Circulating cell-free DNA (cfDNA) currently allows for minimal residual disease detection and treatment monitoring with new clinical data emerging regularly. However, as technology evolves and assays increase in sensitivity, the possibility of earlier detection will become more of a reality. At Cambridge Healthtech Institute’s Third Annual Circulating Cell-Free DNA, leading researchers from academia and industry will come together to address advances as well as existing challenges in this rapidly growing field. This year’s event will place emphasis on early detection, clinical validation of assay sensitivity and specificity, and establishing standards for cfDNA.
Final Agenda
Thursday, March 10
7:30 am Registration and Morning Coffee
8:25 Chairperson’s Opening Remarks
Cloud P. Paweletz, Ph.D., Head, Translational Research Laboratory; Biomarker Lead, Belfer Institute for Applied Cancer Science, Dana Farber Cancer Institute
» 8:30 KEYNOTE PRESENTATION: Deep Sequencing of Circulating Tumor DNA for Personalized Cancer Detection and Monitoring
Maximilian Diehn, M.D., Ph.D., Assistant Professor, Radiation Oncology, Stanford University
I will describe the development and application of CAPP-Seq, a deep sequencingbased method for ultra-sensitive and specific detection of circulating tumor DNA that is broadly applicable to different cancer types and clinical scenarios.
9:10 Clinical Applications of an NGS Assay for Ultrasensitive Measurement of ctDNA
Abhijit Patel, M.D., Ph.D., Assistant Professor, Yale University School of Medicine
Our group has developed an ultrasensitive, multi-target NGS-based assay that can identify and quantify mutant ctDNA using novel error-suppression techniques. Broad coverage of mutation hotspots and warm-spots allows detection of ctDNA without prior knowledge of the tumor’s mutation profile. Data will be presented from various ongoing studies to establish the clinical utility of this technology.
9:40 Assay Characterization for Clinical Applications
Rebecca Leary, Ph.D., Lab Head & Research Investigator, Genomics Group, Next Generation Diagnostics, Novartis
Evaluation and characterization of cell-free DNA technologies is a critical step when introducing these assays for the analysis of clinical specimens.
10:10 Multiplexed ICE COLD PCR Enriches Any Low-Level Mutation Present in DNA Isolated from FFPE and Plasma Samples
Harjit Kullar, Ph.D., MBA, Vice President, Marketing, Transgenomic
Using Multiplexed ICE COLD-PCR, cancer patients’ DNA isolated from FFPE & Plasma was utilized to enrich for any low level mutations present in the samples. The results demonstrated detection of low level mutations in liquid biopsies; thus allowing for disease management with respect to treatment options and/or drug cocktail modifications for cancer patients.
10:40 Coffee Break with Exhibit and Poster Viewing
11:15 Clinical Evidence for the Utility of the cobas® EGFR Mutation Test v2 with Liquid Biopsy Samples from NSCLC Patients
John Palma, Ph.D., Director, Medical Affairs, Roche
Common challenges to ascertaining EGFR mutation status in non-small cell lung cancer are the lack of or inadequacy of biopsy material. The cobas® EGFR Mutation Test v2 was developed to detect the most common mutations in exons 18-21, including T790M, for both tissue and plasma. Using different cutoffs for sample type, the detection of 42 mutations is now possible. While maintaining a high positive predictive value to detect EGFR mutations relative to tissue results, liquid biopsy testing could eventually enable the detection of acquired mutations during treatment.
11:45 Monitoring Cancer through the Blood
Cloud P. Paweletz, Ph.D., Head, Translational Research Laboratory; Biomarker Lead, Belfer Institute for Applied Cancer Science, Dana Farber Cancer Institute
Genomic alterations in genes such as EGFR, ALK, KRAS, and BRAF have been validated as powerful predictive biomarkers in the management of non-small cell lung cancer (NSCLC), colorectal cancer, and melanoma; testing for these mutations is currently standard to personalize treatment decisions. The challenges associated with routine use of NGS include availability of adequate tumor specimens, slow turnaround time, and evolving tumor biology in response to treatment that may necessitate a repeat biopsy to guide subsequent therapy. Here we discuss the use of blood based, non-invasive test to diagnose, monitor and understand lung cancer.
12:15 pm Pharmacodynamic Assessment of Drug Response by Monitoring Mutational Load in Urinary Circulating Tumor DNA
Vlada Melnikova, M.D., Ph.D, Vice President, Research & Development, Trovagene
The concept of liquid biopsies is expanding to include urine as a specimen type. Using DNA extraction process that isolates systemic ctDNA and a quantitative PCR-NGS enrichment method for mutation detection at a single copy level, we demonstrate that drug-induced immediate early changes in ctDNA mutational load correlate with tumor burden and treatment response. As a non-invasive specimen, urine enables development of novel algorithms to inform treatment decisions via frequent monitoring of ctDNA.
12:45 Luncheon Presentation: Precision-Based Circulating Tumor DNA Detection and Monitoring in Gynecologic Cancer Samples
John Martignetti, M.D., Ph.D., Icahn School of Medicine at Mount Sinai, Departments of Genetics and Genomic Sciences, Pediatrics, Obstetrics/Gynecology & Reproductive Sciences and Oncological Sciences
Dr. Martignetti and his team are devloping a rapid and efficient approach for variant discovery in gynecologic cancer samples that couples tumor-specific mutation identification and RainDrop® digital PCR-based ctDNA detection. Tumor mutation profiles have been generated for detection and monitoring of tumor status in ovarian and endometrial cancer samples. All results were compared against current FDA-approved biomarkers and the known clinical status of the patients from which the samples were taken, demonstrating the highly sensitive, specific and robust nature of this approach.
1:15 Session Break
1:50 Chairperson’s Remarks
Nishant Agrawal, M.D., Professor, Surgery; Director, Head and Neck Surgical Oncology, University of Chicago
2:00 Application of Tumor-Derived DNA in the Management of Individuals
with Cancer
Chetan Bettegowda, M.D., Ph.D., Assistant Professor, Neurological Surgery, Johns Hopkins University School of Medicine
The development of non-invasive methods to detect and monitor tumors continues to be a major challenge in oncology. The molecules of tumor derived DNA can be distinguished from the background of normal DNA by the presence of somatic mutations. Using sensitive digital PCR based approaches, we have been able to query bio-fluids from a number of human malignancies for levels of tumor derived DNA (ct-DNA). We will present our efforts at detecting ctDNA in a number of different solid tumors and the potential clinical applications of this approach.
2:30 Deep Learning of Circulating Tumor DNA Genome-Wide Sequencing Data Yields Novel Early Detection Signatures
Gabriel Otte, Ph.D., President & CEO, Freenome Inc.
Adam Drake, CSO, Computational Research, Freenome Inc.
Current methods of cancer detection perform little better than a coin flip due to inadequate sensitivity and specificity. We have sequenced the cell-free DNA (cfDNA) from blood plasma to a shallow depth and designed classification procedures based on deep learning of genomic data that detect cancer with a 96%+ sensitivity and specificity.
3:00 Refreshment Break with Exhibit and Poster Viewing
3:30 Maximizing Next-Generation Sequencing Capabilities
of Circulating, Cell-Free DNA
Timothy Harkins, President & CEO, Swift Biosciences
Swift Biosciences presents NGS methods that are cost effective, sensitive, and specific to assess cfDNA. Methods discussed for whole genome sequencing from PCR-free libraries, point mutation detection with hyb/capture and multiplex amplicons, and methylation patterns all from single liquid biopsy samples.
4:00 PANEL DISCUSSION: Clinical Implementation of cfDNA
Moderator: Daniel Grosu, M.D., MBA, CMO, Sequenom, Inc
Panelists: Sabita Sankar, Ph.D., Director, Business Development, Biodesix
Abhijit Patel, M.D., Ph.D., Assistant Professor, Yale University School of Medicine
Cloud P. Paweletz, Ph.D., Head, Translational Research Laboratory; Biomarker Lead, Belfer Institute for Applied Cancer Science, Dana Farber Cancer Institute
- How to implement in a way that benefits patients
- What information are you getting and how will people use it?
- Who pays? Are we ready for plasma-based NGS tests?
- Lessons learned
5:00 Reception with Exhibit and Poster Viewing
6:00 Close of Day
Friday, March 11
7:00 am Breakfast Presentation: Tracking of Circulating Cell-Free DNA from Plasma for Treatment Response Monitoring
Raj Krishnan, Ph.D., CEO, Biological Dynamics, Inc.
Interest in the isolation, quantification, and analysis of cell-free biomarkers directly from blood has grown significantly. A proprietary platform has been developed for isolating and quantifying circulating biomarkers from physiological solutions using AC Electrokinetics (ACE). Biomarkers, such as high molecular weight cell-free DNA, have been established as indicators of tumor burden, and our TR(ACE) assay is currently being developed as a rapid, inexpensive method to follow treatment response in cancer patients.
7:55 Chairperson’s Remarks
Grace Zhao, Ph.D., Co-Founder and Director, Research, AccuraGen
8:00 An Ultra-Accurate System for Cancer Mutation Detection in Circulating Cell-Free DNA from Plasma
Grace Zhao, Ph.D., Co-Founder and Director, Research, AccuraGen
Rare mutation detection using circulating cfDNA presents two major challenges: low input, and relatively high error rate of current NGS technology. At AccuraGen, we have developed an innovative assay platform to amplify cfDNA, and correct sequencing error to enhance mutation detection at a high accuracy. AccuraGen provides flexible liquid biopsy assays that cover from hundreds to handful of genes with a reliable detection rate of 0.05% in 5000 copies of genome.
8:30 High Sensitivity NGS Analysis of ctDNA: Applications to Non-Small Cell Lung Cancer and Beyond
Tim Forshew, Ph.D., Head, Technology Development, Inivata, Ltd.
It is now established that a broad spectrum of cancers release circulating tumor DNA (ctDNA) into the blood. Detection and quantification of this DNA has numerous potential clinical applications but raises challenges that need to be overcome. We published the first NGS method to detect mutations de novo through ctDNA sequencing (TAm-Seq) and will present the development of our optimized, highly sensitive multi-gene panel method, with supporting clinical data.
9:00 Multipanel Massive Parallel Sequencing cfDNA in Monitoring Cutaneous Melanoma Progression
Dave S.B. Hoon, Ph.D., Director, Molecular Oncology and Sequencing Center, John Wayne Cancer Institute, Providence Health Care
Analysis of cfDNA gene mutation panel in melanoma patients serial bleeds over several years of follow-up can be very informative on events ongoing during tumor progression. The highly sensitive and approach of multiple gene mutation panel assessment by MPS of cfDNA provides a very comprehensive analysis to allow correlation to real-time clinical events of patients. These retrospective studies demonstrate that cfDNA mutations can arise at different time points during tumor progression. This approach of precision medicine monitoring melanoma progression allows a more accurate real-time assessment of what is transpiring in the patient.
9:30 Measuring Donor-Derived Cell-Free DNA in Organ Transplant Recipients as a Dynamic Biomarker of Rejection
John J. Sninsky, Ph.D. CSO, CareDx
A clinical-grade NGS assay was developed to monitor donor-derived cell-free DNA (dd-cfDNA) in plasma from solid organ transplant recipients. Longitudinal samples from heart and kidney transplant patients show elevated dd-cfDNA levels prior to and at the time of acute rejection which are reduced following successful immunosuppressive therapy.
10:00 Blood Testing for Actionable Variants in NSCLC Patients by ddPCR: A CLIA Lab Experience
Hestia Mellert, Ph.D., Senior Scientist, Biodesix Inc.
Clinical validation of cfDNA and cfRNA assays can be challenging due to the low prevalence of some variants. The GeneStrat™ panel is a Lab Developed Test using Droplet Digital PCR performed in a CLIA Laboratory for the detection of EGFR sensitizing and resistance, KRAS and BRAF mutations, and EML4-ALK fusion transcripts using nucleic acids isolated from plasma.
10:30 Coffee Break with Exhibit and Poster Viewing
11:00 Considerations for Establishing Standards for Circulating Cell Free DNA, RNA and Exosome Biomarkers in Cancer Diagnostics
Michael J. Heller, Ph.D., Professor, Nanoengineering & Bioengineering, University of California San Diego
Many considerations exist for establishing standards for circulating cell free DNA, RNA and exosome biomarkers in cancer diagnostics. First, these biomarkers will be used for diagnostic assays and tests that range from early cancer detection to patient management and therapy monitoring. Second, clinical decisions will have to be made as to which and how many DNA/RNA mutations and exosomes surface protein biomarkers are most relevant and predictive at different stages of cancer. Third, it is very likely that different diagnostic criteria will used for the many different types of cancer. Fourth, how will solid tumor biopsy information be used with new liquid biopsy information? Finally, standards will not only be dependent upon detection methods (PCR, sequencing, etc.), but also on sample preparation procedures which will ultimately determine viability of the assays in terms of time and cost.
11:30 Plasma Genotyping as a Novel Biomarker in Non-Small Cell Lung Cancer & Tool for Drug Development
Adrian G. Sacher, M.D., Clinical Fellow, Medicine, Dana-Farber Cancer Institute
Plasma genotyping of cell-free DNA (cfDNA) is quickly evolving as a method to select personalized therapy as well as monitor response to therapy in lung cancer. However, prospective data on the optimal platform and utility of plasma genotyping assays is limited. This talk will review the current state-of-the-art and present results from our ongoing prospective studies of plasma genotyping in lung cancer at the Dana-Farber Cancer Institute.
12:00 pm Quality Assessment of Cell-Free DNA to Guide Downstream Molecular Analyses
Muhammed Murtaza, Research Assistant Professor, Co-Leader, Center for Non-Invasive Diagnostics, Translational Genomics Research Institute
Recent proof-of-principle studies have demonstrated potential utility of sequencing cell-free DNA in cancer diagnostics. However, little is understood about the effect of fragment size distributions in plasma DNA on the performance of sequencing-based assays. We developed a multiplexed assay to perform one-step analysis of DNA quantity and integrity from minute amounts of cell-free DNA using picoliter droplet digital PCR. Our results were predictive of diversity and obtainable depth-of-coverage in next-generation sequencing libraries made from cell-free DNA samples.
12:30 Close of Symposium