Wednesday, December 5th

9:30 AM

Registration and Continental Breakfast

10:30 AM

Welcome and Logistics

John Mudgett, Co-Founder and Chief Scientific Officer

JsM BioScience, LLC

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Welcome and Logistics

10:45 AM

Keynote Presentation
Transitioning Discoveries in ALS Genetics to Therapeutic Treatments

John Landers, Professor of Neurology

University of Massachusetts Medical School

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Transitioning Discoveries in ALS Genetics to Therapeutic Treatments

TBA

11:25 AM

Application of sequencing data in pre-clinical drug development in oncology

Julia Schüler, Research Director

Charles River Discovery Research Services

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Application of sequencing data in pre-clinical drug development in oncology

In the current study, we present a potential workflow for the investigation of novel anti-cancer molecules using the example of EGFR targeting small molecules as well as antibodies evaluated on PDX – in vitro and in vivo – pre-clinical platforms. In a first step, suitable models were selected from the Charles River Compendium by means of EGFR expression. The selection included several high expressing models as well as low expressers for proof of principle.  Subsequently, EGFR expression was confirmed on protein level by immunohistochemistry using a tissue micro array and quantified by the OSANO software for the selected PDX models. A screen on 25 EGFR-targeting small molecules in 74 PDX derived cell lines in 2D revealed 27 highly sensitive and 25 resistant models. Among the sensitive models, gastric cancer and melanoma were statistically over-represented. A subsequent ex vivo 3D screen on 296 PDX models using the 10 most active EGFR-targeting compounds confirmed and extended the data-set of the 2D screen. Again, gastric cancer but as well non-small cell lung and renal cancer were highly sensitive towards treatment. The predictivity of the in vitro platforms was proven by in vivo characterization of Cetuximab, Gefitinib and Erlotinib in 23 PDX models. The predictivity of the 3D clonogenic assay for the PDX in vivo assay was precise for Gefitinib and Erlotinib. For Cetuximab the 3D assay was under-predictive as the antibody was much more active in vivo as in vitro.  Furthermore, we could identify a predictive biomarker for model sensitivity towards an EGFR-targeting drug treatment using the data-set comprising molecular, in vitro and in vivo data. Combining those datasets with clinical data will give us the possibility to validate the biomarkers towards their translational value. Taken together, the compendium-based model selection helps to test potential drug treatments in suitable in vitro and in vivo cancer models and thereby proofs to be a reliable tool in the drug development pipeline, including sensitivity, predictive biomarker, and validation analysis.

Authors
Hagen Klett, Anne-Lise Peille, Armin Maier, Gerhard Kelter, Kerstin Klingner, Daniel Bug, Julia Schüler

11:55 AM

Lunch

TRACK 1: Sequencing as a Tool to Discover and Quantify Gene Signatures for potential Diagnostic and Therapeutic Indication

1:00 PM

Keynote Presentation
High Throughput, Reduction Transcriptomics, using Ultraplex RNA Sequencing

Samuel Rulli, R&D Scientist

QIAGEN

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High Throughput, Reduction Transcriptomics, using Ultraplex RNA Sequencing

Elucidating molecular mechanisms and understanding genetic heterogeneity in drug discovery and complex biology can be achieved through high throughout gene expression analysis and pathway assignment. Recent advances in RNAseq methodologies have enabled accurate gene expression profiling from low input samples, but analyzing hundreds or thousands of samples is an arduous exercise in library prep.

Here we describe a high throughput 3’ RNAseq library prep methodology termed Ultraplex. With QIAseq Ultraplex, Reverse transcription is performed directly on lysed cells, while simultaneously assigning a unique ‘cell index’ to all cDNA synthesized from a sample. All subsequent transcriptome or targeted panel library steps that follow occur in a single pooled sample; with up to 384 samples assigned per sequencing index. Each 384-plex library is assigned a standard sample index, such that up to 384 pooled 384X libraries can be sequenced together.

With this methodology, thousands of transcriptomes or targeted RNA panels can be prepared and sequenced together. With QIAseq UPX, high throughput transcriptomic analysis enables the identification and characterization of gene signatures which divide samples into discrete sub-populations based on vectoral gene expression component

1:40 PM

Immuno-Oncology Biomarkers: Incorporating Tumor Mutational Burden and Micro-Satellite Instability in routine tumor profiling via next-generation sequencing

Maude Champagne, Commercial Development, Oncology

Illumina

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Immuno-Oncology Biomarkers: Incorporating Tumor Mutational Burden and Micro-Satellite Instability in routine tumor profiling via next-generation sequencing

TBA

2:10 PM

SeqPlus Unlocking the true value in FFPE Samples, Enabling More Efficient Genome & Exome Sequencing.

Jim Lund, Director Tumor Product Development

WuXi NEXTCODE

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SeqPlus Unlocking the true value in FFPE Samples, Enabling More Efficient Genome & Exome Sequencing.

Pathology archives with linked clinical data are an invaluable resource for translational research, with the limitation that most cancer samples are formalin-fixed paraffin-embedded (FFPE) tissues.  Therefore, FFPE tissues are an important resource for genomic profiling studies but are under-utilized due to the low amount and quality of extracted nucleic acids. We will discuss how SeqPlus can achieve efficient DNA extraction from FFPE samples, including aged samples as old as 15 years resulting in isolation of high-molecular weight fragments for next-generation sequencing. Whole-genome sequencing data derived from these samples revealed consistent sequencing quality as determined by Phred scores, GC content, duplicate reads, and sequencing coverage.  Variant analysis of these samples led to the identification of several well-known cancer driver genes. We also identified several genomic regions with altered copy number, including those previously reported. For the first time attain 98% sequence alignments covered at a depth of 20x with WGS and WES sequencing.

2:40 PM

Networking Break

TRACK 2: Understanding Genetic Data in the Bigger Picture of Clinical Assays and Personal Family History

3:10 PM

Gene therapy in Charcot-Marie-Tooth type 4J

Cat Lutz, Senior Director, In Vivo Pharmacology

The Jackson Laboratory

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Gene therapy in Charcot-Marie-Tooth type 4J

Rare diseases affect 350 million people globally with costly economic, social and societal implications. An estimated 50-75% of rare diseases affect children, with a 30% mortality rate before 5 years old. With the increasing use of exome sequencing in diagnostic process comes the opportunity to diagnose patients earlier in their disease, providing the best opportunities for therapeutic interventions. More than 250 new rare diseases are described annually and many of these diseases are so rare, the resources to understand the natural history of the patient population is lacking. The ability to rapidly engineer precise genetic mutations using CRISPR/Cas9 in the mouse can help to gain insight into the pathophysiology of these rare diseases and also provides a preclinical platform for testing therapeutics, such as gene therapy. As AAV delivery becomes more common in clinical trial designs, the timeline to move from preclinical testing to potential clinical trials decreases significantly. In this case study, we describe the accelerated pace by which we were able to move from patient diagnosis, to mouse models, to preclinical proof of concept data for an IND application in a rare form of Charcot-Marie Tooth disease.

Authors: Maximiliano Presa, Crystal Davis, Randy Walls, Jenn Cook, Cathleen Lutz

3:35 PM

DNBseq™, a proven & robust high-throughput next-generation sequencing technology

Zonghui Peng, Application Scientist

BGI Americas

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DNBseq™, a proven & robust high-throughput next-generation sequencing technology

DNBseq™ is a new sequencing technology developed by BGI’s subsidiary Complete Genomics™. Using DNA nanoball, combinatorial probe anchor synthesis, rolling-circle replication and patterned nano array technologies, it generates short reads at a large scale. From aspects of sequencing capacity, data quality, accuracy and sensitivity of variants detection and sequencing cost, we present the human whole-genome sequencing (WGS) and other NGS application datasets generated by DNBseq™ technology. We found that either for DNA sequencing or RNA sequencing, the data performances of DNBseq™ technology are highly comparable with Illumina technology, and it even turns out that DNBseq™ technology perform better than Illumina from perspective of some variants type calling, such as InDel of WGS.

Authors: Zonghui Peng

4:00 PM

QIAseq FastSelect: One-step, rapid removal of rRNA during whole transcriptome NGS library prep

Jonathan Shaffer, Associate Director, NGS Assay Product Development

QIAGEN

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QIAseq FastSelect: One-step, rapid removal of rRNA during whole transcriptome NGS library prep

Whole transcriptome NGS enables the characterization of both coding mRNAs and long noncoding RNAs (lncRNAs) from biological samples, including FFPE samples. However, before ultra-sensitive RNAseq can be performed on FFPE samples, cytoplasmic and mitochondrial rRNA should be removed to increase sensitivity and decrease the cost per sample.

To deplete rRNA, various methodologies exist, including hybridization/capture methods both as a pre-treatment or during post-library construction and methods which utilize enzymatic removal with target specific probes. However, these methods are arduous, not ideally suited for fragmented samples and may cause sample loss or distortion of transcriptomic profiles.

To remedy the complexity and the time necessary for rRNA removal in RNAseq applications, we have developed the QIAseq FastSelect RNA Removal kits which utilize a novel, one-step rRNA depletion technology. QIAseq FastSelect is compatible with both fresh and FFPE-fragmented RNA and stranded RNAseq libraries that utilize the dUTP or selective ligation method. Globin depletion kits are also available.

Lung cancer is the second most common cancer in both men and women. In 2018 alone, there were more than 200,000 new cases and over 150,000 deaths. One of the keys for developing new strategies around lung cancer treatment involves understanding the molecular pathology responsible for growth, metastasis and treatment failure. To determine this, whole transcriptome analysis, particularly next-generation sequencing (NGS), becomes a crucial technique in identifying druggable pathways and new biomarkers for patient stratification.

Here we utilized QIAseq FastSelect and strand-specific RNAseq to analyze the whole transcriptome of matched normal and tumor lung cancer FFPE samples. The resulting differentially expressed RNA signatures are being utilized for pathway analysis and biomarker evaluation for sample stratification. QIAseq FastSelect rapidly eliminates rRNA to enable the discovery of gene signatures locked away in FFPE samples. It is faster, more efficient, and more cost effective than existing solutions.

Authors:  Daniel Kim1, Melanie Hussong1, Christopher Howard1, Niels Tolstrup2, Sam Rulli1, Jonathan Shaffer1, Eric Lader1 (2 – QIAGEN Nordic)

 

1QIAGEN Inc, 6951 Executive Way, Frederick, MD 21703, 2QIAGEN Nordic, Skelstedet 16, 2950 Vedbæk, Denmark

4:25 PM

Advances Utilizing the 10x Genomics Chromium™ System for Single-Cell RNA-Seq

Alexander Lemenze, Technical Application Scientist

GENEWIZ

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Advances Utilizing the 10x Genomics Chromium™ System for Single-Cell RNA-Seq

Single-cell sequencing is at the forefront of next generation sequencing, rapidly expanding into a plethora of biological fields. GENEWIZ was one of the first service providers to offer single-cell transcriptomics through the 10x Genomics Chromium™ platform. Since its introduction, we have optimized upstream processes to deliver the most accurate and consistent data possible with this technology.

The success of single-cell experiments is critically dependent on the quantity of viable cells. As such, we developed a comprehensive workflow to best facilitate sample preparation and shipment. With our methods for cryopreservation and dead cell removal, we provide flexibility to scientists pursuing projects with low-input starting materials. Furthermore, we have successfully executed these protocols in experiments for drug discovery and evaluation of therapeutic responses.

With our single-cell workflows, customers can more easily prepare samples and achieve the highest quality results from their RNA-Seq projects.

Authors:  Alexander Lemenze, Chris Mozdzierz, Ryan Gutierrez

4:50 PM

Closing Comments Day 1

John Mudgett, Co-Founder and Chief Scientific Officer

JsM BioScience, LLC

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Closing Comments Day 1

5:00 PM

Reception