Evaluating Splice Site Variants in VarSeq

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To adequately assess a variant's pathogenicity it is crucial to take into account the variant's effect on splicing. While mutations that disrupt the pairs of bases at the beginning of a splice site are straightforward to identify, detection of disrupted splice sites caused by changes to the splice motif is more difficult. In this webcast, we will discuss VarSeq's capabilities for detecting both disrupted and novel splice sites. This will include a deep dive into several splice site variants to examine their effect and assess their pathogenicity.

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Evaluating Splice Site Variants in VarSeq
Nathan Fortier, Ph.D. – Director of Research
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NIH Grant Funding Acknowledgments
▪ Research reported in this publication was supported by the National Institute Of
General Medical Sciences of the National Institutes of Health under:
- Award Number R43GM128485
- Award Number 2R44 GM125432-01
- Award Number 2R44 GM125432-02
- Montana SMIR/STTR Matching Funds Program Grant Agreement Number 19-51-RCSBIR-005
▪ PI is Dr. Andreas Scherer, CEO Golden Helix.
▪ The content is solely the responsibility of the authors and does not necessarily
represent the official views of the National Institutes of Health.

Golden Helix – Who We Are
Golden Helix is a global bioinformatics
company founded in 1998.
GWAS
Genomic Prediction
Large-N Population Studies
RNA-Seq
CNV-Analysis
Variant Warehouse
Centralized Annotations
Hosted Reports
Sharing and Integration
Variant Calling
Filtering and Annotation
Clinical Reports
CNV Analysis
Pipeline: Run Workflows

Cited in over 1,300 peer-reviewed publications

Golden Helix – Who We Are
When you choose a Golden Helix solution, you get more than
just software
▪ REPUTATION
▪ TRUST
▪ EXPERIENCE
▪ INDUSTRY FOCUS
▪ THOUGHT
LEADERSHIP
▪ COMMUNITY
▪ TRAINING
▪ SUPPORT
▪ RESPONSIVENESS
▪ INNOVATION and
SPEED

VSClinical
▪ Complete Support for ACMG Guideline
Workflow:
- Implements a guided workflow for following the ACMG guideline
scoring and classifying
- Place criteria into conceptually related groups, paired with their
opposites, and formatted as answerable question.
▪ Aggregate and Automate:
- Questions have supporting evidence presented with rich and
interactive visuals
- Automatically computed recommendations for questions that have
explicit bioinformatic evidence, with supporting reasons for each
answer.
▪ Expert and Beginner Friendly:
- Start with descriptive summaries and recommendations for a
variant
- Deep dive into Population Catalogs, Gene Impact, Published
Studies and Clinical tabs
- Integrated documentation, readings on scoring criteria and
citations

Splice Sites
▪ Understanding a variant’s impact on
splicing is crucial
▪ Synonymous variants can disrupt
existing splice sites or introduce
novel splice sites
▪ Such variants can cause exon
skipping or truncation

Splice Sites
▪ Introns have distinct nucleotide
pairs at each end
- GT at the 5’ end (Donor Site)
- AG at the 3’ end (Acceptor Site)
▪ Area surrounding nucleotide pair is
defined by a splice motif
▪ Sequences around splice sites are
highly variable
▪ Machine learning and probabilistic
methods are used to identify sites

Algorithms
▪ VSClinical supports four splice site prediction algorithms
- PWM: Uses position weight matrix similar to SpliceSiteFinder and
Human Splice Finder
- MaxEntScan: Approximates sequence motifs using Maximum
Entropy Distribution
- NNSplice: Identifies splice sites using neural networks
- GeneSplicer: Uses Markov models combined with maximal
dependence decomposition

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Comprehensive genomic testing via next generation sequencing (NGS) is being increasingly adapted as part of cancer care in conjunction with molecular and immunohistochemical tests. Comprehensive genomic profiling potentially expands the number of targeted therapies that are available to patients, improves patient diagnosis and prognosis, and increases the number of clinical trials that are relevant to patients. However, with these advancements come challenges such as gaps in expertise resulting in inadequate efforts to interpret genomic data accurately and efficiently, poorly coordinated efforts to implement precision care, patients being diagnosed and treated despite inadequate access to relevant information and subsequent lack of patient exposure to all available treatment options. Golden Helix CancerKB v2.0 provides a means of closing the gap, whether you're a beginner who is trying to capture the vast amount of knowledge in the cancer field or an expert who has high sample volume AND needs to keep up with the ever-evolving knowledge of Tier II and III variants. In this webcast, we will discuss and apply Golden Helix CancerKB in the context of cancer precision medicine. Golden Helix CancerKB is systematically curated and reviewed by experts in the field and contains information about cancer genes, biomarkers, and treatments generated from several trusted cancer resources. With VarSeq 2.3.0’s added support for comprehensive cancer genomic profiling tests, Golden Helix CancerKB has expanded to include interpretations for genomic signatures, combination biomarkers, and more investigational (tier II) biomarkers, among several other additions that will be discussed. With the Golden Helix CancerKB database, users will experience a streamlined automatic matching of biomarkers to available drugs and trials which ultimately saves users massive amounts of time and effort while reducing the possibility for errors.

Maximizing the Benefits of Comprehensive Genomic Testing in Cancer Care with ...

Golden Helix

VarSeq 2.3.0 facilitates the evaluation of a multitude of somatic genomic variations with a more refined user interface to streamline variant evaluation. Our recent webcasts have shown the full range of these newly developed upgrades: VarSeq 2.3.0: Supporting the Full Spectrum of Genomic Variation VarSeq 2.3.0: New TSO-500 and Genomic Signature Support in VSClinical AMP Now, we are showing it all in action from the user’s perspective. This webcast will provide a comprehensive demonstration of performing somatic variation analysis and reporting. We will review how to use workflow automation to expedite the NGS project creation process and report rendering. We will also demonstrate the streamlined capture of knowledge during variant evaluation by leveraging our clinical expert-curated interpretations with the Golden Helix Cancer Knowledge Base (CancerKB). We hope you will join us to see VarSeq 2.3.0 from a user’s perspective, covering: -Somatic variant workflows: necessary algorithms and filtering strategies -Import of all relevant biomarker and genomic signatures data from TSO-500 -Review content and value of clinically curated interpretations and treatments with CancerKB -Interpretation of structural variants in the VSClinical AMP Guidelines workflow -Workflow automation with VSPipeline

A User’s Perspective: Somatic Variant Analysis in VarSeq 2.3.0

Golden Helix

Next Generation Sequencing allows for the detection of a wide variety of genomic alterations. This includes small mutations, copy number variants and complex rearrangements. However, it can be difficult to annotate, filter, and interpret these alterations. As part of our VarSeq 2.3.0 release, we have greatly simplified this process by allowing you to import, annotate, and filter mutations across all spectrums of genomic variation. This supports concurrent importation of small variants and CNVs as well as complex rearrangements. This release also includes strong support for structural variant annotation, filtering, and interpretation, including structural variant effect prediction. After filtering is complete, any clinically relevant structural variants can be interpreted with the VSClinical AMP Guidelines workflow and included in the final clinical report. Come join us for this webcast to discuss VarSeq’s enhanced import and annotation capabilities, including: Concurrent importation of variants CNVs and complex rearrangements Improved multi-threaded import which dramatically speeds up the importation of large VCFs Annotation of structural variants and prediction of effect Interpretation of structural variants in the VSClinical AMP Guidelines workflow Support for visualization and use of CRAM files as input for computing coverage statistics

VarSeq 2.3.0: Supporting the Full Spectrum of Genomic Variation

Golden Helix

Precision medicine for cancer is rapidly accelerating because of the development and approval of targeted molecular therapies. These therapies require new genomic biomarkers as an indication for use, and require evaluating additional mutation types that are available in comprehensive genomic profiling assays as well as the small variants detected by Next-Generation Sequencing gene panels. We are excited to announce VarSeq 2.3.0 which will update the VSClinical AMP workflow to meet the growing needs of labs conducting comprehensive genomic profiling (CGP) of tumors. This includes built-in support for the Illumina TruSight Oncology 500 (TSO-500) kit as well as similar kits from other vendors. The VSClinical AMP workflow has also gained native support for the bioinformatic outputs of CGP kits. Join us to learn about comprehensive genomic profiling in cancer, specifically: Evaluation and clinical reporting of genomic signatures such as Microsatellite (MSI), Tumor mutation burden (TMB), PD-L1, Homologous recombination deficiency (HRD) statuses, and more. Built-in TSO-500 import and expandable import capabilities for new genomic data types through the new advanced workflow scripting system. Golden Helix CancerKB updates with report-ready genomic-signature interpretations written for approved therapies as well as gene interpretations for all 500 genes of the TSO-500 panel. In addition, CancerKB scopes have been extended to reference multiple relevant biomarkers in a single interpretation, capture approved therapies at the tumor type level, and include interpretations for clinically relevant negative findings. Expanded clinical trial support to include international trials and the ability to search within proximity of European postal codes. VSClinical is accessing all active studies in AACT/ClinicalTrials.gov wherein users can search and select trials based on relevant drugs, biomarkers, and the geographic distance to the patient or testing site. VarSeq 2.3.0 will deliver powerful capabilities for genomic profiling in cancer, enabling a new level of personalized and effective care for your patients. We look forward to demonstrating these updates and Golden Helix’s continued innovation making the VarSeq Clinical Suite the NGS analysis platform of choice for germline and cancer testing.

VarSeq 2.3.0: New TSO-500 and Genomic Signature Support in VSClinical AMP

Golden Helix

One major hurdle facing medicine is the need to quickly identify and assess the genetic components contributing to rare diseases. It has been estimated that nearly 350 million people suffer from rare diseases, 140 million of which are children, of whom ~30% do not live past their fifth birthday1,2. The specific issue to overcome is reducing morbidity by facilitating rapid diagnosis and treatment. Fortunately, the cost of whole genome sequencing has dropped below the $1000 mark, which not only makes the NGS approach more affordable but has become the status quo method of comprehensive diagnosis for these rare disorders. Currently, there are limited options in the market when it comes to quality software that can scale to this size of data and handle variant processing and evaluation in a timely fashion. Fortunately, Golden Helix has sought to set the market standard for top-quality NGS analysis with our bioinformatic software VarSeq. The focus of this webcast will be to explore example workflows tailored for rare disorders and elaborate on how best to expedite the NGS pipeline process with our command-line tool VSPipeline. During the webcast, we will address the following: Customizing clinically validated NGS workflows with VarSeq for both single sample and trios Demonstrating the automation of ACMG-based guideline review in the VSClinical variant interpretation hub and rendering of customized clinical reports Expediting the NGS workflow via Golden Helix command-line tool VSPipeline We look forward to you joining us for our presentation, where we can demonstrate the value of our products when building your next-gen workflows. Ultimately, we wish to diminish the intimidation of genome workflow design and leave our future customers feeling confident that there is capable software to suit their needs. Bick D, Jones M, Taylor SL, et al. Case for genome sequencing in infants and children with rare, undiagnosed or genetic disease. J Med Genet 2019; 56:783-791. Owen M, Lefebvre S, Hansen C, et al. An automated 13.5 hour system for scalable diagnosis and acute management guidance for genetic diseases. Nat Commun 2022; 13: 4057. https://doi.org/10.1038/s41467-022-31446-6

Single Sample and Family Based Genome Analysis With VarSeq

Golden Helix

Somatic analysis is a complex and precise process that is constantly evolving. As the volume of available data and the accessibility of sequencing technology increase, so too does the value of a versatile, well-vetted, and efficient workflow solution. In this webcast, we will take a deep dive into the current state of our AMP interpretation software and explore various ways to optimize workflows. For anyone from grizzled VarSeq veterans to those seeing our software for the first time and labs of any size, we will provide a practical overview of our somatic analysis capabilities and how those capabilities scale with improving technology. Throughout this webcast we will be discussing the following: - Universal principles of somatic workflows, providing baseline recommendations - Specific tumor-normal and somatic-only use cases - VSClinical AMP interpretation hub and some variants of interest - Opportunities for automation and how to decrease time to report for increased throughput Join us as we show off the versatility and scalability of our AMP interpretation capabilities!

User perspective for somatic variant analysis in VSClinical AMP

Golden Helix

The automation of clinical NGS workflows provides a number of important benefits for labs. Automation reduces the time required to produce a clinical report, mitigates the possibility of human error, and improves the precision of clinical results. In turn, these benefits create higher profitability from a P&L perspective. Golden Helix software is designed to meet these needs by automating the full analysis workflow from sequencer to clinical report on a fixed annual subscription model. We are looking forward to discussing the best practices of maximizing profitability in your NGS testing lab and how Golden Helix supports these efforts. Join us in this webinar as we cover how to develop repeatable cancer and germline interpretation workflows that scale from panels to whole exomes and genomes.

Maximizing Profitability in your NGS Testing Lab

Golden Helix

VarSeq has become renowned for the accuracy of its CNV Caller, and the ease with which VSClinical takes the user through CNV evaluation. Cited in many publications, this well-validated tool brings our customers the ability to run both variant and CNV interpretations in one program, on data ranging from Gene Panels to whole genomes. What is less well known about CNV analysis through VarSeq, is that our users are not only limited to CNVs called through our software. VarSeq CNV is able to import CNVs in several file formats (VCFs, text, or tsv) generated by a wide variety of secondary callers, allowing the user to analyze their externally derived data. In this webcast, we will take you through the basics of CNV analysis with both the VarSeq CNV caller and from several common external CNV callers. Join us in this webinar as we cover: Leveraging the Copy Number Probability and Segregation Algorithm to add power to a CNV Trio workflow. Importing externally called CNVs. Expediting the path to analysis with CNV specific templates. Utilizing the auto-recommendations to efficiently analyze the pathogenicity of several CNV calls and generate clinical reports.

Handling a Variety of CNV Caller Inputs with VarSeq

Golden Helix

In the era where cloud-based solutions are the default for the modern office, it may not be obvious why many laboratories and testing centers choose to host their data and analysis pipelines on-premises or on self-managed cloud services. Next-generation sequencing enables a precision medicine approach to rare disease diagnostic and cancer therapeutics through its power to detect unique variants in individuals. This data is generated quickly and cheaply but requires a lot of disk space and processing power to arrive at clinically useful insights. When providing a clinical lab service under a regulated environment: data security, long-term affordable storage, and versioning through locked-down pipelines are all factors that must go into the choice of whether to choose a hosted analytics platform versus on-premises solutions or self-managed cloud infrastructure. Join us in this webinar as we cover: The validation and regulatory requirements that inform infrastructure and hosting decisions for NGS labs The cost structure of scaling NGS labs to exomes and genomes Deployment and security architecture for on-premises and self-managed cloud infrastructure Validating and versioning analysis pipelines with clinical tests through self-managed software lifecycles and versioned annotation sources Cybersecurity, patient data privacy, and scalable unit economics play a bigger role than ever before in the planning of NGS lab’s infrastructure choices. We look forward to you joining us as we tackle the trade-offs and choices around these topics and how deployment flexibility is a core feature of the Golden Helix VarSeq Suite.

Evaluating Cloud vs On-Premises for NGS Clinical Workflows

Golden Helix

As our users have come to know, VarSeq serves as a hub for variant annotation and the full interpretation/classification of germline (ACMG) and somatic (AMP) variants. Whether direct annotation or backend variant evidence is being presented to the user via VSClinical for the interpretation process, users greatly benefit from the hosted variant databases being available directly from VarSeq. Our team has automated much of the curation process and hosts the ongoing updates to these tracks so that users no longer suffer manual review of each database via the web or manual curation efforts. Useful databases include ClinVar and ClinGen for classification submissions, gnomAD exomes/genomes for filtering out common variants in the population, RefSeq for gene impact and sequence ontology assessment, and OMIM for phenotypic information. Obviously, there is a large collection of databases out there, and not all of them make it into our automated queue. However, GHI supports the utilization of custom databases in our software. This webcast will expose features of custom database curation/utilization in VarSeq to optimize your NGS workflows even further. During the presentation, we will discuss many different approaches with custom annotations, including: Interval Tracks: Bed files defining target regions for coverage calculations and CNV detection. Assessment catalogs: record keeping of variant classification/interpretations in VSClinical. Frequency catalogs: approaches to capture all variant allele frequencies at a project level and cohort level with VSWarehouse.

VarSeq Custom Database Curation Capabilities.pdf

Golden Helix

The automation of clinical NGS workflows provides a number of important benefits. Automation reduces the time required to produce a clinical report, mitigates the possibility of human error, and improves the precision of clinical results. In this webcast, we will discuss how the VarSeq Suite can be leveraged to automate the full analysis workflow from sequencer to clinical report. Join us as we demonstrate how VarSeq’s automation capabilities can enable your laboratory to: Automatically perform secondary analysis when a new sequence run is complete Go from FASTQ to BAM and high-quality variants in VCFs using Sentieon Automatically start VSPipeline to go from raw VCFs to candidate variants Compute coverage and call CNVs alongside small variants with VS-CNV Efficiently interpret a small set of annotated candidate variants and CNVs Draft reports with VarSeq and VSClinical Join us as we discuss the automation of the clinical analysis process for NGS genetic tests from FASTQ to Clinical Reports using the VarSeq Suite and discover how your laboratory’s NGS workflows may benefit from these automation capabilities.

Automating Clinical Workflows with the VarSeq Suite

Golden Helix

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