70 results on '"Chris Karlovich"'
Search Results
2. TPM, FPKM, or Normalized Counts? A Comparative Study of Quantification Measures for the Analysis of RNA-seq Data from the NCI Patient-Derived Models Repository
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Yingdong Zhao, Ming-Chung Li, Mariam M. Konaté, Li Chen, Biswajit Das, Chris Karlovich, P. Mickey Williams, Yvonne A. Evrard, James H. Doroshow, and Lisa M. McShane
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RNA sequencing ,Quantification measures ,Normalization ,TPM ,FPKM ,Count ,Medicine - Abstract
Abstract Background In order to correctly decode phenotypic information from RNA-sequencing (RNA-seq) data, careful selection of the RNA-seq quantification measure is critical for inter-sample comparisons and for downstream analyses, such as differential gene expression between two or more conditions. Several methods have been proposed and continue to be used. However, a consensus has not been reached regarding the best gene expression quantification method for RNA-seq data analysis. Methods In the present study, we used replicate samples from each of 20 patient-derived xenograft (PDX) models spanning 15 tumor types, for a total of 61 human tumor xenograft samples available through the NCI patient-derived model repository (PDMR). We compared the reproducibility across replicate samples based on TPM (transcripts per million), FPKM (fragments per kilobase of transcript per million fragments mapped), and normalized counts using coefficient of variation, intraclass correlation coefficient, and cluster analysis. Results Our results revealed that hierarchical clustering on normalized count data tended to group replicate samples from the same PDX model together more accurately than TPM and FPKM data. Furthermore, normalized count data were observed to have the lowest median coefficient of variation (CV), and highest intraclass correlation (ICC) values across all replicate samples from the same model and for the same gene across all PDX models compared to TPM and FPKM data. Conclusion We provided compelling evidence for a preferred quantification measure to conduct downstream analyses of PDX RNA-seq data. To our knowledge, this is the first comparative study of RNA-seq data quantification measures conducted on PDX models, which are known to be inherently more variable than cell line models. Our findings are consistent with what others have shown for human tumors and cell lines and add further support to the thesis that normalized counts are the best choice for the analysis of RNA-seq data across samples.
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- 2021
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3. Supplementary Methods, Supplementary Tables 1-3, Supplementary Figure 1 from Exosome-Based Detection of EGFR T790M in Plasma from Non–Small Cell Lung Cancer Patients
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Johan K. Skog, Luis E. Raez, Bakhos A. Tannous, Jihane Tannous, Mikkel Noerholm, Anne Krug, Mitch Raponi, Chris Karlovich, Raajdeep Venkatesan, Mia Sher, Patricia L. Neal, John Healy, James Hurley, Vasisht Tadigotla, Dominik G. Grimm, and Elena Castellanos-Rizaldos
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Supplementary Table 1. Clinical characteristics of patients included in the study (T790M positive/negative and the different pools of plasma from healthy donors); Supplementary Table 2. Mutant admixtures used as part of the analytical validation. These samples were run using three technical replicates; Supplementary Table 3. Analytical validation. Supplementary Table 4. Summary of patient demographics and T790M results for the validation cohort; Supplementary Figure 1. Patient histology for T790M positive and negative patients (A) and disease distribution by stage (B) for all the clinical samples.
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- 2023
4. Supplementary Methods. Supplementary Tables S1-7. Supplementary Figures S1 and S2 from Assessment of EGFR Mutation Status in Matched Plasma and Tumor Tissue of NSCLC Patients from a Phase I Study of Rociletinib (CO-1686)
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Heather Wakelee, Keunchil Park, Andrew R. Allen, Mitch Raponi, Lindsey Rolfe, Darrin Despain, Shannon Matheny, Patrick O'Donnell, Sean Chien, Lin Wu, Cloud Paweletz, Shirish Gadgeel, D. Ross Camidge, Benjamin Solomon, Jean-Charles Soria, David Spigel, Leora Horn, Philipp Angenendt, Wei Wen, Krzysztof Konopa, Lecia V. Sequist, Elaina Mann, Jong-Mu Sun, Jonathan W. Goldman, and Chris Karlovich
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Supplementary Tables S1-7. Supplementary Table S1. Clinical characteristics of patients in the present analysis. Supplementary Table S2. Plasma/tissue concordance results for del19 and L858R mutations. Supplementary Table S3. EGFR mutations identified in tissue and plasma by the cobas® EGFR mutation test. Supplementary Table S4. Concordance between tumor and BEAMing plasma EGFR status. Supplementary Table S5. Overall concordance between cobas® and BEAMing EGFR mutation tests. Supplementary Table S6. Platform Comparison of T790M tests in a sample set enriched for low copy plasma cases. Supplementary Table S7. EGFR mutation detection by cobas® plasma test and NSCLC disease classification (n = 72) Supplementary Figures S1-2. Supplementary Figure S1. Tumor burden is a weak predictor of ability to detect EGFR mutations in plasma. Supplementary Figure S2. T790M to activating mutation ratio in plasma is associated with depth of response to rociletinib.
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- 2023
5. Data from Assessment of EGFR Mutation Status in Matched Plasma and Tumor Tissue of NSCLC Patients from a Phase I Study of Rociletinib (CO-1686)
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Heather Wakelee, Keunchil Park, Andrew R. Allen, Mitch Raponi, Lindsey Rolfe, Darrin Despain, Shannon Matheny, Patrick O'Donnell, Sean Chien, Lin Wu, Cloud Paweletz, Shirish Gadgeel, D. Ross Camidge, Benjamin Solomon, Jean-Charles Soria, David Spigel, Leora Horn, Philipp Angenendt, Wei Wen, Krzysztof Konopa, Lecia V. Sequist, Elaina Mann, Jong-Mu Sun, Jonathan W. Goldman, and Chris Karlovich
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Purpose: The evaluation of plasma testing for the EGFR resistance mutation T790M in NSCLC patients has not been broadly explored. We investigated the detection of EGFR activating and T790M mutations in matched tumor tissue and plasma, mostly from patients with acquired resistance to first-generation EGFR inhibitors.Experimental Design: Samples were obtained from two studies, an observational study and a phase I trial of rociletinib, a mutant-selective inhibitor of EGFR that targets both activating mutations and T790M. Plasma testing was performed with the cobas EGFR plasma test and BEAMing.Results: The positive percent agreement (PPA) between cobas plasma and tumor results was 73% (55/75) for activating mutations and 64% (21/33) for T790M. The PPA between BEAMing plasma and tumor results was 82% (49/60) for activating mutations and 73% (33/45) for T790M. Presence of extrathoracic (M1b) versus intrathoracic (M1a/M0) disease was found to be strongly associated with ability to identify EGFR mutations in plasma (P < 0.001). Rociletinib objective response rates (ORR) were 52% [95% confidence interval (CI), 31 – 74%] for cobas tumor T790M-positive and 44% (95% CI, 25 – 63%) for BEAMing plasma T790M-positive patients. A drop in plasma-mutant EGFR levels to ≤10 molecules/mL was seen by day 21 of treatment in 7 of 8 patients with documented partial response.Conclusions: These findings suggest the cobas and BEAMing plasma tests can be useful tools for noninvasive assessment and monitoring of the T790M resistance mutation in NSCLC, and could complement tumor testing by identifying T790M mutations missed because of tumor heterogeneity or biopsy inadequacy. Clin Cancer Res; 22(10); 2386–95. ©2016 AACR.
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- 2023
6. Data from Exosome-Based Detection of EGFR T790M in Plasma from Non–Small Cell Lung Cancer Patients
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Johan K. Skog, Luis E. Raez, Bakhos A. Tannous, Jihane Tannous, Mikkel Noerholm, Anne Krug, Mitch Raponi, Chris Karlovich, Raajdeep Venkatesan, Mia Sher, Patricia L. Neal, John Healy, James Hurley, Vasisht Tadigotla, Dominik G. Grimm, and Elena Castellanos-Rizaldos
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Purpose: About 60% of non–small cell lung cancer (NSCLC) patients develop resistance to targeted epidermal growth factor receptor (EGFR) inhibitor therapy through the EGFR T790M mutation. Patients with this mutation respond well to third-generation tyrosine kinase inhibitors, but obtaining a tissue biopsy to confirm the mutation poses risks and is often not feasible. Liquid biopsies using circulating free tumor DNA (cfDNA) have emerged as a noninvasive option to detect the mutation; however, sensitivity is low as many patients have too few detectable copies in circulation. Here, we have developed and validated a novel test that overcomes the limited abundance of the mutation by simultaneously capturing and interrogating exosomal RNA/DNA and cfDNA (exoNA) in a single step followed by a sensitive allele-specific qPCR.Experimental Design: ExoNA was extracted from the plasma of NSCLC patients with biopsy-confirmed T790M-positive (N = 102) and T790M-negative (N = 108) samples. The T790M mutation status was determined using an analytically validated allele-specific qPCR assay in a Clinical Laboratory Improvement Amendment laboratory.Results: Detection of the T790M mutation on exoNA achieved 92% sensitivity and 89% specificity using tumor biopsy results as gold standard. We also obtained high sensitivity (88%) in patients with intrathoracic disease (M0/M1a), for whom detection by liquid biopsy has been particularly challenging.Conclusions: The combination of exoRNA/DNA and cfDNA for T790M detection has higher sensitivity and specificity compared with historical cohorts using cfDNA alone. This could further help avoid unnecessary tumor biopsies for T790M mutation testing. Clin Cancer Res; 24(12); 2944–50. ©2018 AACR.
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- 2023
7. PARP Inhibitor Applicability: Detailed Assays for Homologous Recombination Repair Pathway Components
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Geraldine, O'Sullivan Coyne, Chris, Karlovich, Deborah, Wilsker, Andrea Regier, Voth, Ralph E, Parchment, Alice P, Chen, and James H, Doroshow
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Oncology ,Pharmacology (medical) - Abstract
Poly(ADP-ribose) polymerase inhibitors (PARPi) have been in clinical use since 2014 for certain patients with germline
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- 2022
8. Validation of ctDNA Quality Control Materials Through a Precompetitive Collaboration of the Foundation for the National Institutes of Health
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Robert T. McCormack, Daniel Stetson, Dana E. Connors, J. Carl Barrett, Kenneth D. Cole, Steven P. Lund, Chris Karlovich, Thomas Forbes, P. Mickey Williams, Megan H. Cleveland, Laura M. Yee, Christie J. Lau, Cloud P. Paweletz, Gary J. Kelloff, Benoit Destenaves, Hua-Jun He, and Susan M. Keating
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Quality Control ,0301 basic medicine ,Cancer Research ,Engineering ,DNA Copy Number Variations ,media_common.quotation_subject ,Control (management) ,MEDLINE ,Polymerase Chain Reaction ,Circulating Tumor DNA ,03 medical and health sciences ,0302 clinical medicine ,Gene Frequency ,Neoplasms ,Original Reports ,Biomarkers, Tumor ,Humans ,Quality (business) ,Diagnostics ,media_common ,business.industry ,Foundation (engineering) ,High-Throughput Nucleotide Sequencing ,United States ,Engineering management ,030104 developmental biology ,National Institutes of Health (U.S.) ,Oncology ,030220 oncology & carcinogenesis ,Mutation ,business - Abstract
PURPOSEWe report the results from a Foundation for the National Institutes of Health Biomarkers Consortium project to address the absence of well-validated quality control materials (QCMs) for circulating tumor DNA (ctDNA) testing. This absence is considered a cause of variance and inconsistencies in translating ctDNA results into clinical actions.METHODSIn this phase I study, QCMs with 14 clinically relevant mutations representing single nucleotide variants, insertions or deletions (indels), translocations, and copy number variants were sourced from three commercial manufacturers with variant allele frequencies (VAFs) of 5%, 2.5%, 1%, 0.1%, and 0%. Four laboratories tested samples in quadruplicate using two allele-specific droplet digital polymerase chain reaction and three (amplicon and hybrid capture) next-generation sequencing (NGS) panels.RESULTSThe two droplet digital polymerase chain reaction assays reported VAF values very close to the manufacturers’ claimed concentrations for all QCMs. NGS assays reported most single nucleotide variants and indels, but not translocations, close to the expected VAF values. Notably, two NGS assays reported lower VAF than expected for all translocations in all QCM mixtures, possibly related to technical challenges detecting these variants. The ability to call ERBB2 copy number amplifications varied across assays. All three QCMs provided valuable insight into assay precision. Each assay across all variant types demonstrated dropouts at 0.1%, suggesting that the QCM can serve for testing of an assay’s limit of detection with confidence claims for specific variants.CONCLUSIONThese results support the utility of the QCM in testing ctDNA assay analytical performance. However, unique designs and manufacturing methods for the QCM, and variations in a laboratory’s testing configuration, may require testing of multiple QCMs to find the best reagents for accurate result interpretation.
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- 2021
9. Abstract 40: NCI patient derived models repository: PDX, organoid and cell lines from the same patient - bridging the translational pipeline
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Yvonne A. Evrard, Li Chen, Sergio Alcoser, Gareth Bliss, Carrie Bonomi, Suzanne Borgel, John Carter, Ting-Chia Chang, Alice Chen, Kevin Cooper, Biswajit Das, Kelly Dougherty, Lindsay Dutko, Marion Gibson, Michelle M. Ahalt-Gottholm, Tara Grinnage-Pulley, Keegan Kalmbach, Chris Karlovich, Kimberly Klarmann, Shahanawaz Jiwani, Tiffanie Miner, Michael Mullendore, Matthew Murphy, Kevin Plater, Gloryvee Rivera, Jessica Steed, Luke Stockwin, Cindy R. Timme, Dianne L. Newton, Paul Mickey Williams, Melinda G. Hollingshead, and James H. Doroshow
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Cancer Research ,Oncology - Abstract
The National Cancer Institute’s Patient-Derived Models Repository (NCI PDMR; https://pdmr.cancer.gov) has developed a national repository of Patient-Derived Models (PDMs) comprised of patient-derived xenografts (PDXs), in vitro patient-derived tumor cell cultures (PDCs) and cancer associated fibroblasts (CAFs) as well as patient-derived organoids (PDOrg). These PDMs are clinically annotated with molecular information available in an easily accessible database for the extramural community. A key effort in developing these models is to develop matched models sets allowing for larger scale screening efforts using 2D or 3D models to prioritize selection of PDX models for preclinical translational research. To date, over 220 model sets with a PDX, PDOrg, and PDC from a single patient have been developed; 40 of these have matched CAF models allowing for exploration of research questions focused on tumor microenvironment. The largest model sets are in colorectal cancer (COADREAD, n=76), gynecologic cancers (n=33), pancreatic adenocarcinoma (PAAD, n=29), melanoma (MEL, n=19), and head and neck squamous cell carcinomas (HNSCC, n=17). Every model undergoes several quality control assessments that serve as go/no-go criteria including pathology assessment, STR validation, NGS concordance assessment and for PDXs, human:mouse DNA content assessment. It should be noted that not every model is successful in the development or QC phase so additional model sets with only one or two of the model types are also available for researcher requests, for example there are over 125 PDX/PDOrg matched model sets. The NCI is currently performing parallel preclinical screening of PDXs and PDCs or PDOrgs to determine the ability of the in vitro lines to predict in vivo activity. Genetic and histopathologic assessment of these matched model sets have demonstrated a high degree of stability by somatic mutation, copy number alteration (CNA) and gene expression data. Gene expression correlation analysis shows that mean of Spearman r between PDXs 0.89, between matched PDC/PDXs 0.79, and between matched PDOrg/PDXs 0.82. As expected, some variation at the gene expression level when comparing PDX to in vitro cultures by t-SNE can be observed, likely due to the differences in culture conditions. Funded by NCI Contract No. HHSN261200800001E Citation Format: Yvonne A. Evrard, Li Chen, Sergio Alcoser, Gareth Bliss, Carrie Bonomi, Suzanne Borgel, John Carter, Ting-Chia Chang, Alice Chen, Kevin Cooper, Biswajit Das, Kelly Dougherty, Lindsay Dutko, Marion Gibson, Michelle M. Ahalt-Gottholm, Tara Grinnage-Pulley, Keegan Kalmbach, Chris Karlovich, Kimberly Klarmann, Shahanawaz Jiwani, Tiffanie Miner, Michael Mullendore, Matthew Murphy, Kevin Plater, Gloryvee Rivera, Jessica Steed, Luke Stockwin, Cindy R. Timme, Dianne L. Newton, Paul Mickey Williams, Melinda G. Hollingshead, James H. Doroshow. NCI patient derived models repository: PDX, organoid and cell lines from the same patient - bridging the translational pipeline [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 40.
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- 2023
10. Cross-Site Concordance Evaluation of Tumor DNA and RNA Sequencing Platforms for the CIMAC-CIDC Network
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Junko Tsuji, Jin Wang, David Cohen, Jianjun Zhang, Jijun Yu, Ignacio I. Wistuba, Tomas Vilimas, Len Taing, Li Chen, Chunhua Yan, Qing-Rong Chen, Magdalena Thurin, Rebecca A. Enos, Xiaoman Wang, Peng Jiang, Candace Patterson, Mohamed Uduman, Catherine J. Wu, Beatriz Sanchez-Espiridion, Aashna Jhaveri, Yang Liu, Andrew Futreal, Jingxin Fu, Jiexin Zhang, Jack Lee, Donna Neuberg, Curtis Gumbs, Xin Huang, Sylvie Janssens, Collin Tokheim, Zexian Zeng, Avinash Das Sahu, Ming Tang, Carrie Cibulskis, James Lindsay, Cu Nguyen, Jason L. Weirather, Joyce Yu, Stacey Gabriel, Ethan Cerami, Dzifa Y. Duose, Daoud Meerzaman, Chris Karlovich, Sharmistha Sarkar, X. Shirley Liu, Biswajit Das, Sachet A. Shukla, and Jianhua Zhang
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Nonsynonymous substitution ,Cancer Research ,Base Sequence ,Concordance ,RNA ,DNA, Neoplasm ,Computational biology ,Human leukocyte antigen ,Biology ,Article ,chemistry.chemical_compound ,Oncology ,chemistry ,Monitoring, Immunologic ,Immune infiltration ,Neoplasms ,Exome Sequencing ,Fresh frozen ,Humans ,RNA, Neoplasm ,International HapMap Project ,DNA - Abstract
Purpose: Whole-exome (WES) and RNA sequencing (RNA-seq) are key components of cancer immunogenomic analyses. To evaluate the consistency of tumor WES and RNA-seq profiling platforms across different centers, the Cancer Immune Monitoring and Analysis Centers (CIMAC) and the Cancer Immunologic Data Commons (CIDC) conducted a systematic harmonization study. Experimental Design: DNA and RNA were centrally extracted from fresh frozen and formalin-fixed paraffin-embedded non–small cell lung carcinoma tumors and distributed to three centers for WES and RNA-seq profiling. In addition, two 10-plex HapMap cell line pools with known mutations were used to evaluate the accuracy of the WES platforms. Results: The WES platforms achieved high precision (> 0.98) and recall (> 0.87) on the HapMap pools when evaluated on loci using > 50× common coverage. Nonsynonymous mutations clustered by tumor sample, achieving an index of specific agreement above 0.67 among replicates, centers, and sample processing. A DV200 > 24% for RNA, as a putative presequencing RNA quality control (QC) metric, was found to be a reliable threshold for generating consistent expression readouts in RNA-seq and NanoString data. MedTIN > 30 was likewise assessed as a reliable RNA-seq QC metric, above which samples from the same tumor across replicates, centers, and sample processing runs could be robustly clustered and HLA typing, immune infiltration, and immune repertoire inference could be performed. Conclusions: The CIMAC collaborating laboratory platforms effectively generated consistent WES and RNA-seq data and enable robust cross-trial comparisons and meta-analyses of highly complex immuno-oncology biomarker data across the NCI CIMAC-CIDC Network.
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- 2020
11. The Exceptional Responders Initiative: Feasibility of a National Cancer Institute Pilot Study
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Maria F. Cardenas, Kristen M. Leraas, Lalitha K. Shankar, Adrienne Johnson, Benjamin Kim, Jean C. Zenklusen, Thomas J. Giordano, Vincent A. Miller, Julie M. Gastier-Foster, Linghua Wang, Paula M. Jacobs, Brian Rodgers, Irina A. Lubensky, Tracy S. Nolan, Elise C. Kohn, Bhupinder Singh Mann, Richard F. Little, Shakun Malik, Elijah F. Edmondson, Viktoriya Korchina, Lou Staudt, Barbara A. Conley, Manel Esteller, Hui Shen, James V. Tricoli, Sean M Berryman, Carol J. Weil, Geraldine O'Sullivan-Coyne, Jay Bowen, David A. Wheeler, S. Percy Ivy, Lyndsay Harris, Naoko Takebe, Roy Tarnuzzer, Jeffrey White, Paul Williams, Lisa M. McShane, Peter W. Laird, and Chris Karlovich
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Oncology ,0303 health sciences ,Cancer Research ,medicine.medical_specialty ,education.field_of_study ,business.industry ,Population ,MEDLINE ,Exceptional Response ,Disease ,Systemic therapy ,Deep sequencing ,03 medical and health sciences ,0302 clinical medicine ,Germline mutation ,030220 oncology & carcinogenesis ,Internal medicine ,medicine ,education ,business ,Exome sequencing ,030304 developmental biology - Abstract
Background Tumor molecular profiling from patients experiencing exceptional responses to systemic therapy may provide insights into cancer biology and improve treatment tailoring. This pilot study evaluates the feasibility of identifying exceptional responders retrospectively, obtaining pre-exceptional response treatment tumor tissues, and analyzing them with state-of-the-art molecular analysis tools to identify potential molecular explanations for responses. Methods Exceptional response was defined as partial (PR) or complete (CR) response to a systemic treatment with population PR or CR rate less than 10% or an unusually long response (eg, duration >3 times published median). Cases proposed by patients’ clinicians were reviewed by clinical and translational experts. Tumor and normal tissue (if possible) were profiled with whole exome sequencing and, if possible, targeted deep sequencing, RNA sequencing, methylation arrays, and immunohistochemistry. Potential germline mutations were tracked for relevance to disease. Results Cases reflected a variety of tumors and standard and investigational treatments. Of 520 cases, 476 (91.5%) were accepted for further review, and 222 of 476 (46.6%) proposed cases met requirements as exceptional responders. Clinical data were obtained from 168 of 222 cases (75.7%). Tumor was provided from 130 of 168 cases (77.4%). Of 117 of the 130 (90.0%) cases with sufficient nucleic acids, 109 (93.2%) were successfully analyzed; 6 patients had potentially actionable germline mutations. Conclusion Exceptional responses occur with standard and investigational treatment. Retrospective identification of exceptional responders, accessioning, and sequencing of pretreatment archived tissue is feasible. Data from molecular analyses of tumors, particularly when combining results from patients who received similar treatments, may elucidate molecular bases for exceptional responses.
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- 2020
12. Pembrolizumab in Relapsed and Refractory Mycosis Fungoides and Sézary Syndrome: A Multicenter Phase II Study
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Chris Karlovich, Richard Shine, Jennifer H. Yearley, Martin A. Cheever, Pierluigi Porcu, Francine M. Foss, Steven M. Horwitz, Youn H. Kim, Alison J. Moskowitz, Michael S. Khodadoust, Wendy M. Blumenschein, P. Mickey Williams, Andrei R. Shustov, Vivekananda Datta, Erin Cantu, Biswajit Das, Lubomir Sokol, Yi Yang, Sophia Fong, Holden T. Maecker, Shufeng Li, Priyanka B. Subrahmanyam, Nirasha Ramchurren, Alain H. Rook, Elad Sharon, Satish Shanbhag, Jinah Kim, Robert H. Pierce, Steven P. Fling, Holbrook E Kohrt, Asa Davis, Justine N. McCutcheon, and Rajesh Patidar
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Adult ,Male ,Cancer Research ,medicine.medical_specialty ,Skin Neoplasms ,Refractory Mycosis Fungoides ,Phases of clinical research ,Pembrolizumab ,Antibodies, Monoclonal, Humanized ,B7-H1 Antigen ,Drug Administration Schedule ,Antineoplastic Agents, Immunological ,Mycosis Fungoides ,Recurrence ,Biomarkers, Tumor ,medicine ,Humans ,Sezary Syndrome ,In patient ,Infusions, Intravenous ,Aged ,Neoplasm Staging ,Aged, 80 and over ,business.industry ,ORIGINAL REPORTS ,Middle Aged ,Dermatology ,Clinical trial ,Oncology ,Multicenter study ,Monoclonal ,Female ,Neoplasm staging ,business - Abstract
PURPOSE To assess the efficacy of pembrolizumab in patients with advanced relapsed or refractory mycosis fungoides (MF) or Sézary syndrome (SS). PATIENTS AND METHODS CITN-10 is a single-arm, multicenter phase II trial of 24 patients with advanced MF or SS. Patients were treated with pembrolizumab 2 mg/kg every 3 weeks for up to 24 months. The primary end point was overall response rate by consensus global response criteria. RESULTS Patients had advanced-stage disease (23 of 24 with stage IIB to IV MF/SS) and were heavily pretreated with a median of four prior systemic therapies. The overall response rate was 38% with two complete responses and seven partial responses. Of the nine responding patients, six had 90% or more improvement in skin disease by modified Severity Weighted Assessment Tool, and eight had ongoing responses at last follow-up. The median duration of response was not reached, with a median response follow-up time of 58 weeks. Immune-related adverse events led to treatment discontinuation in four patients. A transient worsening of erythroderma and pruritus occurred in 53% of patients with SS. This cutaneous flare reaction did not result in treatment discontinuation for any patient. The flare reaction correlated with high PD-1 expression on Sézary cells but did not associate with subsequent clinical responses or lack of response. Treatment responses did not correlate with expression of PD-L1, total mutation burden, or an interferon-γ gene expression signature. CONCLUSION Pembrolizumab demonstrated significant antitumor activity with durable responses and a favorable safety profile in patients with advanced MF/SS.
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- 2020
13. TPM, FPKM, or Normalized Counts? A Comparative Study of Quantification Measures for the Analysis of RNA-seq Data from the NCI Patient-Derived Models Repository
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Chris Karlovich, P. Mickey Williams, Mariam M. Konaté, Biswajit Das, James H. Doroshow, Ming-Chung Li, Lisa M. McShane, Li Chen, Yvonne A. Evrard, and Yingdong Zhao
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0301 basic medicine ,Normalization (statistics) ,Intraclass correlation ,Coefficient of variation ,FPKM ,RNA-Seq ,DESeq2 ,Computational biology ,RSEM ,Biology ,TPM ,General Biochemistry, Genetics and Molecular Biology ,03 medical and health sciences ,0302 clinical medicine ,Count ,Humans ,Sequence Analysis, RNA ,Gene Expression Profiling ,Research ,High-Throughput Nucleotide Sequencing ,Reproducibility of Results ,RNA sequencing ,General Medicine ,Replicate ,Hierarchical clustering ,TMM ,Human tumor ,Normalization ,030104 developmental biology ,Patient derived xenograft models ,030220 oncology & carcinogenesis ,Medicine ,RNA ,Quantification measures ,Count data - Abstract
Background In order to correctly decode phenotypic information from RNA-sequencing (RNA-seq) data, careful selection of the RNA-seq quantification measure is critical for inter-sample comparisons and for downstream analyses, such as differential gene expression between two or more conditions. Several methods have been proposed and continue to be used. However, a consensus has not been reached regarding the best gene expression quantification method for RNA-seq data analysis. Methods In the present study, we used replicate samples from each of 20 patient-derived xenograft (PDX) models spanning 15 tumor types, for a total of 61 human tumor xenograft samples available through the NCI patient-derived model repository (PDMR). We compared the reproducibility across replicate samples based on TPM (transcripts per million), FPKM (fragments per kilobase of transcript per million fragments mapped), and normalized counts using coefficient of variation, intraclass correlation coefficient, and cluster analysis. Results Our results revealed that hierarchical clustering on normalized count data tended to group replicate samples from the same PDX model together more accurately than TPM and FPKM data. Furthermore, normalized count data were observed to have the lowest median coefficient of variation (CV), and highest intraclass correlation (ICC) values across all replicate samples from the same model and for the same gene across all PDX models compared to TPM and FPKM data. Conclusion We provided compelling evidence for a preferred quantification measure to conduct downstream analyses of PDX RNA-seq data. To our knowledge, this is the first comparative study of RNA-seq data quantification measures conducted on PDX models, which are known to be inherently more variable than cell line models. Our findings are consistent with what others have shown for human tumors and cell lines and add further support to the thesis that normalized counts are the best choice for the analysis of RNA-seq data across samples.
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- 2021
14. Development and interlaboratory evaluation of a NIST Reference Material RM 8366 for EGFR and MET gene copy number measurements
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Kenneth D. Cole, Biswajit Das, Li Chen, P. Mickey Williams, Steve Lund, Megan H. Cleveland, Christopher R McEvoy, Jamie L. Almeida, Carolyn R. Steffen, Corinne Camalier, Liang-Chun Liu, Kara L. Norman, Chris Karlovich, Hua-Jun He, and Andrew Fellowes
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0301 basic medicine ,EGFR ,Clinical Biochemistry ,Gene Dosage ,Computational biology ,Biology ,Polymerase Chain Reaction ,Proto-Oncogene Mas ,Article ,DNA sequencing ,03 medical and health sciences ,0302 clinical medicine ,Reference genes ,Tumor Cells, Cultured ,Humans ,cancer ,Digital polymerase chain reaction ,Copy-number variation ,Gene ,reference material ,Exome sequencing ,next generation sequencing ,Whole genome sequencing ,digital PCR ,Biochemistry (medical) ,High-Throughput Nucleotide Sequencing ,DNA, Neoplasm ,General Medicine ,Proto-Oncogene Proteins c-met ,Reference Standards ,ErbB Receptors ,genomic DNA ,030104 developmental biology ,030220 oncology & carcinogenesis ,MET - Abstract
Background The National Institute of Standards and Technology (NIST) Reference Material RM 8366 was developed to improve the quality of gene copy measurements of EGFR (epidermal growth factor receptor) and MET (proto-oncogene, receptor tyrosine kinase), important targets for cancer diagnostics and treatment. The reference material is composed of genomic DNA prepared from six human cancer cell lines with different levels of amplification of the target genes. Methods The reference values for the ratios of the EGFR and MET gene copy numbers to the copy numbers of reference genes were measured using digital PCR. The digital PCR measurements were confirmed by two additional laboratories. The samples were also characterized using Next Generation Sequencing (NGS) methods including whole genome sequencing (WGS) at three levels of coverage (approximately 1 ×, 5 × and greater than 30 ×), whole exome sequencing (WES), and two different pan-cancer gene panels. The WES data were analyzed using three different bioinformatic algorithms. Results The certified values (digital PCR) for EGFR and MET were in good agreement (within 20%) with the values obtained from the different NGS methods and algorithms for five of the six components; one component had lower NGS values. Conclusions This study shows that NIST RM 8366 is a valuable reference material to evaluate the performance of assays that assess EGFR and MET gene copy number measurements.
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- 2019
15. Exosome-based detection of activating and resistance EGFR mutations from plasma of non-small cell lung cancer patients
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Johan Skog, Vasisht Tadigotla, Dominik G. Grimm, Chris Karlovich, Xuan Zhang, Luis E. Raez, and Elena Castellanos-Rizaldos
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0301 basic medicine ,biology ,business.industry ,medicine.disease ,Exosome ,respiratory tract diseases ,03 medical and health sciences ,T790M ,Exon ,030104 developmental biology ,0302 clinical medicine ,Oncology ,Cell-free fetal DNA ,030220 oncology & carcinogenesis ,Cancer research ,biology.protein ,Medicine ,Epidermal growth factor receptor ,Liquid biopsy ,business ,Lung cancer ,Gene - Abstract
Non-small cell lung cancer (NSCLC) is the most prevalent form of lung cancer and its molecular landscape has been extensively studied. The most common genetic alterations in NSCLC are mutations within the epidermal growth factor receptor (EGFR) gene, with frequencies between 10-40%. There are several molecular targeted therapies for patients harboring these mutations. Liquid biopsies constitute a flexible approach to monitor these mutations in real time as opposed to tissue biopsies that represent a single snap-shot in time. However, interrogating cell free DNA (cfDNA) has inherent biological limitations, especially at early or localized disease stages, where there is not enough tumor material released into the patient's circulation. We developed a qPCR- based test (ExoDx EGFR) that interrogates mutations within EGFR using Exosomal RNA/DNA and cfDNA (ExoNA) derived from plasma in a cohort of 110 NSCLC patients. The performance of the assay yielded an overall sensitivity of 90% for L858R, 83% for T790M and 73% for exon 19 indels with specificities of 100%, 100%, and 96% respectively. In a subcohort of patients with extrathoracic disease (M1b and MX) the sensitivities were 92% (L858R), 95% (T790M), and 86% (exon 19 indels) with specificity of 100%, 100% and 94% respectively.
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- 2019
16. Abstract 973: Comparative single cell transcriptome profiling of primary tumors, CTCs and metastatic sites from a bladder cancer PDX model
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Tomas Vilimas, Brandie Fullmer, Alyssa Chapman, Rini Pauly, Ting-Chia Chang, Li Chen, Biswajit Das, Chris Karlovich, Yvonne Evrard, Melinda Hollingshead, Howard Stotler, Michelle Ahalt-Gottholm, Tara Grinnage-Pulley, Mickey Williams, and James H. Doroshow
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Cancer Research ,Oncology - Abstract
Background: A PDX bladder cancer model, BL0293-F563, spontaneously metastasizes to the liver and bone, and sheds high numbers of circulating tumor cells (CTCs). This PDX model provides a unique opportunity to explore the relationships between primary tumors, CTCs, and metastases. Methods: BL0293-F563 tumors (available from the NCI Patient-Derived Models Repository [https://pdmr.cancer.gov/] and originally developed by Jackson Laboratories) were implanted into NSG mice, and primary tumors, metastatic nodules in the liver, and blood were collected at maximal allowable tumor burden. Tumor tissue was dissociated using Miltenyi Tumor Dissociation Kit with OctoDissociator, and Human CTCs were enriched from mouse blood through negative selection with anti-mouse CD45 and anti-mouse MHC-1 magnetic beads. Single cell sequencing was done using 10X Genomics 3’ gene expression assay v3.1. Data processing and analysis was done using 10X Genomics’ Cell Ranger pipeline, Seurat, and cNMF. Results: single cell RNAseq data from primary tumors, CTCs, and metastases from 9 mice were aggregated into a single dataset, and cells were classified into 17 clusters using Seurat FindNeighbors. All clusters contained cells from multiple sites (primary tumor, CTCs, metastases), but three clusters were enriched in CTCs and one cluster was composed of mostly primary tumor cells. All clusters exhibited epithelial-like gene expression signature scores, suggesting that CTC shedding was occurring without prominent epithelial-mesenchymal transition. CTC-enriched clusters showed elevated expression of RHO pathway genes, implicating ameboid-like migration in CTC shedding in this PDX model. Consistent with expected differences in oxygenation states, CTC-enriched clusters exhibited a lower hypoxia gene expression score than primary tumor and metastasis-enriched clusters. CTC-enriched clusters also showed higher expression of oxidative phosphorylation genes, suggesting metabolic differences between CTCs and cells from other sites. Additionally, two of three CTC-enriched clusters had elevated expression of mitosis-associated genes, indicating that at least some subpopulations of CTCs are actively cycling. A metastasis suppressor gene KISS1 was expressed in a subset of primary tumor cells but undetectable in CTCs, suggesting that KISS1 expression loss occurs before CTC shedding. Conclusions: Utilizing single cell gene expression profiling, we have linked the gene expression profile of CTCs to specific cell subpopulations in primary tumors and metastases. We show that CTC-enriched cell clusters appear to maintain an epithelial phenotype. Subpopulations of CTC cells exhibit enrichment of motility-associated transcripts and features of active cell cycling. Our results implicate a known metastasis suppressor gene KISS1 in CTC shedding and metastatic dissemination in this PDX model. Citation Format: Tomas Vilimas, Brandie Fullmer, Alyssa Chapman, Rini Pauly, Ting-Chia Chang, Li Chen, Biswajit Das, Chris Karlovich, Yvonne Evrard, Melinda Hollingshead, Howard Stotler, Michelle Ahalt-Gottholm, Tara Grinnage-Pulley, Mickey Williams, James H. Doroshow. Comparative single cell transcriptome profiling of primary tumors, CTCs and metastatic sites from a bladder cancer PDX model [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 973.
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- 2022
17. DIPG-47. TSO500ctDNA sequencing reveals oncogenic mutations and copy number variations in the liquid biome of children with diffuse midline glioma
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Erin R Bonner, Robin Harrington, Augustine Eze, Miriam Bornhorst, Cassie N Kline, Adam Dawood, Biswajit Das, Li Chen, Rini Pauly, P Mickey Williams, Chris Karlovich, Amanda Peach, D'Andra Howell, James Doroshow, Lindsay Kilburn, Roger J Packer, Sabine Mueller, and Javad Nazarian
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Cancer Research ,Oncology ,Neurology (clinical) - Abstract
BACKGROUND: Molecular profiling of childhood CNS tumors is critical for diagnosis and clinical management, yet tissue access is restricted due to sensitive neuroanatomical locations. Moreover, CNS tumors including diffuse midline glioma (DMG) exhibit mutational heterogeneity and clonal evolution, which cannot be captured by upfront diagnostic biopsy alone. To address the lack of tumor visibility, and tprovide opportunity for longitudinal sampling, we validated and optimized a commercially available deep sequencing platform for analysis of circulating tumor DNA (TSO500ctDNATM). METHODS: In a proof-of-concept study, we defined the sensitivity, specificity, and clinical relevance of our novel ctDNA platform via analysis of paired tissue, CSF, and blood from children with DMG (n=10). Paired samples were assessed for concordance and sequencing results were compared to digital droplet PCR (ddPCR) detection of prognostic H3K27M mutation. RESULTS: DMG associated mutations in genes including H3-3A, H3C2, TP53, and ACVR1 were detected in ctDNA, including in CSF samples with low (
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- 2022
18. EGFR Genotyping of Matched Urine, Plasma, and Tumor Tissue in Patients With Non–Small-Cell Lung Cancer Treated With Rociletinib, an EGFR Tyrosine Kinase Inhibitor
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Benjamin Solomon, Chris Karlovich, Ronald B. Natale, Mark G. Erlander, Mitch Raponi, Corey J. Langer, Jean-Charles Soria, Gregory A. Otterson, Heather A. Wakelee, Shirish M. Gadgeel, Mark A. Socinski, Joel W. Neal, Jason B. Litten, Lecia V. Sequist, Vassiliki A. Papadimitrakopoulou, Maurice Pérol, Sai-Hong Ignatius Ou, Vlada Melnikova, D. Ross Camidge, Stephen V. Liu, Jonathan W. Goldman, Karen L. Reckamp, Darrin Despain, Sergey Yurasov, Helena A. Yu, Aleksandra Franovic, and Tarek Mekhail
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0301 basic medicine ,Cancer Research ,medicine.medical_specialty ,biology ,business.industry ,Urology ,Urine ,medicine.disease ,respiratory tract diseases ,03 medical and health sciences ,T790M ,030104 developmental biology ,0302 clinical medicine ,Oncology ,030220 oncology & carcinogenesis ,biology.protein ,Medicine ,In patient ,Epidermal growth factor receptor ,Rociletinib ,business ,Lung cancer ,Genotyping ,Egfr tyrosine kinase - Abstract
Purpose Liquid biopsies represent an attractive alternative to tissue biopsies, particularly rebiopsies, in determining patient eligibility for targeted therapies. Clinical utility of urine genotyping, however, has not been explored extensively. We evaluated epidermal growth factor receptor ( EGFR) T790M detection in matched urine, plasma, and tissue and the clinical outcomes of patients with advanced non–small-cell lung cancer treated with rociletinib. Methods Tissue (n = 540), plasma (n = 482), and urine (n = 213) were collected from evaluable patients enrolled in TIGER-X, a phase I/II study. Genotyping was performed by therascreen EGFR testing in tissue, BEAMing in plasma, and a quantitative short footprint assay (Trovera) in urine, which was used to further examine discordant samples. Results Positive percent agreement with tissue T790M results was similar for urine (82%; 142 of 173) and plasma (81%; 313 of 387) genotyping. Urine and plasma together identified more patients who were T790M positive (92%) than tissue alone (83%) among matched samples (n = 177). The ability to identify mutations in plasma was strongly associated with M stage ( P < .001); rate of T790M detection for patients with M1a/M0 disease increased from 54% for plasma alone to 85% when urine and plasma were both examined. Objective response rates of patients who were T790M positive were comparable between tumor (34%), plasma (32%), and urine (37%). Conclusion Clinical response to rociletinib was comparable irrespective of whether T790M status was identified by liquid or tissue biopsy. Combined, urine and plasma identified a higher percentage of patients who were T790M positive than tumor genotyping alone and improved detection of T790M, particularly in the absence of distant metastases. These findings support the noninvasive analysis of urine and plasma before tumor rebiopsy when assessing T790M status.
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- 2018
19. Molecular Profiling-Based Assignment of Cancer Therapy (NCI-MPACT): A Randomized Multicenter Phase II Trial
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Nancy Moore, James H. Doroshow, Saiama N. Waqar, Christina L. Rosenberger, Mel Simpson, Richard Piekarz, Alida Palmisano, Funda Meric-Bernstam, Stephen Leong, Yingdong Zhao, P. Mickey Williams, Kanwal Pratap Singh Raghav, Richard Simon, Biswajit Das, S. Kummar, Larry Rubinstein, Chris Karlovich, David J. Sims, Mariam M. Konaté, Ming Chung Li, Eric C. Polley, Jared C. Foster, Alice P. Chen, Geraldine O'Sullivan Coyne, and Chih Jian Lih
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Adult ,Male ,0301 basic medicine ,Oncology ,Cancer Research ,medicine.medical_specialty ,Pyridones ,Cancer therapy ,Antineoplastic Agents ,Pyrimidinones ,DNA sequencing ,Carboplatin ,Young Adult ,03 medical and health sciences ,0302 clinical medicine ,Double-Blind Method ,Neoplasms ,Internal medicine ,Temozolomide ,medicine ,Humans ,Everolimus ,Aged ,Aged, 80 and over ,business.industry ,Gene Expression Profiling ,DNA, Neoplasm ,ORIGINAL REPORTS ,Targeted Drug Therapy ,Middle Aged ,Refractory cancer ,030104 developmental biology ,Molecular Diagnostic Techniques ,030220 oncology & carcinogenesis ,Pyrazoles ,Benzimidazoles ,Female ,business - Abstract
PURPOSEThis trial assessed the utility of applying tumor DNA sequencing to treatment selection for patients with advanced, refractory cancer and somatic mutations in one of four signaling pathways by comparing the efficacy of four study regimens that were either matched to the patient's aberrant pathway (experimental arm) or not matched to that pathway (control arm).MATERIALS AND METHODSAdult patients with an actionable mutation of interest were randomly assigned 2:1 to receive either (1) a study regimen identified to target the aberrant pathway found in their tumor (veliparib with temozolomide or adavosertib with carboplatin [DNA repair pathway], everolimus [PI3K pathway], or trametinib [RAS/RAF/MEK pathway]), or (2) one of the same four regimens, but chosen from among those not targeting that pathway.RESULTSAmong 49 patients treated in the experimental arm, the objective response rate was 2% (95% CI, 0% to 10.9%). One of 20 patients (5%) in the experimental trametinib cohort had a partial response. There were no responses in the other cohorts. Although patients and physicians were blinded to the sequencing and random assignment results, a higher pretreatment dropout rate was observed in the control arm (22%) compared with the experimental arm (6%; P = .038), suggesting that some patients may have had prior tumor mutation profiling performed that led to a lack of participation in the control arm.CONCLUSIONFurther investigation, better annotation of predictive biomarkers, and the development of more effective agents are necessary to inform treatment decisions in an era of precision cancer medicine. Increasing prevalence of tumor mutation profiling and preference for targeted therapy make it difficult to use a randomized phase II design to evaluate targeted therapy efficacy in an advanced disease setting.
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- 2021
20. ACTR-61. A RANDOMIZED PHASE 2 TRIAL OF CEDIRANIB IN COMBINATION WITH OLAPARIB VERSUS BEVACIZUMAB IN PATIENTS WITH RECURRENT GLIOBLASTOMA
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Andrew B. Nixon, Douglas E. Ney, Jan Drappatz, Alona Muzikansky, S. Percy Ivy, Mary Welch, Biswajit Das, Isabel Arrillaga-Romany, P. Mickey Williams, John L. Villano, Eudocia Q. Lee, Elizabeth M. Swisher, Robert Aiken, Pierre Giglio, Chris Karlovich, Elizabeth R. Gerstner, Tracy T. Batchelor, David Picconi, Benjamin M. Ellingson, Jian Campian, Kevin P. Becker, and Solmaz Sahebjam
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Oncology ,Cancer Research ,medicine.medical_specialty ,Temozolomide ,Bevacizumab ,business.industry ,Surrogate endpoint ,Phases of clinical research ,Olaparib ,Cediranib ,chemistry.chemical_compound ,chemistry ,Internal medicine ,Adult Clinical Trials - Non-Immunologic ,Medicine ,In patient ,Neurology (clinical) ,Progression-free survival ,business ,medicine.drug - Abstract
BACKGROUND Like most proliferating tumors, GBM relies heavily on accurate DNA repair for maintenance of genome stability. Dysfunction in repair of both single and double strand DNA breaks by PARP inhibition and impairment of homologous recombination, respectively, would be synthetically lethal. In this study we combined the PARP inhibitor olaparib with cediranib, a pan VEGF receptor inhibitor. Cediranib may mediate disruption in the homologous recombination pathway through its antiangiogenic properties. METHODS Through the Experimental Therapeutics Clinical Trials Network, we performed an open-label randomized phase II study of bevacizumab (BEV)- naive adult patients with first or second recurrence of glioblastoma after radiation and temozolomide. Patients were randomized 1:1 to receive either olaparib 200 mg by mouth twice daily with cediranib 30 mg by mouth daily or BEV 10 mg/kg IV every 2 weeks. The primary endpoint was progression-free survival at 6 months (PFS6). Secondary endpoints included safety and overall survival. Exploratory objectives included blood, tissue and imaging-based biomarkers of response to treatment. RESULTS Seventy patients were enrolled. Median age was 60.5 years (range: 19–79), 39% females, median KPS was 90 (range: 60–100). Baseline characteristics were well balanced. With a data cut-off of 5/2/2019, PFS6 was 14% [95% CI 4–30%] in the cediranib/olaparib arm vs 30.9% [95% CI 12.7–51.2%] in the BEV arm. Median OS was 247 days in the cediranib/olaparib arm vs 201 days in the BEV arm, HR 0.816, 95% CI (0.431, 1.546). Related grade 3, 4 or 5 toxicity was experienced in 29% vs 12% of patients for the cediranib/olaparib vs BEV arm. CONCLUSION Treatment with cediranib/olaparib failed to increase PFS and OS in patients with recurrent GBM. Blood, tissue and imaging correlates will be presented to help understand why this treatment combination was unsuccessful.
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- 2019
21. Improved EGFR mutation detection using combined exosomal RNA and circulating tumor DNA in NSCLC patient plasma
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Johan Skog, Chris Karlovich, Jonathan W. Goldman, A. Spiel, Mitch Raponi, K. Brinkmann, S. Bentink, Heather A. Wakelee, J. Emenegger, Lecia V. Sequist, T. Priewasser, Mikkel Noerholm, Shirish M. Gadgeel, Elena Castellanos-Rizaldos, D. Enderle, Anne Krug, D.R. Camidge, Dominik G. Grimm, and J-C. Soria
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0301 basic medicine ,Lung Neoplasms ,Thoracic Tumors ,EGFR ,exosomes ,NSCLC ,exoRNA ,Circulating Tumor DNA ,03 medical and health sciences ,T790M ,0302 clinical medicine ,Germline mutation ,Carcinoma, Non-Small-Cell Lung ,medicine ,Carcinoma ,Humans ,Rociletinib ,Liquid biopsy ,Lung cancer ,liquid biopsy ,business.industry ,Cancer ,Original Articles ,ctDNA ,Hematology ,medicine.disease ,ErbB Receptors ,Editor's Choice ,030104 developmental biology ,Oncology ,Cell-free fetal DNA ,030220 oncology & carcinogenesis ,Mutation ,Cancer research ,RNA ,business - Abstract
Background A major limitation of circulating tumor DNA (ctDNA) for somatic mutation detection has been the low level of ctDNA found in a subset of cancer patients. We investigated whether using a combined isolation of exosomal RNA (exoRNA) and cell-free DNA (cfDNA) could improve blood-based liquid biopsy for EGFR mutation detection in non-small-cell lung cancer (NSCLC) patients. Patients and methods Matched pretreatment tumor and plasma were collected from 84 patients enrolled in TIGER-X (NCT01526928), a phase 1/2 study of rociletinib in mutant EGFR NSCLC patients. The combined isolated exoRNA and cfDNA (exoNA) was analyzed blinded for mutations using a targeted next-generation sequencing panel (EXO1000) and compared with existing data from the same samples using analysis of ctDNA by BEAMing. Results For exoNA, the sensitivity was 98% for detection of activating EGFR mutations and 90% for EGFR T790M. The corresponding sensitivities for ctDNA by BEAMing were 82% for activating mutations and 84% for T790M. In a subgroup of patients with intrathoracic metastatic disease (M0/M1a; n = 21), the sensitivity increased from 26% to 74% for activating mutations (P = 0.003) and from 19% to 31% for T790M (P = 0.5) when using exoNA for detection. Conclusions Combining exoRNA and ctDNA increased the sensitivity for EGFR mutation detection in plasma, with the largest improvement seen in the subgroup of M0/M1a disease patients known to have low levels of ctDNA and poses challenges for mutation detection on ctDNA alone. Clinical Trials NCT01526928
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- 2018
22. Abstract P097: Comparative single cell transcriptome profiling of primary tumors, CTCs and metastatic sites from a bladder cancer PDX model
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Tomas Vilimas, Brandie Fullmer, Alyssa Chapman, Li Chen, Ting-Chia Chang, Rini Pauly, Biswajit Das, Chris Karlovich, Yvonne A. Evrard, Howard Stotler, Michelle M. Gottholm-Ahalt, Tara Grinnage-Pulley, Melinda G. Hollingshead, James H. Doroshow, and P. Mickey Williams
- Subjects
Cancer Research ,Oncology - Abstract
Background: A PDX bladder cancer model, BL0293-F563, grows large subcutaneous tumors, spontaneously metastasizes to the liver and bone, and sheds high numbers of circulating tumor cells (CTCs). This PDX model provides a unique opportunity to explore the relationships between primary tumors, CTCs and metastatic cell subpopulations. Methods: BL0293-F563 tumors (available from the NCI Patient-Derived Models Repository [https://pdmr.cancer.gov/] and originally developed by Jackson Laboratories) were implanted into NSG mice and and primary tumors, metastatic nodules in the liver, and blood were collected at maximal allowable tumor burden. Tumor tissue was dissociated using Miltenyi Tumor Dissociation Kit with OctoDissociator, and Human CTCs were enriched from whole mouse blood through negative selection with anti-mouse CD45 and anti-mouse MHC-1 magnetic beads. Single cell sequencing was done using 10X Genomics 3’ gene expression assay v3.1. Sequencing libraries were prepared using 10X Genomics Chromium and 3’ gene expression kit v3.1. Data processing and analysis was done using 10X Genomics’ Cell Ranger pipeline, Seurat, and consensus non-negative matrix factorization. Results: Using Seurat FindNeighbors, cells in the aggregated dataset were classified into 17 distinct clusters. All clusters were comprised of cells from multiple sites (primary tumor, CTCs, metastases), but three clusters were enriched in CTCs and one cluster was composed of mostly primary tumor cells. All clusters exhibited an epithelial-like gene expression signature score, suggesting that CTC shedding was occurring without prominent epithelial-mesenchymal transition. Consistent with expected differences in oxygenation states, CTC-enriched clusters exhibited a lower hypoxia gene expression score than primary tumor and metastasis-enriched clusters. CTC-enriched clusters also showed higher expression of oxidative phosphorylation genes, suggesting metabolic differences between CTCs and cells from primary tumors and metastases. Based on Human Primary Cell Atlas phenotype prediction, several clusters were associated with stem cell like phenotypes. Additionally, two of three CTC-enriched clusters had elevated expression of mitosis-associated genes, suggesting that at least some populations of CTCs are not quiescent but actively cycling. Conclusions: Utilizing single cell gene expression profiling, we have linked the gene expression profile of CTCs to specific cell subpopulations in primary tumors and metastases. We show that CTC-enriched cell clusters appear to maintain an epithelial phenotype. Subpopulations of CTC cells exhibited enrichment of stemness-associated transcripts and features of active cell cycling. Citation Format: Tomas Vilimas, Brandie Fullmer, Alyssa Chapman, Li Chen, Ting-Chia Chang, Rini Pauly, Biswajit Das, Chris Karlovich, Yvonne A. Evrard, Howard Stotler, Michelle M. Gottholm-Ahalt, Tara Grinnage-Pulley, Melinda G. Hollingshead, James H. Doroshow, P. Mickey Williams. Comparative single cell transcriptome profiling of primary tumors, CTCs and metastatic sites from a bladder cancer PDX model [abstract]. In: Proceedings of the AACR-NCI-EORTC Virtual International Conference on Molecular Targets and Cancer Therapeutics; 2021 Oct 7-10. Philadelphia (PA): AACR; Mol Cancer Ther 2021;20(12 Suppl):Abstract nr P097.
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- 2021
23. Abstract 3012: Patient-derived models of rare cancers in the National Cancer Institute's patient-derived models repository
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Michael Mullendore, Jenna Hull, P. Mickey Williams, Cindy R. Timme, Dianne L. Newton, Tia Shearer, Shahanawaz Jiwani, Li Chen, Chris Karlovich, Michelle Eugeni, Tara Grinnage-Polley, James H. Doroshow, Kristen Cooley, Tom Walsh, Devynn Breen, Emily Delaney, Kimberly Klarmann, Michelle M. Gottholm-Ahalt, Melinda G. Hollingshead, Jesse Stottlemyer, Chelsea McGlynn, Malorie Morris, John Mark Carter, Yvonne A. Evrard, Alice P. Chen, Ting-Chia Chang, Candace Mallow, Sergio Y. Alcoser, Shannon Uzelac, Biswajit Das, and Justine Mills
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Oncology ,Cancer Research ,medicine.medical_specialty ,business.industry ,Internal medicine ,medicine ,Cancer ,business ,medicine.disease - Abstract
There is an unmet need for preclinical models of rare cancers and rare disease sub-types. The National Cancer Institute's Patient-Derived Models Repository (NCI PDMR; https://pdmr.cancer.gov) is developing quality-controlled, early-passage, clinically-annotated patient-derived tumor xenografts (PDXs), in vitro tumor cell cultures (PDCs), cancer associated fibroblasts (CAFs), and patient-derived organoids (PDOrg) and has focused on addressing unmet needs in the preclinical model space including developing models from adult and pediatric patients with rare cancers. To date, NCI has created and molecularly characterized over 150 preclinical models of rare cancer including indications such as Hurthle cell carcinoma, osteosarcomas, Merkel cell carcinomas, salivary gland cancers, synovial sarcomas, and carcinosarcomas. Rare cancer models developed to date will be reviewed and their histopathologic and molecular characteristics compared to that reported in the clinical setting. A pipeline to identify fusion proteins in these rare cancers such as the Ewing sarcoma EWSR1-FLI1 fusion and NAB2-STAT6 fusions in solitary fibrous tumors (SFT) has been implemented. Four malignant peripheral nerve sheath tumors (MPNST) PDX models are available for researches; these models were developed from patients diagnosed between the ages of 37-68. At the time of model development, two patients were treatment naïve and two had prior radiotherapy. Two of the MPNST PDX models have NF1 oncogenic mutations, three have deep deletions in CDKN2A/B, and three have a mutation in either EED or SUZ12 consistent with the reported molecular characteristics of patients with MPNST. Also of clinical relevance, of two mesothelioma models available, one carries an NF2 driver mutation and the other BAP1 and LATS2 and a PDX model for Hurthle cell carcinoma has wide-spread loss of heterozygosity (LOH 80%). Models for other rare cancers are in development, including four cholangiocarcinoma PDXs with histopathologic confirmation that are currently being expanded for molecular characterization and distribution. Funded by NCI Contract No. HHSN261200800001E Citation Format: Cindy R. Timme, Sergio Y. Alcoser, Devynn Breen, John Carter, Ting-Chia Chang, Alice Chen, Li Chen, Kristen Cooley, Biswajit Das, Emily Delaney, Michelle A. Eugeni, Michelle M. Gottholm-Ahalt, Tara Grinnage-Polley, Jenna Hull, Chris Karlovich, Kimberly Klarmann, Shahanawaz Jiwani, Candace Mallow, Chelsea McGlynn, Justine Mills, Malorie Morris, Michael Mullendore, Dianne Newton, Tia Shearer, Jesse Stottlemyer, Shannon Uzelac, Thomas Walsh, P. Mickey Williams, Yvonne A. Evrard, Melinda G. Hollingshead, James H. Doroshow. Patient-derived models of rare cancers in the National Cancer Institute's patient-derived models repository [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 3012.
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- 2021
24. Abstract 3015: Applications of immunohistochemistry in characterization of patient derived xenograft models
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Michelle M. Gottholm Ahalt, Kelly Benauer, Jesse Stottlemyer, Vishnuprabha Rahulkannan, Michelle Eugeni, James H. Doroshow, Gloryvee Rivera, Lindsay Dutko, Tiffanie Chase, Erin Cantu, Melinda G. Hollingshead, Yvonne A. Evrard, Biswajit Das, Suzanne Borgel, Mickey Williams, Emily Delaney, Shahanawaz Jiwani, Chris Karlovich, Howard Stotler, Chelsea McGlynn, and John Mark Carter
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Cancer Research ,Pathology ,medicine.medical_specialty ,Oncology ,business.industry ,medicine ,Immunohistochemistry ,business ,Tumor xenograft - Abstract
Background: Well characterized patient derived xenograft models (PDX) are becoming the preferred pre-clinical tool in translational cancer research for biologic understanding of the disease, development of new treatments, and identifying potential therapy predictive and resistant biomarkers. Characterization of PDX models using a multi-omic approach is most desirable, however such efforts can be expensive and technically demanding. Immunohistochemistry (IHC) has become an indispensable ancillary tool in the accurate classification of tumor types, determination of cell of origin, identification of biologic properties like growth and metastatic potential, and evaluation for the presence/absence of therapeutic or prognostic biomarkers. Methods: 43 IHC assays were validated on the Leica Bond RX automated staining platform to identify common inconsistencies in PDX development including markers for classifying carcinomas, lymphomas, sarcomas, murine tumors, and theragnostic biomarkers. Rabbit antibodies are used rather than mouse antibodies to prevent non-specific staining of murine tissue. Results: 1. IHC evaluation of models within NCI's Patient Derived Models Repository (pdmr.cancer.gov) led to re-classification or sub-classification of 12 tumor models in accordance with WHO guidelines. 2. IHC evaluation of theragnostic markers in 8 breast cancer PDX models showed concordant results throughout passaging, suggesting stability of these biomarkers in our models. 3. We observe malignant transformation of murine or transplanted benign human tissue at a rate of 2.5%. On IHC analysis, 52% were human lymphomas, 20% were murine lymphomas, and 28% were other murine tumors. Conclusions: IHC is a rapid, cost-effective tool that can be used for accurate tumor classification, identifying subclonal outgrowth and tumor evolution, assessing stability of biomarkers and identifying malignant transformation of benign tissue. Funded by NCI Contract No. HHSN261200800001E ANTIBODYCLONEVENDORANTIBODYCLONEVENDORAndrogen Receptor[EPR1535(2)]abcamGATA3[EPR16651]abcamB-Catenin[E247]abcamGCDFP-15[EPR1582Y]abcamCD19polyclonalabcamGFAPpolyclonalDAKO/AgilentCD3polyclonalabcamHER2 ErbB2[SP3]abcamCD20[SP32]abcamKi-67[D2H10]Cell SignalingCD34[EP373Y]abcamKu80[EPR3468]abcamCD45polyclonalabcamMGMTMT3.1MilliporeCD56 (NCAM1)[EPR2566]abcamMitochondria Marker (Biotin)MTC02abcamCD68[EPR20545]abcamMyogenin[EPR4789]abcamCDX2[EPR2764Y]abcamNAPSIN A[EPR6252]abcamChromogranin A[SP12]abcamp63polyclonalGeneTexCK7 (purified)[EPR1619Y]abcamPD-1[EPR4877(2)]abcamCK19[EPR1580Y]abcamPD-L1 (CD274)RBT-PDL1LifeSpan BiosciencesCK20[EPR1622Y]abcamProgesterone Receptor[SP2]abcamCytokeratin wide spectrumpolyclonalabcamProstate Specific Antigen (PSA)[EP1588Y]abcamDesmin[Y66]abcamS100[EPR19013]abcamEBV LMP1[D24-G]abcamSmooth Muscle Actin (SMA)polyclonalabcamERG[EPR3864]abcamSynaptophysin[SP11]abcamEstrogen Receptor[SP1]abcamTTF1[SP141]abcamFOXP1monoclonalLifeSpan BiosciencesVimentin[EPR3776]abcamFOXP3(5H10L18)Invitrogen Citation Format: Lindsay Dutko, Gloryvee Rivera, Erin Cantu, Vishnuprabha Rahulkannan, Kelly Benauer, Tiffanie Chase, Emily Delaney, Jesse Stottlemyer, Chelsea McGlynn, Howard Stotler, John Carter, Suzanne Borgel, Michelle M. Gottholm Ahalt, Michelle Eugeni, Melinda Hollingshead, Yvonne Evrard, Chris Karlovich, Biswajit Das, Mickey Williams, James H. Doroshow, Shahanawaz Jiwani. Applications of immunohistochemistry in characterization of patient derived xenograft models [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 3015.
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- 2021
25. Abstract 3139: Study of tumor heterogeneity and subclonality in primary pancreatic and metastatic sites from rapid autopsy patients in PDMR
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Lindsay Dutko, Shahanawaz Jiwani, Dianne L. Newton, Li Chen, Chris Karlovich, Michelle Eugeni, James H. Doroshow, Kelly Benauer, P. Mickey Williams, Nikitha Nair, Chapman Alyssa, Anna L. Fong, Biswajit Das, Luis E. Romero, Melinda G. Hollingshead, Yvonne A. Evrard, Ting-Chia Chang, Tomas Vilimas, Kelly Dougherty, Amanda Peach, and Marianne Morton
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Cancer Research ,Somatic cell ,business.industry ,Cancer ,Drug resistance ,medicine.disease ,medicine.disease_cause ,Tumor heterogeneity ,Metastasis ,Germline mutation ,Oncology ,Cancer research ,medicine ,Adenocarcinoma ,KRAS ,business - Abstract
Background: A major set of preclinical models derived from specimens acquired from rapid autopsy patients in the National Cancer Institute Patient-Derived Models Repository (NCI PDMR, https://pdmr.cancer.gov) were from pancreatic adenocarcinoma (PAAD) patients, with metastatic specimens originating from liver, colon, omentum, and lung. Genomic characterization of these preclinical models provides a unique opportunity to study tumor heterogeneity and subclonality associated with the metastatic process and potential treatment resistance. Methods: To date, 30 rapid autopsy patient-derived xenograft (PDX)/patient-derived cell (PDC)/patient-derived organoid (PDOrg ) models derived from pancreatic adenocarcinoma patients (n = 9) have been sequenced using whole-exome sequencing (WES) and RNASeq. Tumor heterogeneity between primary and metastatic sites was studied based on somatic mutation, copy number alteration (CNA) and gene expression data. A bioinformatics workflow was developed to stably infer and visualize the tumor subclonality by integrating the tools of PyClone, SCHISM, and TIMESCAPE, using somatic mutations and site-specific copy number data of multiple samples generated from PDX models in primary and metastatic sites. Results: Among 30 rapid autopsy preclinical models from primary and metastatic sites, liver is the most common metastatic site in PAAD (9/19=47%) compared to other sites. Driver mutations are conserved in all preclinical model specimens derived from a given patient. KRAS p.G12D is present in 28 PDX/PDC/PDOrg models as well as the corresponding patient specimens, and BRAF p.V600E is present in other preclinical models. The fraction of the genome affected by CNA remains stable within a PDX model across passages (n=18, mean=7.63%, sd=5.90%). However, we found that this increased when comparing PDX models derived from metastatic sites versus the primary site (n=16, mean=19.47%, sd=9.69%). This indicates the presence of tumor heterogeneity between metastatic and primary sites. Site-specific subclones were identified in PDX models from two patients (521955 and 485368) and a phylogenetic tree of primary and metastasis sites indicates that one liver metastasis had a unique seeding event compared to the other metastatic sites for both patients. Conclusion: Tumor heterogeneity and subclonality was observed in preclinical models generated from PAAD patients in the NCI PDMR. These models provide a unique resource for preclinical studies in tumor evolution, metastatic spread mediators, and drug resistance. Citation Format: Li Chen, Biswajit Das, Yvonne A. Evrard, Chris A. Karlovich, Tomas Vilimas, Amanda Peach, Chapman Alyssa, Nikitha Nair, Anna L. Fong, Luis Romero, Ting-Chia Chang, Shahanawaz Jiwani, Lindsay Dutko, Kelly Benauer, Marianne Morton, Kelly Dougherty, Michelle A. Eugeni, Dianne Newton, Melinda G. Hollingshead, P. Mickey Williams, James H. Doroshow. Study of tumor heterogeneity and subclonality in primary pancreatic and metastatic sites from rapid autopsy patients in PDMR [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 3139.
- Published
- 2021
26. Profiling 523 cancer associated genes in circulating tumor DNA of children with CNS tumors
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Chris Karlovich, D'Andra Howell, Maria Saeed, Robin D. Harrington, Erin R. Bonner, Biswajit Das, Amanda Peach, Javad Nazarian, and Paul Williams
- Subjects
Surgical resection ,Cancer Research ,business.industry ,Central nervous system ,Cancer ,medicine.disease ,medicine.anatomical_structure ,Oncology ,Circulating tumor DNA ,Cancer research ,medicine ,CNS TUMORS ,business ,Gene - Abstract
3023 Background: Pediatric central nervous system (CNS) cancers often pose unique challenges including tumor ‘invisibility’, where surgical resection is restricted due to the sensitive tumor location and tissue biopsy is not always feasible. Detecting cancer associated mutations and copy number variations (CNV) at diagnosis is increasingly important, as the WHO classification of pediatric CNS cancers has incorporated molecular signatures with tumor grade. To achieve CNS tumor molecular ‘visibility’, we previously established a liquid biopsy platform for detecting single nucleotide variants in circulating tumor DNA (ctDNA). However, our method was limited by the restricted number of genes that can be monitored and the inability to detect genomic events including CNVs. To address this, we developed a deep sequencing liquid biopsy approach to profile alterations across selected genes. Our platform provides an opportunity for multi-gene monitoring, to assess tumor subclonal evolution and response to treatment in the absence of repeat tissue biopsies. Methods: We tested the performance of our platform using paired tissue, CSF, and plasma/serum from 10 children with diffuse midline glioma (DMG). ctDNA was analyzed using the TruSight Oncology 500 (TSO500) ctDNA targeted panel covering 523 genes. Matched tumor, CSF, and blood were assessed for concordance and sequencing results were compared to digital droplet PCR (ddPCR) detection of H3K27M mutation. Results: The median exons with ³500X coverage was 96% for 7 CSF samples with optimal input (³60ng), 0.01% for 3 CSF samples with < 5ng input, and 74.5% for plasma/serum samples. ctDNA was more readily detectable in CSF, yet concordance between paired tumor, CSF and plasma/serum was observed. DMG associated mutations in genes including H3F3A, HIST1H3B, TP53, and ACVR1 were detected in ctDNA. Of 9 H3K27M mutations identified in tumor, 8 were present in CSF and 3 in plasma/serum, for a positive percent agreement of 89% and 33%, respectively, with the tumor results. Among CSF samples, H3.3K27M was detected in 6/6 cases, and H3.1K27M in 2/3 cases, with variant allele frequencies comparable to ddPCR results. CNVs including PDGFRA/B and MDM4 amplifications were present in CSF and confirmed by analysis of paired tumor. Additional events, including PIK3CA p.E545Q, PPM1D truncation, and KRAS amplification, were detected in CSF but absent from paired tumor, indicating tissue heterogeneity. Strategies to optimize ctDNA detection, including optimization of ctDNA isolation and adjustment of library QC metrics, were identified. Conclusions: This proof-of-concept study demonstrates the feasibility of our high depth, targeted sequencing approach for detecting clinically relevant mutations in ctDNA from children with CNS tumors. This approach may aid in diagnosis of CNS tumor molecular subtype, and monitoring of tumor evolution and response to therapy in serially collected ctDNA.
- Published
- 2021
27. A Highly Sensitive and Quantitative Test Platform for Detection of NSCLC EGFR Mutations in Urine and Plasma
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David Berz, Vladislava O. Melnikova, Peter J. P. Croucher, Errin Samuelsz, Jennifer Geis, Shiloh Guerrero, Mitch Raponi, Geoffrey R. Oxnard, D. Ross Camidge, Heather A. Wakelee, Elaina Mann, Chris Karlovich, Shannon Matheny, Mark G. Erlander, Maurice Pérol, Karen L. Reckamp, Lindsey Rolfe, Shirish M. Gadgeel, Cecile Rose T. Vibat, Karena Kosco, and Lecia V. Sequist
- Subjects
Adult ,Male ,0301 basic medicine ,Pulmonary and Respiratory Medicine ,medicine.medical_specialty ,Lung Neoplasms ,Urine ,NSCLC ,medicine.disease_cause ,T790M ,Gastroenterology ,03 medical and health sciences ,Exon ,0302 clinical medicine ,Double-Blind Method ,Carcinoma, Non-Small-Cell Lung ,Internal medicine ,medicine ,Humans ,Rociletinib ,Liquid biopsy ,Aged ,Retrospective Studies ,Aged, 80 and over ,Circulating tumor DNA ,Mutation ,business.industry ,Middle Aged ,Resistance mutation ,EGFR mutations ,Molecular biology ,respiratory tract diseases ,ErbB Receptors ,030104 developmental biology ,Oncology ,Egfr mutation ,030220 oncology & carcinogenesis ,Female ,business - Abstract
Introduction In approximately 60% of patients with NSCLC who are receiving EGFR tyrosine kinase inhibitors, resistance develops through the acquisition of EGFR T790M mutation. We aimed to demonstrate that a highly sensitive and quantitative next-generation sequencing analysis of EGFR mutations from urine and plasma specimens is feasible. Methods Short footprint mutation enrichment next-generation sequencing assays were used to interrogate EGFR activating mutations and the T790M resistance mutation in urine or plasma specimens from patients enrolled in TIGER-X (NCT01526928), a phase 1/2 clinical study of rociletinib in previously treated patients with EGFR mutant–positive advanced NSCLC. Results Of 63 patients, 60 had evaluable tissue specimens. When the tissue result was used as a reference, the sensitivity of EGFR mutation detection in urine was 72% (34 of 47 specimens) for T790M, 75% (12 of 16) for L858R, and 67% (28 of 42) for exon 19 deletions. With specimens that met a recommended volume of 90 to 100 mL, the sensitivity was 93% (13 of 14 specimens) for T790M, 80% (four of five) for L858R, and 83% (10 of 12) for exon 19 deletions. A comparable sensitivity of EGFR mutation detection was observed in plasma: 93% (38 of 41 specimens) for T790M, 100% (17 of 17) for L858R, and 87% (34 of 39) for exon 19 deletions. Together, urine and plasma testing identified 12 additional T790M-positive cases that were either undetectable or inadequate by tissue test. In nine patients monitored while receiving treatment with rociletinib, a rapid decrease in urine T790M levels was observed by day 21. Conclusions DNA derived from NSCLC tumors can be detected with high sensitivity in urine and plasma, enabling diagnostic detection and monitoring of therapeutic response from these noninvasive "liquid biopsy" samples.
- Published
- 2016
28. Genomic profiling of three pathways through molecular profiling-based assignment of cancer therapy (NCI- MPACT)
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Yingdong Zhao, Chih-Jian Lih, G. O'Sullivan Coyne, L. Rubinstein, Kanwal Pratap Singh Raghav, Paul Williams, Funda Meric-Bernstam, Sabrina S. Khan, James H. Doroshow, Chris Karlovich, Shivaani Kummar, Saiama N. Waqar, Nancy Moore, Naoko Takebe, Vivekananda Datta, Alice P. Chen, Alida Palmisano, Elad Sharon, David J. Sims, and Stephen Leong
- Subjects
0301 basic medicine ,medicine.medical_specialty ,Genomic profiling ,business.industry ,education ,Disease progression ,Cancer therapy ,Stock options ,Hematology ,Genetic pathways ,Clinical trial ,03 medical and health sciences ,030104 developmental biology ,0302 clinical medicine ,Oncology ,030220 oncology & carcinogenesis ,Family medicine ,medicine ,Tumor biopsy ,business ,health care economics and organizations ,After treatment - Abstract
Background Emerging clinical data show prediction of response to therapies targeting specific genetic aberrations have unexpectedly variable outcomes. This multicenter, double-blind, randomized trial opened in 2013 to compare response rates (RR) between 2 groups of patients (pts) identified to have an actionable mutation of interest (aMOI) in one of 3 genetic pathways (DNA repair, PI3K, or RAS/RAF/MEK): group A) Pts treated with agent(s) targeting one selected pathway (experimental arm-A) and B) Pts treated with agent(s) not targeting that pathway (control arm-B). Based on the data available at the time, the aMOI selection criteria encompassed alterations throughout the entire selected pathway instead of specific genetic changes. Methods Primary objective is to compare the RR (CR and PR) and 4 months PFS between treatments arms A and B. A CLIA-certified genetic analysis of a fresh tumor biopsy was performed at entry. Pts with an aMOI were randomized 2:1 to arm A vs. B. Study drugs were: 1) DNA repair-a) veliparib & temozolomide (VT); b) AZD1775 & carboplatin (AC); pts with p53 mutations were preferentially selected for AC; 2) PI3K- everolimus (E); 3) RAS- trametinib(T). At disease progression, Arm B pts could cross over to their target arm (A). Results 193 pts underwent biopsies; >90% of samples completed DNA sequencing. 96 pts (50%) had an aMOI and were randomized to treatment. Cohort VT had insufficient accrual on the experimental arm to be evaluable. AC, E and T cohorts were closed due to futility. Enrollment rate after treatment assignment was 77% for Arm A and 53%, for arm B. Attrition analysis between arms A and B are ongoing. Conclusions The increasing availability of genetic sequencing and bias toward expected benefit of highly specific treatment agents may account for the large number pt withdrawal from Arm A. This imbalance made comparison of arms A and B uninterpretable. The trial has been amended to employ a non-randomized design to complete the assessment of VT’s activity. Analysis of the aMOIs are ongoing to develop a more stringent selection criteria for future precision medicine trials. Clinical trial identification NCT01827384. Editorial acknowledgement Christina Rosenberger, PhD. Legal entity responsible for the study NCI. Funding NCI. Disclosure S. Kummar: Advisory / Consultancy: Corvus Pharmaceuticals; Advisory / Consultancy: MedTree; Advisory / Consultancy: Nodus Therapeutics; Advisory / Consultancy: Genentech; Advisory / Consultancy: ShangPharma Innovation; Advisory / Consultancy: Seattle Genetics; Travel / Accommodation / Expenses: Bayer; Shareholder / Stockholder / Stock options: Dhrishti Inc.; Research grant / Funding (institution): Bristol-Myers Squibb; Research grant / Funding (institution): Dynavax; Research grant / Funding (institution): Pfizer; Research grant / Funding (institution): Loxo; Research grant / Funding (institution): Corvus Pharmaceuticals; Research grant / Funding (institution): Plexxikon; Research grant / Funding (institution): Jounce Therapeutics; Research grant / Funding (institution): ADC Therapeutics; Research grant / Funding (institution): Advenchen Laboratories; Research grant / Funding (institution): Incyte; Research grant / Funding (institution): Taiho Pharmaceutical. N. Moore: Licensing / Royalties: Nestle Nutrition. P. Williams: Licensing / Royalties, I was a co-inventor of the DLBCL cell of origin patent recently filed by the NIH: NIH; Research grant / Funding (institution): Illumina. K.P.S. Raghav: Advisory / Consultancy: Bayer; Travel / Accommodation / Expenses: TRACON Pharma; Honoraria (self): Bayer; Honoraria (self): Eisai. F. Meric-Bernstam: Advisory / Consultancy: Genentech; Advisory / Consultancy: Inflection Biosciences; Advisory / Consultancy: Pieris Pharmaceuticals; Advisory / Consultancy: Clearlight Diagnostics; Advisory / Consultancy: Darwin Health; Advisory / Consultancy: Samsung Bioepis; Advisory / Consultancy: Spectrum Pharmaceuticals; Advisory / Consultancy: Aduro Biotech; Advisory / Consultancy: Origimed; Advisory / Consultancy: Xencor; Advisory / Consultancy: Debiopharm Group; Advisory / Consultancy: Mersana; Honoraria (self): Sumitomo Group; Honoraria (self): Dialectica; Research grant / Funding (self): Novartis; Research grant / Funding (self): AstraZeneca; Research grant / Funding (self): Taiho Pharmaceutical; Research grant / Funding (self): Genentech; Research grant / Funding (self): Calithera Biosciences; Research grant / Funding (self): Debiopharm Group. S. Leong: Shareholder / Stockholder / Stock options: Antares Pharmaceuticals; Shareholder / Stockholder / Stock options: Spectrum Pharmaceuticals; Honoraria (self): Lilly; Research grant / Funding (institution): Deciphera; Research grant / Funding (institution): Karyopharm Therapeutics; Research grant / Funding (institution): Bristol-Myers Squibb; Research grant / Funding (institution): Lilly. S. Waqar: Research grant / Funding (institution): Spectrum Pharmaceuticals; Research grant / Funding (institution): Lilly; Research grant / Funding (institution): Pfizer; Research grant / Funding (institution): Genentech/Roche; Research grant / Funding (institution): Daiichi Sankyo; Research grant / Funding (institution): Newlink Genetics; Research grant / Funding (institution): EMD Serono; Research grant / Funding (institution): Puma Biotechnology; Research grant / Funding (institution): Novartis; Research grant / Funding (institution): Xcovery; Research grant / Funding (institution): Synermore biologics; Research grant / Funding (institution): Celgene; Research grant / Funding (institution): Vertex; Research grant / Funding (institution): Bristol-Myers Squibb; Research grant / Funding (institution): Stem CentRx; Research grant / Funding (institution): Hengrui Therapeutics; Research grant / Funding (institution): Checkpoint Therapeutics; Research grant / Funding (institution): Ignyta; Research grant / Funding (institution): AstraZeneca; Research grant / Funding (institution): ARIAD. All other authors have declared no conflicts of interest.
- Published
- 2019
29. Exosome-based detection of activating and resistance
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Elena, Castellanos-Rizaldos, Xuan, Zhang, Vasisht R, Tadigotla, Dominik G, Grimm, Chris, Karlovich, Luis E, Raez, and Johan K, Skog
- Subjects
liquid biopsy ,exoNA ,exosomes ,ctDNA ,NSCLC ,respiratory tract diseases ,Research Paper - Abstract
Non-small cell lung cancer (NSCLC) is the most prevalent form of lung cancer and its molecular landscape has been extensively studied. The most common genetic alterations in NSCLC are mutations within the epidermal growth factor receptor (EGFR) gene, with frequencies between 10-40%. There are several molecular targeted therapies for patients harboring these mutations. Liquid biopsies constitute a flexible approach to monitor these mutations in real time as opposed to tissue biopsies that represent a single snap-shot in time. However, interrogating cell free DNA (cfDNA) has inherent biological limitations, especially at early or localized disease stages, where there is not enough tumor material released into the patient’s circulation. We developed a qPCR- based test (ExoDx EGFR) that interrogates mutations within EGFR using Exosomal RNA/DNA and cfDNA (ExoNA) derived from plasma in a cohort of 110 NSCLC patients. The performance of the assay yielded an overall sensitivity of 90% for L858R, 83% for T790M and 73% for exon 19 indels with specificities of 100%, 100%, and 96% respectively. In a subcohort of patients with extrathoracic disease (M1b and MX) the sensitivities were 92% (L858R), 95% (T790M), and 86% (exon 19 indels) with specificity of 100%, 100% and 94% respectively.
- Published
- 2019
30. Clinical Applications of Next-Generation Sequencing in Precision Oncology
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P. Mickey Williams and Chris Karlovich
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0301 basic medicine ,Cancer Research ,medicine.medical_treatment ,MEDLINE ,Computational biology ,Medical Oncology ,DNA sequencing ,Article ,Targeted therapy ,03 medical and health sciences ,0302 clinical medicine ,Biomarkers, Tumor ,Medicine ,Humans ,Precision Medicine ,Reimbursement ,business.industry ,Computational Biology ,High-Throughput Nucleotide Sequencing ,Genomics ,Molecular analysis ,Patient management ,030104 developmental biology ,Oncology ,Precision oncology ,030220 oncology & carcinogenesis ,Molecular Profile ,business - Abstract
The ability of next-generation sequencing (NGS) to comprehensively assess the molecular profile of a tumor specimen has transformed the clinical testing landscape in oncology. Accordingly, recent years have seen broad uptake of clinical NGS to inform cancer patient management. However, significant challenges remain. The annotation and clinical interpretation of variants identified by NGS tests often require rigorous review and may vary between laboratories. While a clearer regulatory path has emerged, reimbursement for NGS tests remains a subject of continuing debate. Basket clinical studies such as the National Cancer Institute Molecular Analysis of Therapy Choice are evaluating the degree to which matching of a targeted therapy to tumor molecular profile by NGS can be applied independently of tissue histology. Newer applications of NGS such as for circulating tumor DNA testing and to identify novel RNA fusion driver events continue to expand its clinical utility.
- Published
- 2019
31. Molecular Features of Cancers Exhibiting Exceptional Responses to Treatment
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Hui Shen, Louis M. Staudt, Viktoriya Korchina, Katherine A. Hoadley, Roy Tarnuzzer, Richard F. Little, Manuel Castro de Moura, Maria F. Cardenas, Irina A. Lubensky, Manel Esteller, Jianhong Hu, Julie M. Gastier-Foster, Jeffrey White, Elise C. Kohn, Paula M. Jacobs, Ninad Dewal, Chris Karlovich, Carol J. Weil, Donna M. Muzny, Geraldine O'Sullivan-Coyne, Lalitha K. Shankar, Adrienne Johnson, Kristen M. Leraas, Alice P. Chen, Jean C. Zenklusen, Shakun Malik, Peter W. Laird, Barbara A. Conley, Harshavardhan Doddapaneni, Lisa M. McShane, Jay Bowen, Vincent A. Miller, S. Percy Ivy, James V. Tricoli, David Piñeyro, Toshinori Hinoue, Tracy S. Nolan, Brian Rodgers, Lyndsay Harris, James H. Doroshow, David A. Wheeler, Paul Williams, Linghua Wang, Alina M Hamilton, Elijah F. Edmondson, and Naoko Takebe
- Subjects
Male ,0301 basic medicine ,Cancer Research ,Biopsy ,Antineoplastic Agents ,Synthetic lethality ,Epigenesis, Genetic ,03 medical and health sciences ,0302 clinical medicine ,Immune system ,Neoplasms ,Combination cancer therapy ,Tumor Microenvironment ,medicine ,Humans ,Gene Regulatory Networks ,Epigenetics ,Exome sequencing ,Tumor microenvironment ,business.industry ,Genetic Variation ,Cancer ,Genomics ,Cell Biology ,Prognosis ,medicine.disease ,Exceptional Responder ,Survival Analysis ,Treatment Outcome ,030104 developmental biology ,Oncology ,030220 oncology & carcinogenesis ,Cancer research ,Female ,business - Abstract
A small fraction of cancer patients with advanced disease survive significantly longer than patients with clinically comparable tumors. Molecular mechanisms for exceptional responses to therapy have been identified by genomic analysis of tumor biopsies from individual patients. Here, we analyzed tumor biopsies from an unbiased cohort of 111 exceptional responder patients using multiple platforms to profile genetic and epigenetic aberrations as well as the tumor microenvironment. Integrative analysis uncovered plausible mechanisms for the therapeutic response in nearly a quarter of the patients. The mechanisms were assigned to four broad categories���DNA damage response, intracellular signaling, immune engagement, and genetic alterations characteristic of favorable prognosis���with many tumors falling into multiple categories. These analyses revealed synthetic lethal relationships that may be exploited therapeutically and rare genetic lesions that favor therapeutic success, while also providing a wealth of testable hypotheses regarding oncogenic mechanisms that may influence the response to cancer therapy. Profiling multi-platform genomics of 110 cancer patients with an exceptional therapeutic response, Wheeler et al. identify putative molecular mechanisms explaining this survival phenotype in ���23% of cases. Therapeutic success is related to rare molecular features of responding tumors, exploiting synthetic lethality and oncogene addiction.
- Published
- 2021
32. Circulating tumour DNA profiling reveals heterogeneity of EGFR inhibitor resistance mechanisms in lung cancer patients
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Maximilian Diehn, Jonathan W. Goldman, Chris Karlovich, Heather A. Wakelee, W. Thomas Purcell, Henning Stehr, Florian Scherer, Kathleen A. Durkin, David M. Kurtz, Alexander F. Lovejoy, Joel W. Neal, Mohammad Shahrokh Esfahani, Andrew Simmons, Lecia V. Sequist, Jacob J. Chabon, Zofia Piotrowska, Gregory A. Otterson, D. Ross Camidge, Ash A. Alizadeh, Thomas Harding, Henry J. Haringsma, and Aaron M. Newman
- Subjects
0301 basic medicine ,Tumour heterogeneity ,Science ,General Physics and Astronomy ,medicine.disease_cause ,Bioinformatics ,General Biochemistry, Genetics and Molecular Biology ,03 medical and health sciences ,T790M ,0302 clinical medicine ,medicine ,Osimertinib ,Rociletinib ,Epidermal growth factor receptor ,neoplasms ,EGFR inhibitors ,Multidisciplinary ,Crizotinib ,biology ,General Chemistry ,3. Good health ,030104 developmental biology ,030220 oncology & carcinogenesis ,Cancer research ,biology.protein ,KRAS ,medicine.drug - Abstract
Circulating tumour DNA (ctDNA) analysis facilitates studies of tumour heterogeneity. Here we employ CAPP-Seq ctDNA analysis to study resistance mechanisms in 43 non-small cell lung cancer (NSCLC) patients treated with the third-generation epidermal growth factor receptor (EGFR) inhibitor rociletinib. We observe multiple resistance mechanisms in 46% of patients after treatment with first-line inhibitors, indicating frequent intra-patient heterogeneity. Rociletinib resistance recurrently involves MET, EGFR, PIK3CA, ERRB2, KRAS and RB1. We describe a novel EGFR L798I mutation and find that EGFR C797S, which arises in ∼33% of patients after osimertinib treatment, occurs in MET copy number is the most frequent rociletinib resistance mechanism in this cohort and patients with multiple pre-existing mechanisms (T790M and MET) experience inferior responses. Similarly, rociletinib-resistant xenografts develop MET amplification that can be overcome with the MET inhibitor crizotinib. These results underscore the importance of tumour heterogeneity in NSCLC and the utility of ctDNA-based resistance mechanism assessment.
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- 2016
33. Abstract 3916: Patient-derived organoid and cell culture models from the NCI Patient-Derived Models Repository (NCI PDMR) preserve genomic stability and heterogeneity of patient tumor specimens
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Luis E. Romero, Kaitlyn Arthur, Robin D. Harrington, Justine N. McCutcheon, Brandie A. Fullmer, Gloryvee Rivera, Li Chen, Savanna Styers, Matthew R. Murphy, Lindsay Dutko, Rajesh Patidar, Kelly Benauer, Alyssa K. Chapman, Marion Gibson, Abigail Walke, Carrie Bonomi, Vishnuprabha R. Kannan, Luke H. Stockwin, Paul Williams, Tomas Vilimas, Kelly Dougherty, Amanda Peach, Jenna Moyer, Biswajit Das, Michelle M. Gottholm-Ahalt, Peng Wang, James H. Doroshow, Nikitha Nair, Erin Cantu, Kelsey A. Conley, Melinda G. Hollingshead, Anna Wade, Thomas Forbes, Anna J. Lee Fong, Kevin Plater, Joseph P. Geraghty, Mariah Baldwin, Dianne L. Newton, Yvonne A. Evrard, Shahanawaz Jiwani, Chris Karlovich, and Michael Mullendore
- Subjects
Cancer Research ,Cancer ,Variant allele ,Early passage ,Biology ,medicine.disease ,Tumor tissue ,Tumor heterogeneity ,Genomic Stability ,Oncology ,Copy Number Alteration ,Cell culture ,medicine ,Cancer research - Abstract
Background: The National Cancer Institute (NCI) has developed a Patient-Derived Models Repository (PDMR; https://pdmr.cancer.gov) of preclinical models including patient-derived xenografts (PDX), organoids (PDOrg) and patient-derived cell cultures (PDC). Extensive clinical annotation and genomic datasets are available for these preclinical models. However, it is unclear if the molecular profiles of the corresponding patient tumors are stably propagated in these models. We have previously demonstrated that PDX models from the NCI PDMR faithfully represent the patient tumors both in terms of genomic stability and tumor heterogeneity. Here, we conduct an in-depth investigation of genomic representation of patient tumors in the PDOrgs and PDCs. Methods: PDOrgs (n=64) and PDCs (n=94) were established from tumor fragments (i.e., initiator specimens) obtained either from patient specimens or from PDX specimens of early passage. For some models (n=19), both PDOrgs and PDCs were generated from the same tumor tissue; in fewer cases (n=4), PDCs were established from organoids derived from patient specimens. Whole Exome Sequencing and RNA-Seq were performed on all PDCs and PDOrgs, and data were compared with patient specimens or early passage PDXs. Results: A majority of the PDOrgs and PDCs have stably inherited the genome of the corresponding patient specimens based on the following observations: (1) >87% of PDOrgs and PDCs maintained similar copy number alteration profiles compared with the initiator specimens of the preclinical model; (2) the variant allele frequency (VAF) of clinically relevant mutations remained consistent between the PDOrgs, PDCs, and the initiator specimens, with none of the PDCs or PDOrgs deviating by >15% VAF; and (3) clinically relevant biomarkers (e.g., MSI, LOH, mutational signatures etc.) are concordant amongst the PDOrgs, PDCs, and the initiator specimens. We observed that the majority of SNVs and indels present in the initiator specimens were also found in the PDOrgs and PDCs, suggesting almost all the tumor heterogeneity was preserved in these preclinical models. Conclusions: This large and histologically diverse set of PDOrgs and PDCs from the NCI PDMR exhibited genomic stability and faithfully represented the tumor heterogeneity observed in corresponding patient specimens. These preclinical models thus represent a valuable resource for researchers interested in pre-clinical drug or other studies. Citation Format: Biswajit Das, Yvonne A. Evrard, Li Chen, Rajesh Patidar, Tomas Vilimas, Justine N. McCutcheon, Amanda L. Peach, Nikitha V. Nair, Thomas D. Forbes, Brandie A. Fullmer, Anna J. Lee Fong, Luis E. Romero, Alyssa K. Chapman, Kelsey A. Conley, Robin D. Harrington, Shahanawaz S. Jiwani, Peng Wang, Michelle M. Gottholm-Ahalt, Erin N. Cantu, Gloryvee Rivera, Lindsay M. Dutko, Kelly M. Benauer, Vishnuprabha R. Kannan, Carrie A. Bonomi, Kelly M. Dougherty, Joseph P. Geraghty, Marion V. Gibson, Savanna S. Styers, Abigail J. Walke, Jenna E. Moyer, Anna Wade, Mariah L. Baldwin, Kaitlyn A. Arthur, Kevin J. Plater, Luke Stockwin, Matthew R. Murphy, Michael E. Mullendore, Dianne L. Newton, Melinda G. Hollingshead, Chris A. Karlovich, Paul M. Williams, James H. Doroshow. Patient-derived organoid and cell culture models from the NCI Patient-Derived Models Repository (NCI PDMR) preserve genomic stability and heterogeneity of patient tumor specimens [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 3916.
- Published
- 2020
34. Abstract 3554: Genomic landscape of acquired uniparental disomy in NCI PDMR patient derived xenograft models
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Li Chen, Yvonne A. Evrard, Michelle Eugeni, Kelly Benauer, Amanda Peach, Michelle M. Gottholm Ahalt, James H. Doroshow, Shahanawaz Jiwani, Biswajit Das, Chris Karlovich, John Carter, Candace Mallow, Tara Grinnage-Pulley, Lindsay Dutko, Tiffanie L. Miner, Sergio Y. Alcoser, Dianne L. Newton, Devynn Breen, Thomas Forbes, Emily Delaney, Alyssa K. Chapman, Marianne Radzyminski, Anna J. Lee Fong, Shannon Uzelac, Chelsea McGlynn, Tomas Vilimas, Brandie A. Fullmer, Luis E. Romero, Gloryvee Rivera, Suzanne Borgel, Robin D. Harrington, Justine N. McCutcheon, Rajesh Patidar, Jesse Stottlemyer, Vishnuprabha R. Kannan, Nikitha Nair, Erin Cantu, Peng Wang, Melinda G. Hollingshead, Kelsey A. Conley, and Paul Williams
- Subjects
Cancer Research ,Haplotype ,Chromosome ,Cancer ,Biology ,medicine.disease ,Uniparental disomy ,Loss of heterozygosity ,Clear cell renal cell carcinoma ,Oncology ,Cancer research ,Carcinoma ,medicine ,Exome sequencing - Abstract
Background: Acquired Uniparental Disomy (aUPD) is relatively common in cancer. Occurrence of aUPD is more frequent in some tumor histologies (e.g., serous ovarian, colorectal) and may be relevant for choice of therapy. The Patient-Derived Models Repository (PDMR; https://pdmr.cancer.gov) developed by The National Cancer Institute (NCI) includes patient-derived xenograft (PDX) models from multiple tumor histologies with different passages and lineages. The associated clinical annotation and genomic data make it possible to assess the prevalence of aUPD in the PDMR cohort and the stability of aUPD in different passages and lineages within a PDX model. Methods: High tumor purity in the PDX specimens (after removal of mouse reads representing the stroma) enabled highly accurate assessment of loss of heterozygosity (LOH). Variants called by GATK Haplotype caller from whole exome sequencing (WES) data were used to identify segments of homozygosity using BCFtools/RoH (runs of homozygosity). The RoH segments were then intersected with the bed file for chromosome arms to get %LOH at the arm level. If %LOH on a chromosome arm was >90%, we considered the sample to have aUPD at the arm level. WES was also used to look for associations between DNA damage repair (DDR) pathway alterations and aUPD. Results: We made the following observations: a) aUPD was observed most frequently in chr18q (75/427, 17.6%) and chr3p (69/427, 16%) of PDX models; b) aUPD was observed more frequently in certain tumor histologies, e.g., clear cell renal cell carcinoma (6/8), small cell lung cancer (3/4) and non-small cell lung cancer (25/38); c) extensive aUPD was observed in 4 PDMR models (>50% of evaluated chromosome arms in these models have aUPD); d) aUPD was not observed in some tumor histologies, i.e., synovial sarcoma, uterine endometrioid carcinoma; e) in the vast majority of PDMR models (>90%), aUPD is maintained faithfully across lineages and through multiple passaging; f) subclonal aUPD events were observed in some models across different lineages; g) significant enrichment of double strand DNA break repair (DSBR) pathway alterations was observed in PDMR models without aUPD (p=0.0007, Fisher's exact test) suggesting defects in DSBR are not associated with aUPD; and h) aUPD was rarely observed in MSI-high models (1/30) suggesting mutual exclusivity of mismatch repair (MMR) pathway defects and aUPD. Conclusion: We observed a relatively high frequency of UPD in the PDMR models (at least 1 arm of a chromosome). UPD was more frequently observed in specific chromosomal arms. The frequency of aUPD was higher in some tumor histologies and absent in others. aUPD was stably maintained across passages and lineages, although some heterogeneity was observed. Our data suggest aUPD is not associated with defects in DSBR and MMR pathways. Preclinical drug studies using NCI PDMR models may suggest appropriate therapeutic options for cancers with aUPD. Citation Format: Rajesh Patidar, Li Chen, Chris A. Karlovich, Biswajit Das, Yvonne A. Evrard, Tomas Vilimas, Justine N. McCutcheon, Amanda L. Peach, Nikitha V. Nair, Thomas D. Forbes, Brandie A. Fullmer, Anna J. Lee Fong, Luis E. Romero, Alyssa K. Chapman, Kelsey A. Conley, Robin D. Harrington, Shahanawaz S. Jiwani, Peng Wang, Michelle M. Gottholm Ahalt, Erin N. Cantu, Gloryvee Rivera, Lindsay M. Dutko, Kelly M. Benauer, Vishnuprabha R. Kannan, Suzanne D. Borgel, John P. Carter, Jesse M. Stottlemyer, Tiffanie L. Miner, Devynn R. Breen, Emily T. Delaney, Chelsea A. McGlynn, Candace N. Mallow, Marianne Radzyminski, Shannon N. Uzelac, Sergio Y. Alcoser, Tara L. Grinnage-Pulley, Michelle A. Eugeni, Dianne L. Newton, Melinda G. Hollingshead, Paul M. Williams, James H. Doroshow. Genomic landscape of acquired uniparental disomy in NCI PDMR patient derived xenograft models [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 3554.
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- 2020
35. Abstract 5056: Quality control efforts in a large-scale, preclinical trial of rare cancer PDXs by the National Cancer Institute's patient-derived models repository (NCI PDMR)
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John Carter, Dianne L. Newton, Shannon Uzelac, Gloryvee Rivera, Michael Mullendore, Malorie Morris, Abigail Walke, Lily Chen, Thomas Forbes, Kyle Georgius, Emily Delaney, Sergio . Y. Alcoser, Debbie Trail, Tom Walsh, Raymond Divelbiss, Tara Grinnage-Pulley, Shahanawaz Jiwani, Chris Karlovich, Yvonne A. Evrard, Lyndsay Dutko, Sierra Hoffman, Devynn Breen, Biswajit Das, Howard Stotler, Rajesh Patidar, Thomas Vilimas, Peng Wang, Justine Mills, Suzanne Borgel, Nicole Walters, Melinda G. Hollingshead, James Doroshow, Savanna Styers, Jesse Stottlemyer, Tiffanie Chase, Michelle M. Gottholm-Ahalt, Alice P. Chen, Kristen Cooley, and P. Mickey Williams
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Cancer Research ,medicine.medical_specialty ,Scale (ratio) ,business.industry ,media_common.quotation_subject ,Cancer ,medicine.disease ,Rare cancer ,Oncology ,Medicine ,Quality (business) ,Medical physics ,business ,media_common - Abstract
The National Cancer Institute's Patient-Derived Models Repository (NCI PDMR; https://pdmr.cancer.gov) is performing a large-scale multi-year preclinical study with 39 PDX models of rare cancers (mesothelioma, MPNST, osteosarcoma, Merkel cell carcinoma, etc) treated with 56 novel therapeutic combinations in an exploratory, n-of-4 arm, study design. Combinations that show promising responses (e.g., regression or durable inhibition of tumor growth) will be repeated along with the single agent arms to determine if the response is driven by the combination or only one of the agents. In order to do this in a timely fashion, relatively speaking, the PDX tumors are serially passaged and each passage is treated with a set of 8 combinations plus relevant vehicle control(s) while in parallel enough PDXs are retained to be expanded for the next passage and drug set. Every serial passage undergoes several quality control assessments that serve as go/no-go criteria including pathology assessment, human:mouse DNA content assessment, and low pass whole genome sequencing to determine the average fraction of genome changed compared to the original donor material. If there is a QC failure, the PDX model is restarted from early passage cryo-material (passage 1-2). An additional quality control effort is to bookend the combination studies with the first set of agents to see if tumor response is similar across passages. To date, most of the models have demonstrated a high degree of stability, though a couple of models have moved toward murine content and have been restarted from early passage material so all drug combinations can be tested. DNA and RNA are retained from all passages so a full NGS evaluation can be performed at a later date. This effort has been ongoing for over a year and the first bookend studies are beginning to be tested to determine if response at first and last passage of the study are consistent with each other, given the constraints of the inherent heterogeneity of the models themselves. Single agent studies of drug combinations that demonstrated a response in 30%-50% of the models tested are also underway to determine which combinations have a more than additive effect compared to the single agents. Promising combinations will be moved forward to early phase clinical trials for these rare cancers. Funded by NCI Contract No. HHSN261200800001E Citation Format: Yvonne A. Evrard, Biswajit Das, Sergio Y. Alcoser, Suzanne Borgel, Devynn Breen, John Carter, Tiffanie Chase, Alice Chen, Lily Chen, Kristen Cooley, Emily Delaney, Raymond Divelbiss, Lyndsay Dutko, Thomas Forbes, Kyle Georgius, Michelle Gottholm-Ahalt, Tara Grinnage-Pulley, Sierra Hoffman, Chris Karlovich, Shahanawaz Jiwani, Justine Mills, Malorie Morris, Michael Mullendore, Dianne Newton, Rajesh Patidar, Gloryvee Rivera, Howard Stotler, Jesse Stottlemyer, Savanna Styers, Debbie Trail, Shannon Uzelac, Thomas Vilimas, Abigail Walke, Thomas Walsh, Nicole Walters, Peng Wang, P. Mickey Williams, Melinda Hollingshead, James H. Doroshow. Quality control efforts in a large-scale, preclinical trial of rare cancer PDXs by the National Cancer Institute's patient-derived models repository (NCI PDMR) [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 5056.
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- 2020
36. Genomic characterization of preclinical models derived from primary and metastatic sites from rapid autopsy patients in PDMR
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Paul Williams, Lindsay Dutko, Marianne Radzyminski, Li Chen, Yvonne A. Evrard, Melinda G. Hollingshead, Anna Lee Fong, Luis Romero, James H. Doroshow, Tomas Vilimas, Dianne L. Newton, Rajesh Patidar, Shahanawaz Jiwani, Kelly Benauer, Chris Karlovich, Michelle Eugeni, Nikitha Nair, Biswajit Das, Kelly Dougherty, and Amanda Peach
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Oncology ,Cancer Research ,medicine.medical_specialty ,business.industry ,Internal medicine ,medicine ,Cancer ,Rapid autopsy ,medicine.disease ,business - Abstract
e13506 Background: The National Cancer Institute has developed a repository of preclinical models [Patient-Derived Models Repository (NCI PDMR, https://pdmr.cancer.gov )] including patient derived xenografts (PDXs), organoids (PDOrgs) and in vitro tumor cultures (PDCs) from patients with solid tumor cancer histologies. A subset of these preclinical models is derived from post-mortem collections from rapid autopsies representing the end point in disease progression. Clinical annotations and genomic datasets associated with these models provide a unique opportunity to study tumor evolution, mechanistic insights into the metastatic process, and treatment resistance. Methods: To date, 43 PDXs, 21 PDCs, and 23 PDOrgs using rapid autopsy specimens from 8 primary and 35 metastatic sites of 18 patients have been developed by the Biological Testing Branch (DTP, DCTD, NCI Frederick, MD) for the PDMR. Whole exome (WES) and total transcriptome (RNASeq) data were processed to generate mutation, copy number alteration (CNA) and gene expression data. Multi-model lineage trees were reconstructed based on putative somatic variants for all the models derived from all patients. The fraction of the genome affected by CNA was compared both within and across PDX models. Results: Most of the rapid autopsy PDX models (32/43) are derived from pancreatic adenocarcinoma (PAAD) patients (13/18), with metastatic specimens originating from sites including liver, colon, omentum, and lung. Driver mutations are present in all preclinical model specimens derived from the same patient. For instance, KRAS p.G12D is present in all patient-derived model specimens derived from PAAD patient 521955. The fraction of the genome affected by CNA remains stable within a PDX model across passages (n = 24, mean = 6.39%, sd = 5.90%). However, we found that this increased when comparing PDX models derived from metastatic sites versus the primary site (n = 19, mean = 16.92%, sd = 10.46%). This indicates presence of tumor heterogeneity between metastatic and primary sites. The lineage tree for models from patient 521955 indicates that one liver metastasis has a unique seeding event compared to the other 4 metastatic sites. Unsupervised clustering analysis on gene expression data also confirms the observed tumor site relationships. Conclusions: Our data demonstrate the potential use of these preclinical models available from the NCI PDMR. These models provide a unique resource for preclinical studies in tumor evolution, metastatic spread mediators, and drug resistance.
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- 2020
37. Abstract A079: National Cancer Institute Molecular Analysis for Therapy Choice (NCI-MATCH): A successful precision medicine signal-seeking trial in patients (pts) with rare variants and refractory malignancies
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Barbara A. Conley, James V. Tricoli, Keith T. Flaherty, Richard F. Little, Lyndsay Harris, Alice P. Chen, Carlos L. Arteaga, Paul Williams, Robert Gray, Shuli Li, Stanley R. Hamilton, Larry Rubinstein, Edith P. Mitchell, Chris Karlovich, Peter J. O'Dwyer, Nci-Match team, David R. Patton, and Lisa M. McShane
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Oncology ,Trametinib ,Cancer Research ,medicine.medical_specialty ,education.field_of_study ,business.industry ,Afatinib ,Population ,Cancer ,Dabrafenib ,medicine.disease ,Precision medicine ,Clinical trial ,Internal medicine ,medicine ,Nivolumab ,business ,education ,medicine.drug - Abstract
Introduction: NCI-MATCH, developed by ECOG-ACRIN & NCI, is the largest precision medicine study for pts with refractory malignancy. Over 1100 clinical sites in the National Clinical Trials Network enrolled pts. The purpose of the study is to identify potentially beneficial targeted treatments across tumor types with similar molecular abnormalities. Methods: The NCI Central IRB approved NCI-MATCH. Pts with refractory/no treatment available solid tumors, lymphomas or myelomas had a fresh biopsy profiled by next generation sequencing (143 genes, > 4000 single nucleotide variants, indels, amplifications & targeted fusions). Pts are assigned by a defined algorithm to treatments with evidence of activity against tumors with the relevant molecular alteration. Pts are excluded if a treatment is FDA approved or known to be ineffective for their malignancy. After successfully sequencing fresh biopsies from 5540 pts, subprotocols with extremely rare variants lacked sufficient accrual. To address actionable variants with a prevalence of < 1.5%, we decided to accept clinical sequencing results from 30 commercial and academic laboratories vetted by NCI-MATCH to address relevant variants. These labs notify clinicians participating in NCI-MATCH if their pt’s tumor contains an actionable variant. Treatment continues until tumors became refractory, pt intolerance or withdrawal of consent. An objective response rate (ORR by RECIST) of > 16% among 31 eligible patients is considered a positive signal. Results: After screening 5540 pts, 37.6% had an actionable variant. After histology and treatment-specific exclusions, 17.8% were assigned and 69.5% enrolled on the assigned subprotocol. 11 of the initial 30 subprotocols reached completion with adequate follow-up. Of the first 11 evaluable subprotocols, 3 addressing rare variants had a positive signal: Nivolumab in pts with loss of expression of MLH1 or MSH2 (ORR 36%), capivasertib in pts with AKT mutations (ORR 23%), and dabrafenib + trametinib in pts with BRAF V600 mutations (ORR 33%). These molecular variants were found in 2%, 1.2% and 1.9% respectively, of screened pts. Two other subprotcocols (afatinib in ERBB2 mutations and AZD4547 in FGFR abnormalities) showed responses in rare tumors or specific variant subsets, respectively. As of July 15, 2019, an additional 378 of 432 (88%) pts have been assigned to a treatment with a clinical sequencing assay; 83% of these pts enrolled to 1 of 24 subprotocols, allowing completion of an additional 9 of the original 30 subprotocols and complete accrual to 2 of 5 recently added subprotocols. Four of 35 subprotocols closed for lack of accrual, 10 continue accruing and 4 are planned. Conclusions: Platform precision medicine trials can identify potentially useful targeted treatments for diverse malignancies in pts with uncommon tumors & rare actionable variants, an unmet need. In a population of pts with refractory cancers, lymphomas and myelomas, 30-40% will have an actionable variant for targeted treatment (investigational or standard). Of the first 11 subprotocols with adequate follow-up, 3 (27%) showed a positive signal and an additional 2 showed responses in rare tumors or in a molecular subset, suggesting that the NCI-MATCH trial approach identifies useful targets for further exploration. Citation Format: Lyndsay N Harris, Robert J Gray, Barbara A Conley, Alice P Chen, Keith T Flaherty, Stanley R Hamilton, Paul M Williams, Chris Karlovich, David Patton, Shuli Li, Lisa M McShane, Larry V Rubinstein, Edith P Mitchell, James V Tricoli, Richard F Little, Carlos L Arteaga, Peter J O'Dwyer, NCI-MATCH team. National Cancer Institute Molecular Analysis for Therapy Choice (NCI-MATCH): A successful precision medicine signal-seeking trial in patients (pts) with rare variants and refractory malignancies [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference on Molecular Targets and Cancer Therapeutics; 2019 Oct 26-30; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2019;18(12 Suppl):Abstract nr A079. doi:10.1158/1535-7163.TARG-19-A079
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- 2019
38. The PRIME Trial: PARP Inhibition in IDH Mutant Effectiveness Trial. a Phase II Study of Olaparib in Isocitrate Dehydrogenase (IDH) Mutant Relapsed/Refractory Acute Myeloid Leukemia and Myelodysplastic Syndrome
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Namrata S Chandhok, Chris Karlovich, Michael E. Berens, Thomas Prebet, Jing Li, Ranjit S. Bindra, S. Percy Ivy, Stephanie Halene, Yu Shyr, and Wei Wei
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Oncology ,medicine.medical_specialty ,business.industry ,Immunology ,Phases of clinical research ,Myeloid leukemia ,Cell Biology ,Hematology ,medicine.disease ,Biochemistry ,Olaparib ,Clinical trial ,Leukemia ,chemistry.chemical_compound ,medicine.anatomical_structure ,chemistry ,Internal medicine ,PARP inhibitor ,medicine ,Clinical endpoint ,Bone marrow ,business - Abstract
Background: Recurring mutations have been identified in acute myeloid leukemia (AML) and myelodysplastic syndrome (MDS) which translate to therapeutic targets. Isocitrate dehydrogenase-1 and -2 (IDH1/2) mutations occur in ~20% of AML, and up to 12% of patients with MDS. Three conserved mutational hotspots in the IDH enzymes alter their function and lead to the production of (R)-2-hydroxyglutarate (2HG), an oncometabolite with numerous downstream effects, including impaired DNA damage repair. Specifically, homologous recombination (HR) is impaired by inhibiting the function of histone demethylases that are critical for HR and recruitment of the HR machinery to sites of DNA damage. In HR deficient tumors poly-ADP ribose polymerase (PARP) enzymes mediate a key salvage pathway. PARP inhibition in HR deficient tumors leads to synthetic lethality via simultaneous inhibition of HR and SSB mediated DNA repair. Our group previously demonstrated synthetic lethality with PARP inhibition in IDH mutant cells lines, and other IDH mutant models including primary patient-derived cell lines and genetically-matched tumor xenografts. Study Design and Methods: The PRIME trial (NCI10264) is a proof of concept, biomarker-driven, multi-institution, phase II open label clinical trial to assess the overall response of IDH1/2 mutant relapsed/refractory AML and MDS to PARP inhibitor monotherapy with olaparib. The clinical trial is executed by the Experimental Therapeutics Clinical Trials Network of the NCI. The Cancer Therapy Evaluation Program will provide olaparib. Eligibility criteria include documented IDH1 or IDH2 mutation in blood or bone marrow within 30 days of enrollment based on mutational testing by PCR or sequencing in a CLIA certified laboratory and willingness to undergo a bone marrow biopsy. Patients will be treated with olaparib 300 mg q12hrs each day of a 28-day cycle, using a tablet formulation, until disease progression, unacceptable toxicity, withdrawal of consent or death. Blood and bone marrow samples for 2-HG analysis will be collected prior to starting therapy and after 1 cycle (28 days), cycle 2, 3, 6, 9, 12 or when there is concern for disease progression (Figure 1). A Simon two-stage optimal design will be used to test the null (ORR=10%) versus the alternative hypothesis (ORR=40%) in each arm. In the first stage, 9 patients will be accrued in each arm. If one or fewer responses are observed in these 9 patients, that arm will be stopped early for futility. Otherwise, 11 additional patients will be accrued for a total of 18 in each arm. We reject the null hypothesis if at least 5 responses are observed in these 20 patients. In each arm, we have approximately 90% power to detect a 30% increase in ORR at a one-sided type I error rate of 0.05. Primary endpoint: Overall response rate (ORR) of 40%, i.e., a 30% ORR improvement (40% vs. historical control ORR = 10%) based on MDS International Working Group 2006 criteria and AML MDS International Working Group 2003 criteria after 6 cycles of treatment. Cumulative ORR will include complete remission, complete remission with incomplete blood count recovery, partial response, and bone marrow complete remission. Secondary endpoints: Progression-free survival (the interval between the time of initiation of olaparib to the time of documentation of olaparib failure or last follow-up) and overall survival (the interval between the time of initiation of olaparib to the time of death or last follow-up) for the trial. Exploratory studies: The PRIME trial will also test the utility of 2-HG and DNA damage markers such as γ-H2AX as potential biomarkers of response to olaparib. Using multiple viability assays on leukemia cell lines and bone marrow cultures we will assess synergistic therapeutic combinations to further improve outcomes in this patient population. To confirm efficacy in vivo without undue toxicity, promising combination therapies will be confirmed in cytokine-humanized immunodeficient "MISTRG" mice. We will also examine the impact of PARP inhibitors on the genomic, proteomic, metabolomic and immunologic landscape of IDH 1/2-mutant hematologic malignancies using DNA whole exome sequencing (WES), RNA-Seq, and liquid chromatography-mass spectrometry assessment of oncometabolites. Disclosures Bindra: Cybrexa: Consultancy, Equity Ownership. Prebet:pfizer: Honoraria; pfizer: Honoraria; pfizer: Honoraria; Boehringer Ingelheim: Research Funding; pfizer: Honoraria; Tetraphase: Consultancy; novartis: Honoraria; novartis: Honoraria; Genentech: Consultancy; Boehringer Ingelheim: Research Funding; novartis: Honoraria; Boehringer Ingelheim: Research Funding; Agios: Consultancy, Research Funding; Jazz Pharmaceuticals: Consultancy, Honoraria, Research Funding; pfizer: Honoraria; novartis: Honoraria; Bristol-Myers Squibb: Honoraria, Research Funding; novartis: Honoraria. OffLabel Disclosure: We will be using PARP inhibitors as a novel therapy for patients with relapsed or refractory AML and high risk MDS based on preclinical data.
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- 2019
39. Cell-Free DNA Next-Generation Sequencing Prediction of Response and Resistance to Third-Generation EGFR Inhibitor
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Chris Karlovich, Helena A. Yu, A. Karin Choquette, D. Ross Camidge, Thomas Harding, Andrew Simmons, Richard B. Lanman, Alexander I. Spira, Elena Helman, Minh Nguyen, and Darrin Despain
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0301 basic medicine ,Pulmonary and Respiratory Medicine ,Adult ,Male ,Cancer Research ,Lung Neoplasms ,medicine.drug_class ,Drug resistance ,DNA sequencing ,Tyrosine-kinase inhibitor ,03 medical and health sciences ,T790M ,0302 clinical medicine ,Carcinoma, Non-Small-Cell Lung ,Medicine ,Humans ,Rociletinib ,Neoplasm Metastasis ,Lung cancer ,EGFR inhibitors ,Aged ,Neoplasm Staging ,Aged, 80 and over ,Acrylamides ,business.industry ,Antinematodal Agents ,High-Throughput Nucleotide Sequencing ,Middle Aged ,medicine.disease ,Prognosis ,respiratory tract diseases ,ErbB Receptors ,030104 developmental biology ,Pyrimidines ,Treatment Outcome ,Oncology ,Cell-free fetal DNA ,Drug Resistance, Neoplasm ,030220 oncology & carcinogenesis ,Mutation ,Cancer research ,Female ,business ,Cell-Free Nucleic Acids - Abstract
Introduction The genomic alterations driving resistance to third-generation EGFR tyrosine kinase inhibitors (TKIs) are not well established, and collecting tissue biopsy samples poses potential complications from invasive procedures. Cell-free circulating DNA (cfDNA) testing provides a noninvasive approach to identify potentially targetable mechanisms of resistance. Here we utilized a 70-gene cfDNA next-generation sequencing test to interrogate pretreatment and progression samples from 77 EGFR-mutated non-small cell lung cancer (NSCLC) patients treated with a third-generation EGFR TKI. Patients and Methods Rociletinib was evaluated in advanced or metastatic (second line or higher) disease with EGFR T790M-positive NSCLC in the TIGER-X ( NCT01526928 ) and TIGER-2 ( NCT02147990 ) studies. Plasma samples were collected at baseline and at the time of systemic progression while receiving rociletinib. The critical exons in 70 genes were sequenced in cfDNA isolated from plasma samples to elucidate a comprehensive genomic profile of alterations for each patient. Results Plasma-based cfDNA analysis identified 93% of the initial EGFR activating and 85% of the EGFR T790M resistance mutations in pretreatment samples with detectable tumor DNA. Profiling of progression samples revealed significant heterogeneity, with different variant types (eg, mutations, amplifications, and fusions) detected in multiple genes (EGFR, MET, RB1) that may be driving resistance in patients. Novel alterations not previously described in association with resistance to third-generation TKIs were also detected, such as an NTRK1 fusion. Conclusion cfDNA next-generation sequencing identified initial EGFR activating and secondary T790M resistance mutations in NSCLC patients with high sensitivity, predicted treatment response equivalent to tissue analysis, and identified multiple novel and established resistance alterations.
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- 2018
40. Exosome-Based Detection of
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Elena, Castellanos-Rizaldos, Dominik G, Grimm, Vasisht, Tadigotla, James, Hurley, John, Healy, Patricia L, Neal, Mia, Sher, Raajdeep, Venkatesan, Chris, Karlovich, Mitch, Raponi, Anne, Krug, Mikkel, Noerholm, Jihane, Tannous, Bakhos A, Tannous, Luis E, Raez, and Johan K, Skog
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Lung Neoplasms ,Biopsy ,Reproducibility of Results ,Exons ,Exosomes ,Sensitivity and Specificity ,Circulating Tumor DNA ,ErbB Receptors ,ROC Curve ,Carcinoma, Non-Small-Cell Lung ,Mutation ,Biomarkers, Tumor ,Humans ,Alleles ,Neoplasm Staging - Published
- 2017
41. Heterogeneity Underlies the Emergence of EGFRT790 Wild-Type Clones Following Treatment of T790M-Positive Cancers with a Third-Generation EGFR Inhibitor
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Chris Karlovich, Joel W. Neal, Andrew R. Allen, A. John Iafrate, Anuj Kalsy, Elizabeth L. Lockerman, Heather A. Wakelee, Dora Dias-Santagata, Angel R. Garcia, Aaron N. Hata, Hillary E. Mulvey, Subba R. Digumarthy, Alona Muzikansky, Linnea Fulton, Lecia V. Sequist, Richard H. DiCecca, Mitch Raponi, Mari Mino-Kenudson, Jeffrey A. Engelman, Matthew J. Niederst, Melissa Parks, Zofia Piotrowska, and Alice T. Shaw
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Lung Neoplasms ,Drug resistance ,Biology ,Article ,Genetic Heterogeneity ,T790M ,Cell Line, Tumor ,Biopsy ,medicine ,Humans ,Neoplasm ,Prospective Studies ,Rociletinib ,Protein Kinase Inhibitors ,EGFR inhibitors ,Acrylamides ,medicine.diagnostic_test ,Genetic heterogeneity ,Gene Amplification ,DNA, Neoplasm ,medicine.disease ,Small Cell Lung Carcinoma ,respiratory tract diseases ,ErbB Receptors ,Gene Expression Regulation, Neoplastic ,Pyrimidines ,Oncology ,Drug Resistance, Neoplasm ,Mutation ,Immunology ,Cancer research ,EGFR Activating Mutation - Abstract
Rociletinib is a third-generation EGFR inhibitor active in lung cancers with T790M, the gatekeeper mutation underlying most first-generation EGFR drug resistance. We biopsied patients at rociletinib progression to explore resistance mechanisms. Among 12 patients with T790M-positive cancers at rociletinib initiation, six had T790–wild-type rociletinib-resistant biopsies. Two T790–wild-type cancers underwent small cell lung cancer transformation; three T790M-positive cancers acquired EGFR amplification. We documented T790–wild-type and T790M-positive clones coexisting within a single pre-rociletinib biopsy. The pretreatment fraction of T790M-positive cells affected response to rociletinib. Longitudinal circulating tumor DNA (ctDNA) analysis revealed an increase in plasma EGFR-activating mutation, and T790M heralded rociletinib resistance in some patients, whereas in others the activating mutation increased but T790M remained suppressed. Together, these findings demonstrate the role of tumor heterogeneity when therapies targeting a singular resistance mechanism are used. To further improve outcomes, combination regimens that also target T790–wild-type clones are required. Significance: This report documents that half of T790M-positive EGFR-mutant lung cancers treated with rociletinib are T790–wild-type upon progression, suggesting that T790–wild-type clones can emerge as the dominant source of resistance. We show that tumor heterogeneity has important clinical implications and that plasma ctDNA analyses can sometimes predict emerging resistance mechanisms. Cancer Discov; 5(7); 713–22. ©2015 AACR. See related commentary by Ichihara and Lovly, p. 694. This article is highlighted in the In This Issue feature, p. 681
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- 2015
42. Correction: Corrigendum: Circulating tumour DNA profiling reveals heterogeneity of EGFR inhibitor resistance mechanisms in lung cancer patients
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Thomas Harding, D. Ross Camidge, Jonathan W. Goldman, Chris Karlovich, Henry J. Haringsma, Gregory A. Otterson, Henning Stehr, Alexander F. Lovejoy, W. Thomas Purcell, Mohammad Shahrokh Esfahani, Kathleen A. Durkin, Joel W. Neal, Aaron M. Newman, David M. Kurtz, Ash A. Alizadeh, Maximilian Diehn, Florian Scherer, Andrew Simmons, Lecia V. Sequist, Heather A. Wakelee, Jacob J. Chabon, and Zofia Piotrowska
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0301 basic medicine ,Multidisciplinary ,business.industry ,Science ,General Physics and Astronomy ,General Chemistry ,EGFR Tyrosine Kinase Inhibitors ,medicine.disease ,medicine.disease_cause ,Corrigenda ,General Biochemistry, Genetics and Molecular Biology ,03 medical and health sciences ,030104 developmental biology ,DNA profiling ,Cancer research ,Medicine ,Non small cell ,KRAS ,business ,Lung cancer ,EGFR inhibitors - Abstract
Nature Communications 7: Article number: 11815 (2016); Published 10 June 2016; Updated 14 November 2016 Previous work by Del Re et al. describing the emergence of KRAS mutations following treatment of non-small cell lung cancer patients with EGFR tyrosine kinase inhibitors was inadvertently omitted from the reference list of this Article and should have been cited as follows.
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- 2016
43. Abstract 3385: Comparison of genomic biomarkers identified by the whole exome, RNASeq and whole genome sequencing pipelines developed for the PDMR
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Li Chen, Rajesh Patidar, Biswajit Das, Chris Karlovich, Tomas Vilimas, Corinne Camalier, Vivekananda Datta, Shahanawaz Jiwani, William Walsh, Palmer Fliss, Sean McDermott, Justine N. McCutcheon, Amanda Peach, Michelle Ahalt-Gottholm, Carrie Bonomi, Kelly Dougherty, John Carter, Sergio Y. Alcoser, Tiffanie Chase, Raymond Divelbiss1, Marion Gibson, Kelly Hedger, Candace Mallow, Chelsea McGlynn, Malorie Morris, Marianne Radzyminski, Howard Stotler, Jesse Stottlemyer, Debbie Trail, Yvonne Evrard, Melinda G. Hollingshead, Mickey Williams, and James H. Doroshow
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Cancer Research ,Oncology - Abstract
Background: The National Cancer Institute (NCI) has developed a Patient-Derived Models Repository (PDMR; www.pdmr.cancer.gov) of patient-derived xenografts (PDXs) with clinical annotation and comprehensive genomic characterization using whole exome sequencing (WES) and RNASeq. An in-house data analysis pipeline has been developed and validated to call germline and somatic variants and to perform transcriptional profiling in these models. There is a need to incorporate additional biomarkers into standard data analysis pipeline, including loss of heterozygosity (LOH), microsatellite instability (MSI), structure variants (SVs)/fusions and copy number variation (CNV) for identifying appropriate PDX models for preclinical drug studies. Validation of the methods used for the assessment of these and other genomic biomarkers is a crucial aspect in the development of the PDMR data analysis pipeline. Methods: WGS, WES and RNASeq were conducted on 58 PDX samples and genomic biomarkers were derived from different assays. For LOH calling, a set of ~800,000 heterozygous SNPs was first constructed from a population level genomic database (gnomAD) and a specific list of ~3000 highly heterozygous SNPs from a previous study. LOH regions were detected using Runs of Homozygosity (BCFtools/RoH) based on the genotypes of ~800,000 SNPs. Finally, percent of genomic LOH was calculated as the percent of eligible LOH regions in the whole genome. For MSI calling. mSINGS was used to assign a microsatellite instability score based on the fraction of unstable microsatellite loci. Gene fusions were detected using Tophat-fusion and Fusion-catcher from RNASeq data and Manta from WGS. CNVs were derived from WGS and WES using CNVkit. Results: Genomic biomarkers derived from WES and RNASeq were highly concordant with the ones derived from WGS. Specifically, we found 1) the percent of genomic LOH was highly correlated between WGS and WES across 52 samples with R2=0.99, where LOH% ranged from Conclusions: We observed excellent consistency between WGS, WES and RNASeq data in the assessment of percent of LOH, MSI score, SVs/fusions and CNVs. Our data analysis pipeline can accurately call genomic biomarkers from WES and RNASeq data, which facilitates the molecular characterization and prioritization of PDMR models for preclinical drug treatment. Citation Format: Li Chen, Rajesh Patidar, Biswajit Das, Chris Karlovich, Tomas Vilimas, Corinne Camalier, Vivekananda Datta, Shahanawaz Jiwani, William Walsh, Palmer Fliss, Sean McDermott, Justine N. McCutcheon, Amanda Peach, Michelle Ahalt-Gottholm, Carrie Bonomi, Kelly Dougherty, John Carter, Sergio Y. Alcoser, Tiffanie Chase, Raymond Divelbiss1, Marion Gibson, Kelly Hedger, Candace Mallow, Chelsea McGlynn, Malorie Morris, Marianne Radzyminski, Howard Stotler, Jesse Stottlemyer, Debbie Trail, Yvonne Evrard, Melinda G. Hollingshead, Mickey Williams, James H. Doroshow. Comparison of genomic biomarkers identified by the whole exome, RNASeq and whole genome sequencing pipelines developed for the PDMR [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 3385.
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- 2019
44. Abstract 1056: Xenograft-associated B cell lymphoproliferative disease (XABLD) as a surrogate model to study Epstein-Barr virus (EBV) driven B cell Diseases
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Amanda Peach, John Carter, Rajesh Patidar, Brandie A. Fullmer, Howard Stotler, Pallavi Galera, Michelle M. Gottholm-Ahalt, Corinne Camalier, Suzanne Borgel, P. Mickey Williams, Liqiang Xi, Vivekananda Datta, Raymond Divelbiss, Gloryvee Rivera, Biswajit Das, Jesse Stottlemyer, Tomas Vilimas, Wiem Lassoued, Melinda G. Hollingshead, Chris Karlovich, Mark Raffeld, Elaine S. Jaffe, Sean P. McDermott, James H. Doroshow, William Jacob, Li Chen, Yvonne A. Evrard, Lindsay Dutko, and Michelle A. Crespo-Eugeni
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Cancer Research ,business.industry ,Cell of origin ,T cell ,medicine.disease ,medicine.disease_cause ,Epstein–Barr virus ,medicine.anatomical_structure ,Immunophenotyping ,Circulating tumor cell ,Oncology ,Monoclonal ,medicine ,Cancer research ,business ,Diffuse large B-cell lymphoma ,B cell - Abstract
Background: Patient-derived tumor xenografts (PDX) are powerful tools to study cancer biology, cancer genomics and developmental therapeutics. A common problem in the development of PDX models is proliferation of atypical lymphocytes at the implantation site, which often overtake or limit the growth of the original tumor. This atypical lymphocyte proliferation has been described as XABLD in our PDX models. In this study, we characterized XABLD cases by morphology, immunophenotyping and genomic profiling. We hypothesize that XABLD tumors are morphologically and phenotypically similar to EBV-driven post-transplant lymphoproliferative disease (PTLD) and diffuse large B cell lymphoma (DLBCL). XABLD is a surrogate model to study EBV-driven PTLD and DLBCL. Materials and Methods: Models were generated from patient tissue collected under NCI Tissue Procurement Protocol (clinicaltrials.gov: NCT00900198) and CIRB Tissue Procurement Protocol 9846 for development of models for NCI’s Patient-Derived Models Repository (https://pdmr.cancer.gov). Specimens were implanted subcutaneously in NOD/SCID/IL2Rg null (NSG) mice and animal health was monitored throughout the study. Tumors in mice with suspected XABLD were harvested and reviewed by histology and immunohistochemical analysis for CD45, B and T cell markers, EBV status, B-cell clonality assay. All samples were also classified by the Lymph2Cx NanoString cell of origin assay and transcriptome profiling. Results: XABLD cases were found to originate from both solid tumor and circulating tumor cell implants. XABLD is a rapidly growing tumor positive for CD45, CD20, and LMP1 stains, 36 of 42 cases are strongly positive for PD-L1 stain. 39 of 42 cases exhibited an activated B cell (ABC) phenotype with evidence of elevated NF-kB signaling. Most cases were monoclonal for IGK/IGH and contained high numbers of tumor infiltrating CD8-positive T-cells with associated high mRNA expression of activated T cell markers. Conclusion: The clinical presentation, morphology and molecular characteristics of XABLD cases were similar to EBV-driven DLBCL. As the XABLD models exhibited frequent PD-L1 expression and marked infiltration of CD8-positive T cells, they may be useful for in vitro evaluation of checkpoint inhibitor response and T cell antitumor activity. Grant Support: This project has been funded in part with federal funds from the National Cancer Institute, National Institutes of Health, under Contract No. HHSN261200800001E. The content of this publication does not necessarily reflect the views or policies of the Department of Health and Human Services, nor does mention of trade names, commercial products, or organizations imply endorsement by the U.S. Government. Citation Format: Tomas Vilimas, Gloryvee Rivera, Brandie Fullmer, Wiem Lassoued, Lindsay Dutko, Amanda Peach, Corinne Camalier, Li Chen, Rajesh Patidar, Suzanne Borgel, John Carter, Howard Stotler, Raymond Divelbiss, Jesse Stottlemyer, Michelle M. Gottholm-Ahalt, Michelle Crespo-Eugeni, Sean McDermott, William Jacob, Liqiang Xi, Pallavi Galera, Yvonne A. Evrard, Melinda G. Hollingshead, Elaine S. Jaffe, Mark Raffeld, Biswajit Das, Chris Karlovich, Vivekananda Datta, James H. Doroshow, P. Mickey Williams. Xenograft-associated B cell lymphoproliferative disease (XABLD) as a surrogate model to study Epstein-Barr virus (EBV) driven B cell Diseases [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 1056.
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- 2019
45. Abstract 4524: Comparison of PDX, PDC, and PDOrg models from the National Cancer Institute’s Patient-Derived Models Repository (PDMR)
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Jenna Moyer, Howard Stotler, James H. Doroshow, Tom Walsh, Lily Chen, Abigail Walke, Mike Mullendore, Matt Murphy, Tara Grinnage-Pulley, Luke H. Stockwin, Marion Gibson, Yvonne A. Evrard, John Mark Carter, Wiem Lassoued, Suzanne Borgel, Carrie Bonomi, Kelly Dougherty, Kevin Plater, Biswajit Das, Raymond Divelbiss, Joe Geraghty, Vivekananda Datta, Nikki E. Craig, Emily Delaney, Marianne Radzyminski, Alice P. Chen, Kaitlyn Arthur, Mariah Baldwin, Rajesh Patidar, Dianne L. Newton, Sergio Y. Alcoser, Debbie Trail, Kyle Georgius, Shahanawaz Jiwani, Chris Karlovich, Tiffany Chase, Candace Mallow, Mallorie Morris, Sierra Hoffman, Thomas P. Forbes, P. Mickey Williams, Nicki Scott, Savanna Styers, Anna Wade, Jesse Stottlemyer, Chelsea McGlynn, Tomas Vilimas, Melinda G. Hollingshead, Michelle M. Gottholm-Ahalt, and Kelly Hedger
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Cancer Research ,Research use ,Extramural ,Genetic stability ,Cancer ,medicine.disease ,Rare cancer ,Genealogy ,Unmet needs ,Data sequences ,Oncology ,Multiple criteria ,medicine ,Psychology - Abstract
The National Cancer Institute (NCI) has developed a Patient-Derived Models Repository (PDMR) comprised of quality-controlled, early-passage, clinically-annotated patient-derived tumor xenografts (PDXs), in vitro tumor cell cultures (PDCs), cancer associated fibroblasts (CAFs), and patient-derived organoids (PDOrg). NCI has focused on generating models to complement existing PDX collections and address unmet needs in the preclinical model space. These models are offered to the extramural community for research use (https://pdmr.cancer.gov), along with clinical annotation and molecular information (whole exome sequence, gene expression using RNASeq), via a publicly accessible database. Currently, over 200 PDX models, 50 PDC models, and 100 CAF models are available for distribution to the US research community. Approximately 50 PDOrg models will be released in early 2019. As part of its rare cancer initiative, the NCI is also targeting the collection of infrequently-observed tumor histologies to advance both biological investigations and drug development efforts for under-studied malignancies. Comparison of matched models, models where more than one model type are available (e.g., PDX and PDC), demonstrate a high degree of concordance across the model types. Genetic stability across the models is assessed using multiple criteria including genetic assessment of CNVs and presence of driver mutations. Optimal CNV assessment uses whole exome sequence data corrected for cellularity in the patient specimen using germline reads and corrected for cellularity in the PDX specimens by subtraction of the mouse reads. Histomorphologic comparison of PDXs and cell line xenografts (CLX) generated from in vitro PDCs and PDOrgs also overall show a high degree of concordance, though loss of features and dedifferentiation can be observed in some models. Overall these models demonstrate a high degree of conservation at the genetic and pathologic level when compared to the patient tumor. These models can provide researchers the ability to perform high- or mid-throughput screening in 2D or 3D culture followed by targeted selection of PDX models for in vivo studies. Funded by NCI Contract No. HHSN261200800001E Citation Format: Yvonne A. Evrard, Dianne Newton, Biswajit Das, Sergio Y. Alcoser, Kaitlyn Arthur, Mariah Baldwin, Carrie Bonomi, Suzanne Borgel, John Carter, Tiffany Chase, Alice Chen, Lily Chen, Nikki E. Craig, Vivekananda Datta, Emily Delaney, Raymond Divelbiss, Kelly Dougherty, Thomas Forbes, Kyle Georgius, Joe Geraghty, Marion Gibson, Michelle M. Gottholm-Ahalt, Tara Grinnage-Pulley, Kelly Hedger, Sierra Hoffman, Chris Karlovich, Wiem Lassoued, Shahanawaz Jiwani, Candace Mallow, Chelsea McGlynn, Mallorie Morris, Jenna Moyer, Mike Mullendore, Matt Murphy, Rajesh Patidar, Kevin Plater, Marianne Radzyminski, Nicki Scott, Luke H. Stockwin, Howard Stotler, Jesse Stottlemyer, Savanna Styers, Debbie Trail, Tomas Vilimas, Anna Wade, Abigail Walke, Thomas Walsh, P. Mickey Williams, Melinda G. Hollingshead, James H. Doroshow. Comparison of PDX, PDC, and PDOrg models from the National Cancer Institute’s Patient-Derived Models Repository (PDMR) [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 4524.
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- 2019
46. Development and analytical validation of a 523-gene clinical assay for cell-free DNA
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Geraldine O'Sullivan Coyne, Chen Zhao, David J. Sims, Thomas Forbes, Biswajit Das, James H. Doroshow, Naoko Takebe, Chris Karlovich, Robin D. Harrington, Rajesh Patidar, Jennifer S. LoCoco, Sigrid Katz, Corinne E. Camalier, Amanda Peach, Alice P. Chen, Paul Williams, Tingting Jiang, Li Chen, Nikitha Nair, and Ting-Chia Chang
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Cancer Research ,Oncology ,Cell-free fetal DNA ,business.industry ,Gene panel ,medicine ,Cancer ,Computational biology ,medicine.disease ,business ,Gene ,Complement (complexity) - Abstract
3039 Background: Liquid biopsies are emerging as a powerful complement to tumor biopsies for the clinical management of cancer patients. A large gene panel with robust analytical performance that accurately assesses variants, tumor mutational burden (TMB), and microsatellite instability in plasma would be of high value for immunotherapy studies, monitoring minimal residual disease and early cancer detection. To this end, we have completed the initial validation for the cell-free DNA (cfDNA) assay, TruSight Oncology 500 (TSO500), which interrogates the full coding region of 523 genes plus selected intronic regions for fusion detection in 23 driver genes. Methods: Cell-free DNA was extracted from plasma collected from Streck or EDTA blood tubes and quantitated to achieve an assay input of ≥10 ng. Libraries were constructed using unique molecular identifiers (UMIs) and duplex barcodes for error correction, then enriched by target capture and sequenced on a NovaSeq 6000. Healthy donor (HD) specificity assessment used matched white blood cell results to filter germline and clonal hematopoiesis variants. Contrived specimens were used to evaluate sensitivity. Single nucleotide variants (SNVs) (n = 36), insertion/deletions (indels) (n = 19), copy number variants (CNVs) (n = 6), and fusions (n = 5) were tested in 2 multi-site replicates. Results: Sensitivity of detection at 0.5% variant allele fraction (VAF) was > 95% and > 97% for SNVs and indels, respectively. All expected CNVs were identified at the targeted threshold of ≥1.3X change and showed strong correlation with matched digital PCR results. All fusions were identified at ≥0.4% VAF. Specificity in HD was > 99.99%. In 22 temporally matched tumor and blood samples from late-stage patients, 58% of all reportable mutations in tumor were identified in cfDNA. Preliminary TMB analysis identified one TMB high case with POLE p.P286R observed in both tissue and cfDNA. Conclusions: In this initial validation study the TSO500 cfDNA assay exhibited high sensitivity and specificity consistent with requirements for clinical applications. Ongoing studies will further evaluate TSO500 as a complement or potential alternative to tissue biopsy for the genomic profiling of cancer patients.
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- 2019
47. Measuring phospho-MET by multiplex immunofluorescence to aid in selection of patients with MET activation in tumors
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Apurva K. Srivastava, John Carter, Raymond Divelbiss, Tony Navas, Biswajit Das, Paul Williams, Chris Karlovich, Rajesh Patidar, Yvonne A. Evrard, Jeevan P Govindharajulu, Susanne Borgel, Ralph E. Parchment, Donald P. Bottaro, Jesse Stottlemyer, James H. Doroshow, Marianne Radzyminski, Li Chen, and Melinda G. Hollingshead
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Cancer Research ,MET Gene Amplification ,medicine.diagnostic_test ,business.industry ,Immunofluorescence ,Protein expression ,03 medical and health sciences ,0302 clinical medicine ,Oncology ,030220 oncology & carcinogenesis ,Cancer research ,medicine ,Multiplex ,business ,Selection (genetic algorithm) ,030215 immunology - Abstract
3131 Background: Currently, patient selection criteria for clinical testing of MET inhibitors are limited. Robust studies selecting patients based on MET protein expression, MET gene amplification, or mutations have not met their efficacy goals. Development of microscopy-based assays to quantify levels of phospho-MET (pMET) in tumors has been hampered by poor antibody specificity. Here, we present the development and validation of a robust, highly specific multiplex immunofluorescence assay (IFA) that measures pY1235-MET and total MET in tumor tissue. Methods: This assay utilizes antibodies to pY1235-MET (NCI-23111), total MET (D1C2), and plasma membrane (PM) marker Na+/K+-ATPase, each conjugated to a different Alexa Fluor dye. We used tumor tissue from crizotinib-treated SNU5 xenograft models to demonstrate pY1235-MET assay fitness-for-purpose and cross-platform assay concordance with our validated pMET ELISA. In addition, this IFA was validated by phospho-peptide competition using custom tissue microarrays (TMA) derived from patients with colorectal carcinoma (CRC). Finally, we developed quantitative algorithms to assess pY1235 MET levels in the plasma membrane and nucleus using PM and DAPI masks, respectively. Patient-derived xenograft models (PDX) were obtained from NCI’s Patient-Derived Models Repository (www.pdmr.cancer.gov). Results: The prevalence of high pY1235-MET expression in CRC patient specimens was greater than expected; of the 64 TMA cores evaluated, 29 (45%) and 19 (29%) had high pY1235-MET and total MET levels, respectively, as defined by mean marker area of ≥ 30 μm2/cell. To address the potential utility of pY1235-MET as a diagnostic biomarker, we examined 15 CRC PDX models by pMET ELISA and IFA. Two CRC tumor models were positive for pY1235-MET expression in both assays. The pY1235-MET IFA results and gene expression data were used to select PDX models for ongoing preclinical trials of potent MET inhibitors. Conclusions: This novel pY1235-MET IFA will enable clinicians to address the utility of activated MET as a biomarker for patient selection and/or prediction of response in clinical trials of MET inhibitors. Funded by NCI Contract No. HHSN261200800001E.
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- 2019
48. Design and development of the molecular analysis for Therapy Choice (NCI-MATCH) Designated Laboratory Network
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Cynthia Winter, Paul M. Williams, Peter J. O'Dwyer, Linda Zane, Gregory J. Tsongalis, A. John Iafrate, Keith T. Flaherty, Chris Karlovich, Jennifer Lee, Ting-Chia Chang, Stanley R. Hamilton, Alice P. Chen, Laura M. Yee, Barbara A. Conley, Kneshay N. Harper, Jeffrey Sklar, Lyndsay Harris, Robin D. Harrington, James V. Tricoli, and David Patton
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Oncology ,Cancer Research ,medicine.medical_specialty ,business.industry ,Internal medicine ,Mutation (genetic algorithm) ,medicine ,Histology ,business ,Precision medicine ,Refractory cancer ,Molecular analysis - Abstract
3016 Background: NCI-MATCH is a precision medicine trial that assigns treatment to refractory cancer patients by tumor mutation profile rather than by histology. After screening fresh tumor biopsies from nearly 6000 patients many treatment arms did not meet accrual due to the low prevalence of the eligible variants. NCI MATCH developed an approach to identify patients for the remaining arms utilizing a network of academic and commercial CLIA-certified labs that perform NGS assays as routine care at MATCH participating sites. Methods: Candidate labs were recruited through a notice in the Federal Register and posted on the NCI and ECOG ACRIN web sites. Twenty-seven labs (17 academic/10 commercial) submitted applications. After acceptance each lab analyzed a common set of 10 DNAs extracted from 8 cell lines and 2 clinical samples for concordance with the central NCI-MATCH NGS assay. Results: For the 17 labs with concordance results, a median of 8 (range 2 – 58) copy number variants (CNVs) were evaluated by the NGS assay of each DL, with the number evaluated depending on each lab’s clinical assay panel content. CNV concordance between central and DL assays, as measured by positive percent agreement (PPA), averaged 98.7% (range 87.5% - 100%) with the central assay as referent and 94.1% (range 77.8% – 100%) with the DL assay as referent. For single nucleotide variants (SNVs) and Insertion/deletions (Indels) combined, a median of 19 variants (range 11 – 26) were evaluated by each DL for concordance. PPA between central and DL assays averaged 98.0% (range 87.5% – 100%) and 98.6% (range 90.0% – 100%) with central and DL assay as referents, respectively. Strong correlations were observed between central and DL assays for both CNVs (median r = 0.93; 0.33 – 1.00) and SNV/Indels (median r = 0.98; 0.67 – 0.99). Conclusions: Our results suggest that different NGS assay platforms using diverse strategies for target enrichment and data analysis may still achieve high concordance if pre-analytical variables are minimized and the common genomic regions interrogated by each assay are well-understood. The designated lab network allows for a wider search for rare variants in tumors and provides a model for conducting future clinical trials. Clinical trial information: NCT02465060.
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- 2019
49. Durable Responses with Pembrolizumab in Relapsed/Refractory Mycosis Fungoides and Sézary Syndrome: Final Results from a Phase 2 Multicenter Study
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Holden T. Maecker, Martin A. Cheever, Michael S. Khodadoust, Alison J. Moskowitz, Sophia Fong, Yi Yang, Priyanka B. Subrahmanyam, Jinah Kim, Steven P. Fling, Lubomir Sokol, Biswajit Das, Chris Karlovich, Holbrook E Kohrt, Satish Shanbhag, Francine M. Foss, Yelena V. Budovskaya, Alain H. Rook, Vivekananda Datta, Shufeng Li, Steven M. Horwitz, Pierluigi Porcu, Asa Davis, Rajesh Patidar, Youn H. Kim, Jennifer H. Yearley, Elad Sharon, and Andrei R. Shustov
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0301 basic medicine ,medicine.medical_specialty ,business.operation ,Immunology ,Population ,Pembrolizumab ,Biochemistry ,03 medical and health sciences ,0302 clinical medicine ,Internal medicine ,medicine ,Clinical endpoint ,education ,health care economics and organizations ,education.field_of_study ,Horizon Pharma ,business.industry ,Surrogate endpoint ,Mallinckrodt ,Cell Biology ,Hematology ,Discontinuation ,030104 developmental biology ,Tolerability ,030220 oncology & carcinogenesis ,business - Abstract
Background: Current treatment of advanced stage Mycosis fungoides (MF) and Sézary syndrome (SS) remains unsatisfactory. Complete responses (CR) are typically Methods: This single-arm, multicenter study by the Cancer Immunotherapy Trials Network (CITN) enrolled 24 patients with MF/SS stages IB-IV, with at least one prior systemic therapy. Pembrolizumab was administered at 2 mg/kg every 3 weeks for up to two years. The primary endpoint was overall response rate (ORR) using global response criteria according to the ISCL/EORTC consensus guidelines. Skin responses were measured by mSWAT. Secondary endpoints were safety/tolerability, time to response (TTR), duration of response (DOR) and progression-free survival (PFS). Correlative studies included immunohistochemistry (IHC), mass cytometry, whole exome sequencing, gene expression profiling, and serum cytokine analysis. Results: Patients had advanced stage disease (23/24 with stage IIB-IV MF/SS), and were heavily pretreated (median of 4 prior systemic therapies). The ORR was 38% with 2 CR and 7 PR. Of the 9 responding patients, 6 had ≥90% improvement in skin disease by mSWAT. The median TTR was 11 weeks. Responses were durable, with 8 of 9 responses ongoing at last follow up (median DOR 64 weeks, range 32-153 weeks). One responding patient progressed 2 months after discontinuing treatment due to an adverse event (AE). The median PFS was not reached. Overall, the toxicity profile was similar to prior studies of pembrolizumab. Four patients discontinued treatment due to treatment related serious AEs of duodenitis, pneumonitis, hepatitis, and corneal ulcer. Skin flare reactions were observed early in the treatment course in 40% of patients with SS, but none with MF. The skin flare reactions did not result in any treatment discontinuation, and did not correlate with subsequent response to treatment. There was no significant association between response and clinical characteristics including stage, disease type (MF vs. SS), or number of prior therapies. IHC assessment of PD-1, PD-L1, and PD-L2 did not predict response. Treatment resulted in an increase of PD-L1 expression by both IHC and nanoString analysis. A nanoString18 gene signature of tumor inflammation that is predictive of response to pembrolizumab in other tumor types was not predictive in this cohort. High dimensional mass cytometry enabled precise identification and phenotyping of malignant T cells. There was a positive correlation between PD-1 expression on the malignant T cells and development of the skin flare reaction. Whole exome sequencing revealed genomic disruptions of PD-1 signaling including copy loss of PD-1. No associations were found between outcomes and genomic events involving the PD1/PD-L1 axis, total mutation number, or neoantigen numbers. Conclusions: The 38% ORR rate in this heavily pretreated population was 38% with a highly encouraging DOR of 64 weeks. No predictive biomarkers have emerged thus far, but additional studies are ongoing. Additional combination studies with pembrolizumab are warranted to improve response rates. Figure. Figure. Disclosures Khodadoust: Innate Pharma: Research Funding. Porcu:Innate Pharma: Consultancy. Foss:Miragen: Consultancy, Speakers Bureau; Seattle genetics: Consultancy; Spectrum: Consultancy; Mallinkrodt: Consultancy. Moskowitz:Bristol Myers-Squibb: Consultancy, Research Funding; Seattle Genetics: Consultancy, Honoraria, Research Funding; Incyte: Research Funding; Merck: Research Funding; Takeda: Honoraria; ADC Therapeutics: Research Funding. Shustov:Seattle Genetics: Research Funding. Sokol:Seattle Genetics: Consultancy; Mallinckrodt Pharmaceuticals: Consultancy; Spectrum Pharmaceuticals: Consultancy. Yearley:Merck: Employment. Horwitz:Mundipharma: Consultancy; Celgene: Consultancy, Research Funding; Seattle Genetics: Consultancy, Research Funding; ADC Therapeutics: Consultancy, Research Funding; Kyowa-Hakka-Kirin: Consultancy, Research Funding; Forty Seven: Consultancy, Research Funding; Millennium/Takeda: Consultancy, Research Funding; Infinity/Verastem: Consultancy, Research Funding; Aileron Therapeutics: Consultancy, Research Funding; Portola: Consultancy; Trillium: Consultancy; Innate Pharma: Consultancy; Spectrum: Research Funding; Corvus: Consultancy. Kim:miRagen: Research Funding; Horizon Pharma: Consultancy, Research Funding; Neumedicine: Consultancy, Research Funding; Takeda: Membership on an entity's Board of Directors or advisory committees, Research Funding; Galderma: Research Funding; Soligenix: Research Funding; Eisai: Membership on an entity's Board of Directors or advisory committees, Research Funding; Tetralogic: Research Funding; Seattle Genetics: Membership on an entity's Board of Directors or advisory committees; Portola: Research Funding; Merck: Research Funding; Medivir: Membership on an entity's Board of Directors or advisory committees; Kyowa-Kirin-Pharma: Membership on an entity's Board of Directors or advisory committees, Research Funding; Innate Pharma: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Forty Seven Inc: Research Funding.
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- 2018
50. Assessment of EGFR Mutation Status in Matched Plasma and Tumor Tissue of NSCLC Patients from a Phase I Study of Rociletinib (CO-1686)
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Lin Wu, Andrew R. Allen, D. Ross Camidge, Lindsey Rolfe, Wei Wen, Heather A. Wakelee, Patrick O'Donnell, Jong-Mu Sun, Philipp Angenendt, Darrin Despain, Leora Horn, Keunchil Park, Shirish M. Gadgeel, Lecia V. Sequist, Jonathan H. Goldman, Jean-Charles Soria, Elaina Mann, Cloud P. Paweletz, Shannon Matheny, Krzysztof Konopa, Chris Karlovich, David R. Spigel, Benjamin Solomon, Sean Chien, and Mitch Raponi
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0301 basic medicine ,Oncology ,Adult ,Male ,Cancer Research ,medicine.medical_specialty ,Pathology ,Lung Neoplasms ,Drug resistance ,Article ,03 medical and health sciences ,T790M ,0302 clinical medicine ,Internal medicine ,Carcinoma, Non-Small-Cell Lung ,Biopsy ,Carcinoma ,Medicine ,Humans ,Rociletinib ,Protein Kinase Inhibitors ,EGFR inhibitors ,Aged ,Aged, 80 and over ,Acrylamides ,medicine.diagnostic_test ,business.industry ,Cancer ,Middle Aged ,Resistance mutation ,medicine.disease ,respiratory tract diseases ,ErbB Receptors ,030104 developmental biology ,Pyrimidines ,Drug Resistance, Neoplasm ,030220 oncology & carcinogenesis ,Mutation ,Female ,business - Abstract
Purpose: The evaluation of plasma testing for the EGFR resistance mutation T790M in NSCLC patients has not been broadly explored. We investigated the detection of EGFR activating and T790M mutations in matched tumor tissue and plasma, mostly from patients with acquired resistance to first-generation EGFR inhibitors. Experimental Design: Samples were obtained from two studies, an observational study and a phase I trial of rociletinib, a mutant-selective inhibitor of EGFR that targets both activating mutations and T790M. Plasma testing was performed with the cobas EGFR plasma test and BEAMing. Results: The positive percent agreement (PPA) between cobas plasma and tumor results was 73% (55/75) for activating mutations and 64% (21/33) for T790M. The PPA between BEAMing plasma and tumor results was 82% (49/60) for activating mutations and 73% (33/45) for T790M. Presence of extrathoracic (M1b) versus intrathoracic (M1a/M0) disease was found to be strongly associated with ability to identify EGFR mutations in plasma (P < 0.001). Rociletinib objective response rates (ORR) were 52% [95% confidence interval (CI), 31 – 74%] for cobas tumor T790M-positive and 44% (95% CI, 25 – 63%) for BEAMing plasma T790M-positive patients. A drop in plasma-mutant EGFR levels to ≤10 molecules/mL was seen by day 21 of treatment in 7 of 8 patients with documented partial response. Conclusions: These findings suggest the cobas and BEAMing plasma tests can be useful tools for noninvasive assessment and monitoring of the T790M resistance mutation in NSCLC, and could complement tumor testing by identifying T790M mutations missed because of tumor heterogeneity or biopsy inadequacy. Clin Cancer Res; 22(10); 2386–95. ©2016 AACR.
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- 2016
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