads, and uniformity were comparable between the three tube types. When DNA variant detection was evaluated using cancer research samples, no significant difference was observed in performance between K2EDTA and PAXgene® samples. Interestingly, sequencing data for RNA shows that PAXgene® samples had slightly higher mapped reads on average, and significantly higher internal process control coverage compared to both K2EDTA and Streck™ samples. Whereas Streck™ samples had significantly lower internal process control coverage. Conclusion: In summary, this study demonstrates the advantages and disadvantages of using different BCTs for liquid biopsy applications, in terms of cfNA yields, isolated DNA profiles, and sequencing performance. Citation Format: Nicholas Siepert, Emilia Ostrowska, Luming Qu He, Madhuri Jasti, Kris Lea, Thilanka Jayaweera, Angie Cheng. Evaluation of different blood collection tubes for liquid biopsy NGS applications [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 5030.
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Liu, Xiaowei, Kritika Krishnamurthy, and D. Yitzchak Goldstein. "P692: Impact of sample age on RNA sequencing metrics from FFPE tissues using ion torrent Genexus Integrated Sequencer." Genetics in Medicine Open 3 (2025): 103061. https://doi.org/10.1016/j.gimo.2025.103061.
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Buglioni, S., E. Gallo, C. Ercolani, et al. "Clinical Feasibility of NGS Liquid Biopsy Analysis in NSCLC Patients Using Oncomine Precision Assay with Genexus™ Integrated Sequencer." Journal of Liquid Biopsy 1 (November 2023): 100017. http://dx.doi.org/10.1016/j.jlb.2023.100017.
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Liu, Xiaowei, Kritika Krishnamurthy, and D. Yitzchak Goldstein. "P744: Comparative analysis of Ion Torrent sequencing platforms: Unveiling enhanced performance and precision with the Genexus integrated sequencer in clinical applications." Genetics in Medicine Open 2 (2024): 101648. http://dx.doi.org/10.1016/j.gimo.2024.101648.
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Ilié, Marius, Jonathan Benzaquen, Véronique Hofman, et al. "Accurate Detection of SARS-CoV-2 by Next-Generation Sequencing in Low Viral Load Specimens." International Journal of Molecular Sciences 24, no. 4 (2023): 3478. http://dx.doi.org/10.3390/ijms24043478.
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As new SARS-CoV-2 variants emerge, there is an urgent need to increase the efficiency and availability of viral genome sequencing, notably to detect the lineage in samples with a low viral load. SARS-CoV-2 genome next-generation sequencing (NGS) was performed retrospectively in a single center on 175 positive samples from individuals. An automated workflow used the Ion AmpliSeq SARS-CoV-2 Insight Research Assay on the Genexus Sequencer. All samples were collected in the metropolitan area of the city of Nice (France) over a period of 32 weeks (from 19 July 2021 to 11 February 2022). In total, 76% of cases were identified with a low viral load (Ct ≥ 32, and ≤200 copies/µL). The NGS analysis was successful in 91% of cases, among which 57% of cases harbored the Delta variant, and 34% the Omicron BA.1.1 variant. Only 9% of cases had unreadable sequences. There was no significant difference in the viral load in patients infected with the Omicron variant compared to the Delta variant (Ct values, p = 0.0507; copy number, p = 0.252). We show that the NGS analysis of the SARS-CoV-2 genome provides reliable detection of the Delta and Omicron SARS-CoV-2 variants in low viral load samples.
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Vivancos Sánchez, Catalina, María Isabel Esteban Rodríguez, Alberto Peláez García, et al. "Clinical Impact of a Next-Generation Sequencing Approach for Glioblastoma Patients." Cancers 17, no. 5 (2025): 744. https://doi.org/10.3390/cancers17050744.
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Objectives: The purpose of this study is to assess the clinical impact of next-generation sequencing (NGS), as an increasingly available and advantageous tool, for glioblastoma patients. Methods: Adult patients aged less than 65, and surgically treated for glioblastoma between 2010–2021, were included. Tumor samples were analyzed with NGS using the Oncomine Comprehensive v3 (OCA) panel and Ion Reporter Genexus v5.9.1 (Thermo Fisher Scientific). Results: Thirty-two patients were included, with a median age of 47.7 years and a median overall survival of 25 months. Identification of mutations by NGS resulted in a change in diagnosis in two cases. In all patients but one, at least one genetic alteration was detected (median of three per patient), most commonly EGFR amplification. In 93.7% of patients, biomarkers that make them potentially eligible for a clinical trial were found. No survival differences were seen regarding genetic alterations, although a trend towards better survival for those patients without CDK4 mutation was observed (p = 0.088). Conclusions: The use of NGS provides useful information for diagnosis, especially in young patients, and it will probably become valuable for clinical decision-making as more therapeutic targets and treatments emerge. For the moment, it is crucial for scientific progress to happen.
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Andrew, Georgia M., Laura W. Dillon, Maria Saeed, et al. "Evaluation of a Rapid Automated Next Generation Sequencing Assay for Precision Medicine in Acute Myeloid Leukemia." Blood 138, Supplement 1 (2021): 4444. http://dx.doi.org/10.1182/blood-2021-148926.
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Abstract Background: In the last decade there have been significant advances in diagnosing and classifying adult acute myeloid leukemia (AML) based on genomic profiling, enabling risk-stratification and targeted therapies. In 2017 the US FDA approved the first gene mutation targeted therapies for AML with multiple additional targeted therapies since approved or in development. Given the typical acuity of AML at initial presentation however and the current turnaround time for next-generation sequencing (NGS) assays, most patients will start definitive initial therapy before all potentially targetable mutations are known. There is, therefore, a significant need for a fast molecular genotyping test to determine eligibility for personalized therapy in AML. The NCI Myeloid Assay (NMA) is a comprehensive targeted NGS assay on the Ion Torrent Genexus System, a fully automated platform that provides a rapid turnaround time from specimen receipt to clinical reporting. NMA utilizes Thermo Fisher Scientific's Oncomine Myeloid Assay GX and appears ideally suited for use in upcoming AML targeted therapy trials but has yet to be extensively tested in a cohort of AML patient diagnostic samples and compared to a standard targeted "myeloid panel" NGS assay platform (s-NGS). Methods: DNA samples (n=173) extracted from pretreatment bone marrow and/or peripheral blood of adult patients (n=112) diagnosed with de novo AML or high-risk myelodysplastic syndrome (MDS), were blindly tested in parallel using the NMA and s-NGS assays. For the NMA assay, 27.75ng of DNA was put into the Genexus System. All runs, controls, and samples were first analyzed for sequencing quality using established quality control (QC) metrics to assess pass/fail status. For all samples that passed QC metrics, variant results generated by the Ion Torrent Genexus pipeline were manually reviewed prior to being called true positive variants. For the s-NGS, using the ArcherDx Myeloid VariantPlex assay, a DNA input of 50ng was used for library preparation on a dual pre- and post-PCR separated automated liquid-handling workflow. Resulting libraries were sequencing on the Novaseq 6000 (Illumina) and the data analyzed using the Archer Analysis software and filtered as previously described (PMID: 34258102). Results from the two assays were compared for mutations with a variant allele fraction (VAF) &gt;5% occurring in genes of interest in small molecule targeted clinical trials including: FLT3, IDH1, IDH2, JAK2, KIT, NPM1, NRAS, KRAS, and TP53. For FLT3-ITD comparison, the presence or absence of a call by the assay was used. Results: Utilizing a 5% VAF reporting threshold, a total of 171 and 174 variants were detected by NMA and s-NGS assays, respectively. A high rate of concordance was observed between the assays, with NMA detecting 96% of s-NGS variants and s-NGS detecting 95% of NMA variants. The VAF of detected single nucleotide variants was highly correlated (r=0.9848, P&lt;0.0001, Figure 1A). NPM1 mutation VAF values trended lower by s-NGS compared to NMA. We investigated the discordant calls (n=15 total in 11 patients). One patient was correctly identified as having an NRAS p.Gly12 mutation by both approaches, but the resulting mutation was incorrectly annotated by the s-NGS pipeline. Samples from two patients (including one with both blood and marrow tested) were correctly identified as being FLT3 tyrosine kinase domain mutated by both sequencing approaches, although only the major of two missense variants identified by s-NGS was reported by the NMA pipeline. None of these patients, however, would be misclassified. The remaining 11 discordant calls were false negatives (including 6 variants detected by s-NGS but not by NMA). All of these "edge case" variants were detectable by lowering the VAF reporting threshold below 5% (Figure 1B). Conclusions: NMA is an automated sample-to-results workflow that can identify myeloid disorder-associated genomic variants in less than 48 hours from library preparation to clinical reporting. We show that NMA is highly concordant with a standard DNA NGS assay for detecting mutations within recurrently mutated AML genes. Accurate rapid genotyping is required for assignment to initial treatment with targeted therapy, and this technology may be a valuable tool for upcoming clinical trials for patients with myeloid malignancies. Figure 1 Figure 1. Disclosures Zhang: Thermo Fisher Scientific: Current Employment. Sedova: Thermo Fisher Scientific: Current Employment. Huang: Thermo Fisher Scientific: Current Employment. Mittal: Thermo Fisher Scientific: Current Employment. Hatch: Thermo Fisher Scientific: Current Employment. Ni: Thermo Fisher Scientific: Current Employment. Kaznadzey: Thermo Fisher Scientific: Current Employment. Sadis: Thermo Fisher Scientific: Current Employment. Smith: Thermo Fisher Scientific: Current Employment. Williams: Illumina: Other: CRADA. Hourigan: Sellas: Research Funding.
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Frady, Lauren, Desiree Unselt, John Pufky, Fernando Torres, Jennifer Sims, and Jennifer Mason. "Abstract 528: Automated liquid biopsy pan-cancer genomic profiling panel demonstrates utility for screening of targeted biomarkers." Cancer Research 82, no. 12_Supplement (2022): 528. http://dx.doi.org/10.1158/1538-7445.am2022-528.
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Abstract Background: Liquid biopsy testing has demonstrated the utility of capturing the heterogeneity of solid tumors and relevant actionable mutations using a less invasive sample collection method. The Genexus™ Integrated Sequencer provides a viable pathway to deploy an automated platform for a sensitive pan-cancer liquid biopsy assay to meet the needs of quick turn-around and cost effectiveness for multi-site implementation. The Oncomine™ Precision Assay GX (OPAGX) is an amplicon-based next-generation sequencing (NGS) assay that enables detection of biomarkers across 50 genes from both solid tissue and liquid biopsy samples. OPAGX uses AmpliSeq™ HD technology which includes unique molecular tags for sequencing error correction and increased sensitivity. The assay’s detection of hotspot mutations was evaluated for liquid biopsy samples. Methods: Automated library preparation was completed on the Genexus™ Integrated Sequencer using 20 ng of cfTNA derived from plasma or from well characterized control samples (Acrometrix, Agilent, Horizon, and SeraSeq). After reverse transcription was completed, gene targets of interest were amplified from DNA and cDNA. Amplicons were labeled with dual molecular barcodes during library preparation. Subsequently, libraries were bead normalized, pooled, and sequenced on a GX5™ chip to a targeted depth of 12,000,000 reads per sample. Sensitivity and positive predictive value were examined by evaluating the correctness of calling true positive variants compared to false negative and false positive variant calls, respectively. Precision and limit of detection (LoD) were examined by evaluating the concordance of variants across replicate samples with the range of AF between 0.1% - 35% for SNVs and 0.2% - 15% for insertions and deletions. Specificity and accuracy were examined by evaluating the correctness of calling true negative variants compared to false positive and all other variant calls, respectively. Results: Accuracy was 98% for SNVs and 100% for insertions and deletions above 0.25% allele frequency (AF). Sensitivity was 95% for SNVs and 81% for insertions and deletions above 0.5% AF. Specificity and PPV were 100% for SNVs and insertions and deletions above 0.1% AF. Intra-run precision was 95% for SNVs and 92% for insertions and deletions above 0.5% and 0.25% AF, respectively. Inter-run precision was 93% for SNVs and 76% for insertions and deletions above 0.5% and 0.25% AF, respectively. The estimated AF LoD was 0.6% for SNVs, which is slightly above the advertised 0.5% provided by the vendor. Additionally, CNV gain calls were 100% for ERBB2 above 0.25% AF, MET above 0.5% AF, and FGFR3 at all AFs. Conclusions: The demonstrated performance of OPAGX presents a viable liquid biopsy platform which lays the foundation for global standardization of genomic profiling liquid biopsy samples with minimal nucleic acid input. Citation Format: Lauren Frady, Desiree Unselt, John Pufky, Fernando Torres, Jennifer Sims, Jennifer Mason. Automated liquid biopsy pan-cancer genomic profiling panel demonstrates utility for screening of targeted biomarkers [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 528.
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Unselt, Desiree, Lauren Frady, John Pufky, Fernando Torres, Jennifer Sims, and Jennifer Mason. "Abstract 2290: Quick turnaround NGS assay for the characterization of biomarkers in FFPE tissue." Cancer Research 82, no. 12_Supplement (2022): 2290. http://dx.doi.org/10.1158/1538-7445.am2022-2290.
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Abstract Background: Genomic profiling of a tumor to characterize clinically relevant variants has become critically important for cancer diagnosis and prognosis, as well as in driving therapeutic decision making. As such, there is a growing need for rapid inline assessment of somatic variants in genes of key interest in oncology. The Oncomine™ Precision Assay on the Ion Torrent Genexus Sequencer (OPAGX) is an amplicon-based automated next-generation sequencing (NGS) assay that enables quick turnaround genomic profiling of biomarkers across 50 genes from solid tissue. The complete workflow, including library prep, templating, sequencing, and final report generation, can be completed in less than 30 hours for up to 32 formalin-fixed, paraffin-embedded (FFPE) samples. This assay uses AmpliSeq™ HD technology which includes unique molecular tags for sequencing error correction and increased sensitivity for low frequency variant detection. The assay’s detection of hotspot mutations (substitutions, insertions, deletions, copy number variants (CNVs), and fusions) was evaluated for FFPE tissue. Method: Fully automated library preparation and sequencing was completed in the Genexus™ Integrated Sequencer using 10 ng of DNA derived from FFPE tissue or from characterized control samples. Following gene specific amplification, AmpliSeq™ HD technology is used to label amplicons with dual molecular barcodes. The resultant libraries are bead normalized, pooled, and sequenced on a GX5™ chip to a targeted depth of 3,000,000 reads per sample. Sensitivity and positive predictive value were examined by evaluating the correctness of calling true positive variants compared to false negative and false positive variant calls, respectively. Precision and limit of detection (LoD) were examined by evaluating the concordance of variants across replicate control samples. Specificity and accuracy were examined by evaluating the correctness of calling true negative variants compared to false positive and all other variant calls, respectively. Results: Accuracy and specificity were 99.9% for SNVs and 100% for insertions and deletions above 1% allele frequency (AF). Sensitivity and intra-run precision were 100% for SNVs and insertions and deletions above 1% and 2% AF, respectively. Inter-run precision and PPV were 100% for SNVs and insertions and deletions above 1% AF. The estimated AF LoD was 3.7% for SNVs. CNV gain calls were 100% for MET and EGFR and loss calls were 100% for PTEN (homozygous and heterozygous) and CDKN2A. Additionally, fusion calling for RET and ROS1 were 100%. Conclusions: OPAGX is a quick turnaround NGS assay enabling precision medicine through automated multi-variant genomic profiling to inform decision making; performance was characterized with analysis of hotspot mutations in FFPE samples in a variety of tumor types, demonstrating utility of implementation in testing of targeted biomarkers across multiple indications. Citation Format: Desiree Unselt, Lauren Frady, John Pufky, Fernando Torres, Jennifer Sims, Jennifer Mason. Quick turnaround NGS assay for the characterization of biomarkers in FFPE tissue [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 2290.
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Heeke, Simon, Dzifa Duose, Srividya Arjuna, et al. "Abstract 3776: Association of baseline ctDNA EGFR mutation detection with clinical outcome in the phase II RAMOSE trial assessing ramucirumab plus osimertinib versus osimertinib in EGFR mutant non-small cell lung cancer." Cancer Research 85, no. 8_Supplement_1 (2025): 3776. https://doi.org/10.1158/1538-7445.am2025-3776.
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Abstract Background: While third-generation EGFR TKIs such as osimertinib have improved outcomes in EGFR-mutant non-small cell lung cancer (NSCLC), responses remain transient, and resistance develops often. We recently reported primary efficacy results from the phase II RAMOSE trial showing that the combination of osimertinib with the VEGFR2 antagonist ramucirumab improved progression-free survival (PFS) over osimertinib monotherapy (24.8 months [mo] vs 15.6 mo, HR 0.55 (95% CI, 0.32-0.93, p=0.023), establishing this regimen as a promising approach to improve clinical outcomes. However, biomarker strategies are needed to guide optimal treatment decisions and allow for personalized therapy stratification. Therefore, we explore EGFR mutation detection from blood plasma at baseline in patients enrolled in the RAMOSE trial. Methods: The randomized, open-label multicenter phase II RAMOSE trial (NCT03909334, HCRN LUN-18-335) compared osimertinib with ramucirumab to osimertinib for frontline treatment of metastatic EGFR-mutant NSCLC. Blood plasma was collected at baseline (BL). Circulating-tumor DNA (ctDNA) for analysis was isolated from up to 4ml of blood plasma using the Genexus purification system. EGFR mutations were analyzed using the Oncomine Precision Assay on the Ion Torrent Genexus System. EGFR mutations were assessed by variant allele frequency (VAF) and correlated to clinical outcomes, including PFS and OS. Results: Eight-six patient samples were sequenced, and 81 (94%) yielded positive sequencing results at BL. No statistically significant differences in BL EGFR detection were detected between the two treatment arms (χ2 p = 0.33). EGFR mutations at any VAF were detected in 68/81 patients (84%). Patients with EGFR VAF &lt; 0.5% had significantly longer PFS (17.9mo vs not reached; p = 0.016; HR = 0.37) and OS (33.4mo vs not reached; p = 0.028; HR = 0.14) compared to patients with VAF &gt; 0.5%. In patients without detectable EGFR mutations at BL, no death occurred during median follow up of 21.4mo (OS p = 0.076). In patients with detectable EGFR mutation at BL, the PFS was 19.9 mo in combination arm versus 16.0 mo in the osimertinib arm (HR = 0.69, p = 0.3). Conclusion: Detection of EGFR mutations from blood plasma at baseline was prognostic of clinical outcome and associated with significantly shorter PFS and OS. Detection of EGFR VAF by liquid biopsy can identify patients with higher risk of treatment progression and warrants further investigation for stratification of patients to future clinical trials. Citation Format: Simon Heeke, Dzifa Duose, Srividya Arjuna, Monique Nilsson, Jyoti Patel, Elaine Shum, Christina Baik, Rachel Sanborn, Catherine Shu, Chul Kim, Mary Jo Fidler, Richard Hall, Jhanelle Gray, Andreas Saltos, John V. Heymach, Xiuning Le. Association of baseline ctDNA EGFR mutation detection with clinical outcome in the phase II RAMOSE trial assessing ramucirumab plus osimertinib versus osimertinib in EGFR mutant non-small cell lung cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 3776.
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Guo, Fei, Yubo Lang, Guannan Long, et al. "Ion Torrent ™ Genexus ™ Integrated Sequencer and ForeNGS Analysis Software—An automatic NGS-STR workflow from DNA to profile for forensic science." Forensic Science International: Genetics 61 (November 2022): 102753. http://dx.doi.org/10.1016/j.fsigen.2022.102753.
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Pickle, Loni, Andrew Hatch, Gokhan Yavas, et al. "Abstract 4416: Rapid, low-input, targeted NGS workflow for DNA methylation." Cancer Research 84, no. 6_Supplement (2024): 4416. http://dx.doi.org/10.1158/1538-7445.am2024-4416.
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Abstract DNA methylation is a fundamental epigenetic process that regulates human gene expression. In cancer, methylation changes promote proliferation networks and metastasis. Development of biomarkers for methylation will be enabled by flexible, fast, low-input next generation sequencing workflows. We describe the first Ion AmpliSeq™ Methylation targeted panel and protocol on the turn-key Genexus™ Integrated Sequencer as part an ongoing collaboration with Ontario Institute for Cancer Research (OICR) to detect and predict response in early stage breast cancer and improve diagnostics for Black and Asian women. A 2-pool Ion AmpliSeq™ Methylation Panel for Breast Cancer Research was developed as a demonstration of design, workflow, and reporting for targeted, low-input methylation assessment in a multiplex setting across a variety of samples sources including FFPE. The panel contains 327 amplicons and was designed to target both strands and 10ng DNA input into bisulfite conversion was used for controls. A complete workflow begins at bisulfite conversion and progresses through the Genexus™ Integrated Sequencer which combines library construction, template preparation, and sequencing into a single run. The bioinformatics pipeline provides DNA methylation calls on both Watson and Crick strands at single base resolution and methylated:unmethylated ratios for each CpG. The entire end-to-end workflow was completed in 2 days, including a full analysis software solution. The panel was evaluated using 2 control gDNA samples. The first had an expected average methylation state across all CpGs of &gt;98% and the second &lt;5%. An equal mixture of these two samples was also tested. The Methylation Panel for Cancer Research performed well on control samples. The single lane Loading, Total Bases, Final Reads, and Raw Read Accuracy were &gt;92%, &gt;1G, &gt;14 Million, and &gt;90% respectively. The workflow was also demonstrated on research FFPE cancer samples. This Methylation Panel protocol offers a 2-day, end-to-end workflow with high resolution, targeted and quantitative methylation analysis from DNA input as low as 10ng into bisulfite conversion. The option to design custom methylation panels for interrogation of targets of interest without the need for whole genome methylation is now available. &lt;For Research Use Only. Not for use in diagnostic procedures. Early access materials were provided by Thermo Fisher Scientific™ to OICR in support of this abstract, but no other financial support. This study was conducted with the support of the Ontario Institute for Cancer Research through funding provided by the Government of Ontario. © 2023 Thermo Fisher Scientific Inc. All rights reserved. All trademarks are the property of Thermo Fisher Scientific and its subsidiaries unless otherwise specified.&gt; Citation Format: Loni Pickle, Andrew Hatch, Gokhan Yavas, Melanie Spears, Louis Gasparini, Vida Talebian, Anna Ying-Wah Lee, Mathieu Lariviere, Jane Bayani, Seth Sadis, Jeffrey M. Smith. Rapid, low-input, targeted NGS workflow for DNA methylation [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 4416.
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Huang, Jiajie, Haigang Gu, Janet Orton, et al. "Abstract 328: A comprehensive genomic profiling of myeloid malignancies demonstrates mutational spectrum of DNA variants, FLT3-ITDs, and gene fusions." Cancer Research 84, no. 6_Supplement (2024): 328. http://dx.doi.org/10.1158/1538-7445.am2024-328.
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Abstract Introduction: Myeloid malignancies encompass diverse hematopoietic disorders, including acute myeloid leukemia (AML), myeloproliferative neoplasms (MPN), myelodysplastic syndromes (MDS), chronic myeloid leukemia (CML), chronic myelomonocytic leukemia (CMML), and juvenile myelomonocytic leukemia (JMML). We developed two assays to address the genetic complexity of myeloid malignancies, OncomineTM Myeloid Assay and OncomineTM Myeloid Assay GX v2, detecting mutations in 45 DNA genes and &gt;30 fusion driver genes, including &gt;700 fusion isoforms. Our panel includes genetic alterations such as FLT3-Internal Tandem Duplications (ITDs), IDH1/2, CEBPA, CALR, and TP53. Methods: Here we describe the genomic profiles of 8503 clinical research samples (including AML, MPN, MDS, CML, CMML, or JMML). A total of 4723 samples were run on the Ion GeneStudioTM S5 System and analyzed using the OncomineTM Myeloid Research workflow on Ion ReporterTM 5.18. A total of 3780 samples were run on the Ion TorrentTM Genexus Software 6.6 and analyzed using the OncomineTM Myeloid Assay GX v2. Results: On Genexus, the turnaround time (the time between starting the run and NGS data report) was 23-25 hours and the hands-on time was around 1 hour. The success rate of the samples was 100%. Frequency of relevant mutations and variant allele frequency by gene: Genes like TET2 (detected in 12.6% of samples), ASXL1 (9.3%), DNMT3A (7.8%), TP53 (7.5%) exhibited high mutation rates. 56.4% of samples had mutations, averaging 2.3 mutations per sample. Common co-occurring mutations included ASXL1-TET2 and SRSF2-TET2. Variant allele frequency varied greatly, e.g., ANKRD26 had a median VAF of 66%, while MYD88 had only 7%. Mutation spectrum of FLT3ITD variants: FLT3-ITD was observed in 3% of samples, with an average VAF of 0.32, featuring a multi-modal length with the highest peak at around 34 bp and maximum length at 151 bp. Mutation spectrums of fusions: 710 (8.4%) samples were fusion-positive with read counts ranging 21-113,000. ABL1 was the most common driver and BCR-ABL1 was the most common gene pair. Other common drivers included KMT2A, MYH11, RARA, and RUNX1. 17 distinct driver genes, 49 gene pairs, and 110 gene isoforms were observed, including rare fusions like ZMYM2-FGFR1 and KAT6A-CREBBP that are not detectable by traditional methods like qPCR (quantitative polymerase chain reaction) or FISH (fluorescence in-situ hybridization). Conclusions: The Oncomine Myeloid Assay is a fast, robust, and reproducible solution for comprehensive genomic profiling of myeloid malignancies. We describe the mutational spectrum of DNA variants and RNA fusions in a range of clinical research samples. (For research use only. Not for use in diagnostic procedures. © 2023 Thermo Fisher Scientific Inc. All rights reserved. All trademarks are the property of Thermo Fisher Scientific and its subsidiaries unless otherwise specified.) Citation Format: Jiajie Huang, Haigang Gu, Janet Orton, Marina Sedova, Sophie Rozenzhak, Fiona Hyland, Guang Liu. A comprehensive genomic profiling of myeloid malignancies demonstrates mutational spectrum of DNA variants, FLT3-ITDs, and gene fusions [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 328.
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Ilié, Marius, Véronique Hofman, Christophe Bontoux, et al. "Setting Up an Ultra-Fast Next-Generation Sequencing Approach as Reflex Testing at Diagnosis of Non-Squamous Non-Small Cell Lung Cancer; Experience of a Single Center (LPCE, Nice, France)." Cancers 14, no. 9 (2022): 2258. http://dx.doi.org/10.3390/cancers14092258.
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The number of genomic alterations required for targeted therapy of non-squamous non-small cell lung cancer (NS-NSCLC) patients has increased and become more complex these last few years. These molecular abnormalities lead to treatment that provides improvement in overall survival for certain patients. However, these treated tumors inexorably develop mechanisms of resistance, some of which can be targeted with new therapies. The characterization of the genomic alterations needs to be performed in a short turnaround time (TAT), as indicated by the international guidelines. The origin of the tissue biopsies used for the analyses is diverse, but their size is progressively decreasing due to the development of less invasive methods. In this respect, the pathologists are facing a number of different challenges requiring them to set up efficient molecular technologies while maintaining a strategy that allows rapid diagnosis. We report here our experience concerning the development of an optimal workflow for genomic alteration assessment as reflex testing in routine clinical practice at diagnosis for NS-NSCLC patients by using an ultra-fast-next generation sequencing approach (Ion Torrent Genexus Sequencer, Thermo Fisher Scientific). We show that the molecular targets currently available to personalized medicine in thoracic oncology can be identified using this system in an appropriate TAT, notably when only a small amount of nucleic acids is available. We discuss the new challenges and the perspectives of using such an ultra-fast NGS in daily practice.
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Gaal, Orsolya I., Andrei Ungureanu, Bogdan Pop, et al. "The Genomic Landscape of Romanian Non-Small Cell Lung Cancer Patients: The Insights from Routine NGS Testing with the Oncomine Dx Target Panel at the PATHOS Molecular Pathology Laboratory." Cancers 17, no. 12 (2025): 1947. https://doi.org/10.3390/cancers17121947.
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Background: Comprehensive molecular profiling is essential for precision oncology in non-small cell lung cancer (NSCLC). However, genomic data from Eastern European populations, including Romania, remain limited. Methods: We analyzed 398 consecutive NSCLC cases tested at the PATHOS Molecular Pathology Laboratory (Cluj-Napoca, Romania) between April 2024 and February 2025 using the Ion Torrent™ Genexus™ System and the Oncomine™ Dx Target Test, which evaluates SNVs/indels in 46 genes, fusions in 23 genes, and CNVs in 19 genes from FFPE samples. Results: The cohort was predominantly male (66%) with a median age of 67 years. Adenocarcinoma represented 70% of cases with known histology. Genomic profiling revealed a high frequency of actionable alterations. KRAS mutations were the most common (29.1%), with p.G12C detected in 10.3% of all the cases. EGFR mutations were present in 14.3% of patients, mostly exon 19 deletions and L858R substitutions. BRAF alterations (5.3%) included both V600E and non-V600E variants. RET alterations were detected as eight missense mutations, two canonical fusions (KIF5B–RET, CCDC6–RET), one amplification, and three transcript imbalances. EML4-ALK fusions (1.77%), ERBB2 mutations/amplifications (3.0%), and FGFR1/FGFR3 amplifications were also observed. Conclusions: This study provides the first large-scale molecular snapshot of NSCLC in Romania. While the overall genomic profiles align with Western populations, the higher frequency of KRAS p.G12C and FGFR amplifications highlights the value of region-specific data to support targeted therapies in Eastern Europe.
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Colac, Svetlana, Olga Burduniuc, and Mariana Apostol. "GENETIC SIGNIFICANCE AND TRACKING OF CIRCULATING SARS-COV-2 VARIANTS IN THE REPUBLIC OF MOLDOVA." Romanian Archives of Microbiology and Immunology 83, no. 3 (2024): 148–54. https://doi.org/10.54044/rami.2024.03.02.
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Tracking the circulating variants of the SARS-CoV-2 virus and identifying new genetic variations provide a high-resolution perspective with many advanced applications, ranging from developing diagnostic tests and vaccines to real-time data for public health measures. This study aimed to investigate and assess the genetic variants of SARS-CoV-2 circulating in the Republic of Moldova to improve COVID-19 response strategies. A retrospective descriptive study was carried out in the National Public Health Agency virology laboratory. The study analysed 417 biological samples collected from patients infected with COVID-19, with SARS-CoV-2 presence confirmed by molecular biology techniques (Reverse transcription polymerase chain reaction (RTPCR)). Viral fragment sequencing was performed via the Ion Torrent Genexus sequencer, Pangolin, and GISAID (Global Initiative on Sharing All Influenza Data) software. The Omicron variant underwent multiple genetic changes throughout 2022. Thus, these changes were divided into four distinct periods of SARS-CoV-2 genome evolution. The first period (January–March) was characterized by the predominance of BA.1* and its subvariants; the second period (March–June) by the predominance of the BA.2* sublineage and its subvariants; the third period (June–October) by the predominance of BA.4*/BA.5* and their subvariants; and, finally, in the fourth-period various sublineages, including BQ.1*, BN.1*, CC.1*, and XBB.1.5, became predominant. In conclusion, the implementation of advanced SARS-CoV-2 genome sequencing technologies in the Republic of Moldova has proven the importance of detecting novel viral mutations and their impact on the epidemic progression of the COVID-19 infection.
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Nicholas, Champica, Andrea Beharry, Anna M. Bendzsak, et al. "Point of Care Liquid Biopsy for Cancer Treatment—Early Experience from a Community Center." Cancers 16, no. 14 (2024): 2505. http://dx.doi.org/10.3390/cancers16142505.
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Liquid biopsy is rapidly becoming an indispensable tool in the oncologist’s arsenal; however, this technique remains elusive in a publicly funded healthcare system, and real-world evidence is needed to demonstrate utility and feasibility. Here, we describe the first experience of an in-house point of care liquid biopsy program at a Canadian community hospital. A retrospective review of consecutive cases that underwent plasma-based next-generation sequencing (NGS) was conducted. Liquid biopsy was initiated at the discretion of clinicians. Sequencing followed a point of care workflow using the Genexus™ integrated sequencer and the Oncomine precision assay, performed by histotechnologists. Results were reported by the attending pathologist. Eligible charts were reviewed for outcomes of interest, including the intent of the liquid biopsy, results of the liquid biopsy, and turnaround time from blood draw to results available. A total of 124 cases, with confirmed or suspected cancer, underwent liquid biopsy between January 2021 and November 2023. The median turnaround time for liquid biopsy results was 3 business days (range 1–12 days). The sensitivity of liquid biopsies was 71%, compared to tissue testing in cases with matched tissue results available for comparison. Common mutations included EGFR (29%), in 86 lung cancer patients, and PIK3CA (22%), identified in 13 breast cancer patients. Healthcare providers ordered liquid biopsies to inform diagnostic investigations and treatment decisions, and to determine progression or resistance mechanisms, as these reasons often overlapped. This study demonstrates that rapid in-house liquid biopsy using point of care methodology is feasible. The technique facilitates precision treatment and offers many additional advantages for cancer care.
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Kropf, Jacqueline, Elim Kuo, Jorge Cusco, Jennifer Tseng, Amy Iarrobino Laughlin, and Wang L. Cheung. "Impact of tissue biomarker testing by in-house platform on result times in metastatic non-small cell lung cancer." JCO Oncology Practice 19, no. 11_suppl (2023): 49. http://dx.doi.org/10.1200/op.2023.19.11_suppl.49.
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49 Background: Biomarker testing for actionable mutations is standard of care for patients diagnosed with metastatic non-small cell, non-squamous cell lung cancer. However, testing via third-party laboratories can take weeks to result, leading to treatment decision delays identified at our own institution. Methods: We conducted a retrospective chart review of metastatic non-small cell, non-squamous cell lung cancer patients treated at Orlando Regional Medical Center before (6/1/2021-5/31/2022) as well as after (1/1/2023-5/31/2023) the initiation of in-house next generation sequencing (NGS) tissue testing using Genexus. Endpoints included average and median time to result of plasma and tissue NGS both before and after initiation of in-house molecular testing. An independent two sample t-test was conducted with p<0.05 considered significant. Results: A total of 49 and 18 patients were included pre and post initiation of in house NGS, respectively. For patients with tissue testing, the average time from collection to result was 31.3 versus 13.9 days (p<0.01), median 29 versus 7 days. For patients who underwent plasma testing (n=49, n=10), average time from collection to result was 13.3 versus 9.6 days (p=0.38), median 9 versus 9 days. Conclusions: Time to comprehensive tissue testing result significantly decreased after the commencement of in-house NGS. Rapid turnaround time allows clinicians to select appropriate first-line treatment without delay in care. Further efforts are needed to expedite molecular testing in all stages of non-small cell lung cancer and evaluate sequential versus concurrent plasma and tissue testing.
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Huang, Ping, Chia Chou, and Ronghui Huang. "Seasonal Modulation of Tropical Intraseasonal Oscillations on Tropical Cyclone Geneses in the Western North Pacific." Journal of Climate 24, no. 24 (2011): 6339–52. http://dx.doi.org/10.1175/2011jcli4200.1.
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Abstract The seasonal modulation of tropical intraseasonal oscillation (TISO) on tropical cyclone (TC) geneses over the western North Pacific Ocean (WNP) is investigated in three periods of the WNP TC season: May–June (MJ), July–September (JAS), and October–December (OND). The modulation of the TISO–TC geneses over the WNP is strong in MJ, while it appears weaker in JAS and OND. In MJ, TISO propagates northward via two routes, the west route through the South China Sea and the east route through the WNP monsoon trough region, which are two clustering locations of TC geneses. TISO can synchronously influence most TC geneses over these two regions. In JAS, however, the modulation is out of phase between the monsoon trough region and the East Asian summer monsoon region, as well as the WNP subtropical high region, as a result of further northward propagation of TISO and scattered TC geneses. The TISO–TC genesis modulation in each individual region is comparable to that in MJ, although the modulation over the entire WNP in JAS appears weaker. In OND, TISO has a stronger influence on TC geneses west than east of 150°E because TISO decays and its convection center located at the equator is out of the TC genesis region when propagating eastward into east of 150°E. Midlevel relative humidity is the primary contribution to the modulations of TISO on the genesis environment, while vorticity could contribute to the modulation over the subtropics in JAS.
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Chen, Tsing-Chang, Jenq-Dar Tsay, Jun Matsumoto, and Jordan Alpert. "Impact of the Summer Monsoon Westerlies on the South China Sea Tropical Cyclone Genesis in May." Weather and Forecasting 32, no. 3 (2017): 925–47. http://dx.doi.org/10.1175/waf-d-16-0189.1.
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Abstract After the onset of the Southeast Asian summer monsoon in mid-May, the South China Sea (SCS) trough is deepened by the intensified monsoon westerlies to facilitate the development of a synoptic cyclonic shear flow. This shear flow forms an environment favorable for the SCS tropical storm (TS)/typhoon (TY) genesis triggered by the surge of this monsoon circulation. This genesis mechanism has not been well documented. Seventeen named SCS TS/TY geneses in May over 1979–2016 occurred under the following environmental conditions/processes: 1) with its maximum located south of 15°N, the intensified monsoon westerlies are extended eastward beyond 120°E, 2) the synoptic SCS cyclonic shear flow is developed by the tropical easterlies fed by a northeast Asian cold surge (or a North Pacific cold-air outbreak) and the intensified monsoon westerlies, and 3) SCS TS/TY genesis is triggered by the surge of monsoon flow. The accuracy of the monthly mean forecasts is limited. However, it is found that SCS TS/TY genesis only occurs after the existence of persistent, strong, monsoon westerlies lasting for at least 5 days. Forecasts from the National Centers for Environmental Prediction Global Forecast System (2004–16) and the Global Ensemble Forecast System (1985–2003) cover these 15 SCS TS/TY geneses. The requirements for SCS TS/TY genesis in May described above are met by the 5-day-mean Southeast Asian summer monsoon circulation. Based on a statistical analysis of 5-day forecasts for these TS/TY geneses, a four-step forecast advisory is introduced. The forecasts for SCS TS/TY genesis can be made 3 days prior to occurrence.
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Raskin, William, Parneet Cheema, Kirstin Perdrizet, Marco Iafolla, Shaan Dudani, and Brandon Sheffield. "Rapid point of care NGS in colorectal cancer." Journal of Clinical Oncology 40, no. 4_suppl (2022): 172. http://dx.doi.org/10.1200/jco.2022.40.4_suppl.172.
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172 Background: Next generation sequencing (NGS) is the laboratory cornerstone of precision oncology treatment. In advanced colorectal cancer (CRC), current guidelines recommend testing RAS, BRAF and MMR biomarkers as standard of care. The added value of comprehensive genomic profiling is so far unclear. Traditional NGS operations are complicated, requiring specialized equipment and personnel. In many jurisdictions, cancer patients are treated in publicly-funded community hospitals, where NGS is not typically utilized and access to testing via send-out services is associated with lengthy turnaround times. Here, we have validated and implemented one of the world's first "point of care" NGS services. Our early experience on NGS implementation and impact in CRC patients is described. Methods: All NGS studies were performed using the Oncomine Precision Assay (OPA) on the genexus integrated sequencer. NGS was performed at the request of the treating physician. All NGS was performed in a local community pathology lab by histotechnologists, simultaneously responsible for IHC testing (such as MMR) and interpreted by anatomic pathologists in conjunction with routine diagnostic pathology services. Retrospective chart review was performed for all patients undergoing sequencing studies and key data, including turnaround time and NGS findings were extracted from the electronic medical record for analysis. Results: A total of 51 cases with CRC were tested using point of care NGS from November 2020-August 2021, initiated by treating physicians. The median turnaround time for results was 3 days. Oncogenic driver events were identified in 46 (90%) cases, including canonical mutations in KRAS, NRAS and BRAF (Table). Actionable mutations were identified in 13 (25%) samples that would not have been identified with single-gene testing. Conclusions: Here, we show that comprehensive NGS can reveal occult resistance mechanisms to standard therapy and identify actionable biomarkers in a substantial proportion of patients with CRC. NGS added valuable information compared to guideline-recommended testing standards. Our study demonstrates that local testing can have rapid turnaround times. To our knowledge, this is the first report of “point of care” NGS in CRC. Further follow up is needed to explore the utility of these expanded roles for NGS testing. [Table: see text]
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Chen, Guanghua, and Chia Chou. "Joint Contribution of Multiple Equatorial Waves to Tropical Cyclogenesis over the Western North Pacific." Monthly Weather Review 142, no. 1 (2014): 79–93. http://dx.doi.org/10.1175/mwr-d-13-00207.1.
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Abstract A composite study is performed to examine the differences in equatorial wave behaviors and large-scale background patterns during tropical cyclone (TC) genesis. After removing TC contamination, Madden–Julian oscillation (MJO), equatorial Rossby (ER) wave, mixed Rossby–gravity (MRG) wave, and tropical depression (TD)-type disturbance (jointly referred to as the MT wave) are quantified to evaluate the attribution of TC geneses. Given that TC geneses are attributed to a single wave or multiple waves, the eight categories are specified based on the moderate thresholds. The TC geneses related to multiple waves are roughly twice as many as those related to a single wave. The MT wave alone accounts for a minor proportion of TC geneses without collaboration with other larger-scale waves. The mean TC genesis location related to ER wave shifts to higher latitudes, and the TC geneses attributed to both of MJO and MT waves are more concentrated at the west. The single-wave categories are characterized by a zonally propagating component with a large spatial scale. In contrast, the joint contribution from more than one wave type favors creating a coherent environment with enhanced low-level cyclonic vorticity, horizontal convergence, or vertical easterly shear in a preferred region. Consequently, the waves have a more robust structure and a more northwestward-propagating component. Correspondingly, the TC geneses in the MJO–ER category tend to occur within a monsoon trough dominated by cyclonic circulation. For the MJO–MT category, the background field exhibits a confluence pattern with a monsoon trough to the west and easterly flows to the east. The collaboration of the ER and MT waves facilitates TC geneses within an easterly environment in the southern flank of the subtropical high.
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Pérez-Alarcón, Albenis, Rogert Sorí, José Carlos Fernández-Alvarez, Raquel Nieto, and Luis Gimeno. "Moisture Sources for Tropical Cyclones Genesis in the Coast of West Africa through a Lagrangian Approach." Environmental Sciences Proceedings 4, no. 1 (2020): 3. http://dx.doi.org/10.3390/ecas2020-08126.
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Atmospheric moisture transport plays an important role in the genesis of tropical cyclones (TCs). In this study, the moisture sources associated with the genesis of TCs in the tropical Atlantic Ocean near West Africa, from June to November in the period 1980–2018, were identified. To detect the location of the TCs geneses, the HURDAT2 database from the National Hurricane Center was used. Additionally, global outputs of the Lagrangian FLEXPART model were used to determine the moisture sources that provided water vapor for the genesis of TCs. This model permitted us to track backward in time the air masses from the genesis region of the TCs and identify regions where air masses uptake moisture before reach the target regions. The results reveal that 18.1% (108 TC) of the total number of TCs that formed in the North Atlantic basin were originated in the region of study. The largest frequency for the TCs geneses was observed in August and September, with each one representing approximately 45% of the total. The transport of moisture associated with the genesis of TCs mainly comes from the east of the North and South Atlantic Ocean, as well as from West Africa and the Sahel region. The patterns of moisture uptake confirmed an interhemispheric moisture transport. Finally, during the El Niño, the moisture uptake is more intense over the Atlantic Ocean close to West Africa around 15 °N of latitude, while during La Niña, the pattern is slightly weaker but covers a wider area over the Atlantic Ocean and the north of Africa.
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D’Agnano, Vito, Fabio Perrotta, Giulia Maria Stella, et al. "Molecular Diagnostic Yield and Safety Profile of Ultrasound-Guided Lung Biopsies: A Cross-Sectional Study." Cancers 16, no. 16 (2024): 2860. http://dx.doi.org/10.3390/cancers16162860.
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Background: The recent advances in precision oncology for lung cancer treatment has focused attention on the importance of obtaining appropriate specimens for tissue diagnosis as well as comprehensive molecular profiling. CT scan-guided biopsies and bronchoscopy are currently the main procedures employed for tissue sampling. However, growing evidence suggests that ultrasound-guided biopsies may represent an effective as well as safe approach in this diagnostic area. This study explores the safety and the diagnostic yield for cancer molecular profiling in ultrasound-guided percutaneous lung lesion biopsies (US-PLLB). Methods: One hundred consecutive patients with suspected lung cancer, between January 2021 and May 2024, who had ultrasound-guided lung biopsies have been retrospectively analyzed. Molecular profiling was conducted with next-generation sequencing Genexus using Oncomine precision assay or polymerase chain reaction according to specimen quality. Qualitative immunohistochemical assay of programmed death ligand 1 (PD-L1) expression was evaluated by the Dako PD-L1 immunohistochemistry 22C3 pharmDx assay. The co-primary endpoints were the molecular diagnostic yield and the safety profile of US-guided lung biopsies. Results: From January 2021 to May 2024, 100 US-guided lung biopsies were carried out and 95 were considered for inclusion in the study. US-PLLB provided informative tissue for a histological evaluation in 93 of 95 patients with an overall diagnostic accuracy of 96.84% [Sensitivity: 92.63%; Specificity: 96.84%; PPV: 100%; NPV: 100%]. Sixty-Six patients were diagnosed with NSCLC (69.47%) and were considered for molecular diagnostic yield evaluation and PD-L1 testing. Four patients had malignant lymphoid lesions. US-PLLB was not adequate to achieve a final diagnosis in three patients (3.16%). Complete molecular profiling and PD-L1 evaluation were achieved in all patients with adenocarcinoma (molecular diagnostic yield: 100%). PD-L1 evaluation was achieved in 28 of 29 patients (96.55%) with either SCC or NOS lung cancer. The overall complication rate was 9.47% (n = 9). Six patients (6.31%) developed pneumothorax, while three patients (3.16%) suffered mild haemoptysis without desaturation. Conclusions: According to our findings, US-guided lung biopsy is a safe, minimally invasive procedure in patients with suspected lung malignancies, providing an excellent diagnostic yield for both comprehensive molecular profiling and PD-L1 testing. In addition, our results suggest that US-guided biopsy may also be an effective diagnostic approach in patients with suspected lung lymphoma.
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Chen, Tsing-Chang, Jenq-Dar Tsay, and Ming-Chang Yen. "Genesis and Development of Spring Rainstorms in Northern Southeast Asia: Southwest China–Northern Indochina and the Northern South China Sea." Monthly Weather Review 145, no. 12 (2017): 4949–76. http://dx.doi.org/10.1175/mwr-d-17-0059.1.
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During May and June, the monsoon rainfall in northern Southeast Asia is primarily produced by rainstorms. At the mature stage, these storms, coupled with a midtropospheric subsynoptic-scale trough, produce rainfall ≥50 mm (6 h)−1and exhibit a cyclonic surface vortex. With a scale ~ O(102) km, rainstorms during the period of 1979–2016 are identified with station and satellite observations, along with assimilation data. Several dynamic processes of rainstorm geneses are disclosed by an extensive analysis. 1) Maximum occurrence of rainstorm geneses is located in the midtroposphere of two regions (northern Vietnam–southwestern China and the northern South China Sea), but eventually penetrates downward to the surface. 2) The environment favorable for rainstorm genesis is a southwest–northeast-oriented narrow trough formed by the confluence of the midtropospheric northeasterly around the eastern Tibetan Plateau and the lower-tropospheric monsoon southwesterlies. Because the criterion for Charney–Stern instability is met by the shear flow of this narrow trough, rainstorms are likely initiated by this instability. 3) The majority of rainstorm geneses occurs during the evening over the land and the morning at sea. This timing preference is caused by the modulation of the clockwise rotation of the East Asia continent circulation in response to the diurnal variation of the land–sea thermal contrast. These new findings from this study offer not only a new perspective for the genesis mechanism of the late spring–early summer rainstorms in northern Southeast Asia, but also a new initiative to develop medium-range forecasts for these rainstorms.
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Sarda, Shrutii, David Merrill, Heesun Shin, et al. "680 Automated Ion torrent based solution enables accurate gut microbiome quantification of bacterial species relevant to research in cancer and its response to immunotherapy." Journal for ImmunoTherapy of Cancer 8, Suppl 3 (2020): A719. http://dx.doi.org/10.1136/jitc-2020-sitc2020.0680.
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BackgroundA low-cost targeted solution to profiling gut microbial diversity is sequencing of the 16S rRNA gene; however, it is often insufficient to gain species level resolution due to high homology across different bacteria. Therefore, we developed a first-of-its-kind targeted sequencing solution that supplements 16S gene targets, with highly species-specific primers for a cohort of 73 bacteria associated with research in diabetes, cancer and its response to immunotherapy, gastrointestinal and auto-immune disorders. This assay performs at 100% sensitivity and specificity for the species-level detection (Ion AmpliSeq Microbiome Health Research Kit: www.thermofisher.com/ngsmicrobiome) of these bacteria and is hence better suited for gut microbiome profiling in the context of the above phenotypes, as compared to other existing solutions.MethodsTo assess the utility of the panel in cancer immunotherapy research, we sequenced DNA from 15 stool samples from subjects with Non-Small Cell Lung Carcinoma (NSCLC) undergoing immunotherapy, and compared their microbiome profiles to 26 healthy stool samples collected internally. With our post-sequencing workflow on Ion Reporter™, we automatically generate a report with taxonomic classifications, sample diversity metrics through QIIME2 integration, and relative abundance visualizations for bacteria across multiple samples.ResultsWe identified significant microbiome composition differences between the healthy samples and cancer/treated samples, as evidenced by (i) a clear separation between the two cohorts based on a beta diversity principal coordinate analysis (PCoA) plot, driven by large abundance changes in Clostridium, Lachnoclostridium, Subdolinigranulum and Oscillibacter (P < 0.05), (ii) grouping into distinct classes based on overall microbiome profiles (Analysis-of-Similarities ANOSIM P = 0.003), and (iii) differences in abundances of specific bacteria with anti-tumor effects1 such as F. prausnitzii (P = 0.02).ConclusionsWe have created a highly multiplexed, sensitive and specific assay for robust characterization of gut microbiota, with compatibility on both (i) the Ion GeneStudio S5™ with a 48-hr sample-to-result turnaround, and (ii) the new Ion Genexus™ System, a fully integrated platform featuring a hands-off, automated sample-to-report workflow that delivers results in a single day. It enables an efficient and affordable means for conducting extensive analyses of the human microbiome having applications in the study of phenotypic variability, and the potential relationship to disease.For research use only. Not for use in diagnostic procedures.ReferenceMa J, Sun L, Liu Y. et al. Alter between gut bacteria and blood metabolites and the anti-tumor effects of Faecalibacterium prausnitzii in breast cancer. BMC Microbiol 2020; 20:1–19.
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Rappenglück, Michael A. "Natural Iron Silicides: A Systematic Review." Minerals 12, no. 2 (2022): 188. http://dx.doi.org/10.3390/min12020188.
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This review systematically presents all finds of geogenic, impact-induced, and extraterrestrial iron silicide minerals known at the end of 2021. The respective morphological characteristics, composition, proven or reasonably suspected genesis, and possible correlations of different geneses are listed and supported by the available literature (2021). Artificially produced iron silicides are only dealt with insofar as the question of differentiation from natural minerals is concerned, especially regarding dating to pre-industrial and pretechnogenic times.
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JASTI, MADHU, Sergio Hernandez, Nick Siepert, et al. "Abstract 5018: Evaluation of whole blood fractionation conditions for optimal liquid biopsy NGS applications." Cancer Research 84, no. 6_Supplement (2024): 5018. http://dx.doi.org/10.1158/1538-7445.am2024-5018.
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Abstract “For research use only. Not for diagnostic procedures.” Introduction: Liquid biopsy is a non-invasive alternative to tissue biopsies for cancer profiling using cell-free total nucleic acids (cfTNA; including cell free DNA (cfDNA) and cell free RNA (cfRNA)), the biological analytes derived from tumor cells floating in blood. However, it is crucial to optimize the whole blood fractionation workflows, since limited access to a suitable centrifuge at blood collection sites poses a challenge to this critical fractionation step, causing delays in processing the blood, leading to decreased sensitivity of variant detection and increased occurrence of false positives. Current study shows a systematic comparison of the effects of 8 different blood fractionation conditions for spin speeds, temperatures and duration on the levels of cfTNA derived from K2-EDTA-derived blood from multiple healthy and non-small cell lung cancer (NSCLC) donors, using next generation sequencing (NGS). Methods: CfTNA from K2-EDTA derived plasma was isolated using the MagMAXTM Cell-Free Total Nucleic Acid kit and the Genexus™ Cell free total nucleic acid kit, utilizing the KingFisher TM Flex and GenexusTMpurification systems. Yield and integrity of cfTNA isolated was evaluated by Qubit™, Agilent Tapestation and quantitative real time PCR (qRT-PCR). Furthermore, the impact of these test conditions on quality control specifications of DNA and RNA was assessed by performing sequencing on an Ion TorrentTM GenexusTM sequencing instrument using the AmpliSeqTM HD target amplification assay, that reports sequence variations in DNA for 42 cancer related genes and RNA in 7 cancer related genes. Results: Results showed comparable performance for all the 8 blood fractionation conditions tested, for yield and cfDNA profiles compared to control condition, indicating that there is no significant impact of these conditions on either yield or integrity of the cfDNA isolated. In addition, NGS data also showed that on target reads and median molecular coverage for DNA are comparable for all the conditions tested. However, sequencing data demonstrated that one test condition showed significantly better performance for RNA quality control specifications in terms of RNA mapped reads and molecular coverage for process controls used in the assay, compared to all other conditions tested. Conclusion: Data suggests that one of the whole blood fractionation conditions tested impact the cfRNA recovery from plasma without negatively affecting the yield, and integrity of isolated cfDNA and shows increased sensitivity for RNA process controls in NGS. Citation Format: MADHU JASTI, Sergio Hernandez, Nick Siepert, Lokesh Kodandaramaswamy, Thilanka Jayaweera, Jeoffrey Schageman, Angie Cheng, Susanne Lopez, Miao Der Chen. Evaluation of whole blood fractionation conditions for optimal liquid biopsy NGS applications [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 5018.
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O Reilly, Mary, Bruce Moran, Kieran Sheahan, et al. "Oncomine panel testing in the clinical management of colorectal cancer: An Irish experience." Journal of Clinical Oncology 43, no. 4_suppl (2025): 212. https://doi.org/10.1200/jco.2025.43.4_suppl.212.
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212 Background: Early onset colorectal cancer (EOCRC) is defined as colorectal cancer (CRC) in those under the age of 50. It has been hypothesized that the molecular signatures of EOCRC could differ from those of late onset CRC (LOCRC). Next Generation Sequencing (NGS) using the Oncomine Precision Assay is a useful tool to guide treatment decisions based on the presence or absence of oncogene drivers. While not all of these mutations currently influence the clinical management of CRC patients, ongoing research on the function of these genes and the development of new drugs could offer insights into patient prognosis, treatment choices and prediction of therapy resistance. Methods: Of >1500 CRC identified between 2010 and 2024, we investigated the clinical and genetic characteristics of 38 EOCRC and 41 LOCRC cases. All cases were microsatellite stable and staged I-III. NGS was performed with the Ion Torrent Genexus integrated platform using the Oncomine 45-gene Precision Assay panel. DNA hotspot mutations and copy number variations (CNVs) were assessed, followed by the classification of clinically relevant mutations using the genetic variant OncoKB database. Results: There were 38 EOCRC and 41 LOCRC cases. The age range was 27 to 49 and 50 to 91, with median ages of 44 and 72, in the EOCRC and LOCRC groups, respectively. Based on the stratification of NGS mutation levels as per the OncoKB classification, all patients in both groups (100%) had level 1 actionable mutations. NRAS wild type (WT) status was present in 100% of patients in both groups. However, differences were observed in KRAS status: 84% (32/38) of EOCRC were KRAS WT, while only 65% (27/41) of LOCRC were KRAS WT. While 3 % (n=1) of the EOCRC cohort had an ERBB2 amplification, none were detected in the LOCRC group. BRAF V600E mutations were present in 11% (4/38) of EOCRC and 12% (5/41) of LOCRC. Level 4 mutations were detected in 18% (n=7) of EOCRC, all of which were PIK3CA mutations, while these were seen in 24% (n=10) of the LOCRC group, including 9 PIK3CA mutations and 1 FGFR1 mutation. Regarding concurrent mutations, 2 patients in the EOCRC group had two mutations each: one with concurrent KRAS and PIK3CA mutations, and another with concurrent JAK2 and KRAS mutations. The LOCRC group had a higher incidence of concurrent mutations, with 8 patients affected: 6 had concurrent KRAS and PIK3CA mutations, and 2 had concurrent BRAF and PIK3CA mutations. Conclusions: Our study demonstrates that EOCRC have higher KRAS WT status and fewer concurrent mutations compared to LOCRC, who exhibit more frequent KRAS non-G12C oncogenic mutations. It also underscores the importance of routine NGS mutation panel analysis in CRC management. Additionally, identifying specific mutations in patients that relapse can guide targeted therapies, potentially improving outcomes and offering personalized treatment options.