Performing a Trio Analysis in VSClinical

Editor's Notes

• #4: Hi. My name is Steven Hystad. I am a FAS at Golden Helix. We are a bioinformatics software and analytics company where we strive to enable the genetics research and translational genomics community by creating high quality software to analyze large genomic datasets. Today I am going to introduce you to own of our products VarSeq. Varseq is a platform for annotating, filtering, and prioritizing variants from DNA sequencing experiments. I'm going to introduce you to the Varseq interface by running through a gene panel workflow. We are a bioinformatics support company with over 17 years of expertise in the field of bioinformatics. Founded in 1998 Founder spun off from early work in pharmacogenomics at GlaxoSmith Kline, who was a key investorand still remain a solid partner with us. We have two flagship products Varseq and SNP and Variation Suite (SVS) for short. SVS is our research application platform. It’s a powerful analytic tool created to enable researchers to perform complex analysis and visualizations on genomic and phenotypic data. SVS has a broad range of tools to easily perform GWAS, Genomic Prediction, Differential expression analysis on RNA-Seq Data and has the ability to process CNV analysis. VarSeq is our testing platform for filtering and annotating variants of interest. Within VarSeq, We have the option to automatically create customizable clinical reports from the results of various workflows. We also have an add-in function called VSPipeline – take workflows that you created and automate the process with very little human interaction. Our pipeline function can be integrated a secondary analysis pipeline, so you can take VCF files coming out of your secondary pipeline and take it all through filtering, annotation, and creating a clinical report. The Data Warehousing add-in is designed to be installed on a server location. It’s a way of organizing samples within projects. So if your working on gene panels, whole genome/exome data you are going to have different samples for each one of those types projects. The warehouse function can take those samples and organize them in one central repository into each one of those projects. Projects can be queried, used as frequency annotations, host clinical reports, build assessment catalogs.
• #5: Our software has been very well received by the industry. We have been cited in over 1100 peer-reviewed publications and that’s a testament to our customer base.
• #6: Global We work with over 350 organizations comprising of pharmaceutical companies, top-tier institutions, government organizations, clinics, and genetic testing labs using our software. Approaching 10,000 installs of our products and 1,000’s of unique users.
• #7: When you purchase our software you don’t just get a product key and a goodbye and good luck. We do pride ourselves in having a great relationship with our customers. We want to work with you to ensure the software is staying relevant and is meeting all of your requirements that you have. We are a nimble organization and we can add features, functions, and algorithms rather quickly. Our executives attend and often speak at conferences within our industry. They really do a great job in staying relevant and being in the forefront of the industry. We are well known for quality products and excellent customer service. We have been developing software for over 17 years and have a wide range of talented experts on staff including biostatisticians, mathematicians and software engineers. We want to work with you to ensure the software is staying relevant and is meeting all of your requirements that you have. Service Training and Support – Our team of experts is available to provide the resources needed to help complete your analysis, from training, to technical support. We aim to support our customers every step of the way. Product Transparency - All major methods and algorithms are documented (including the math, use cases, and article citations). We completely shunned the “black box” model. We are not aware of another commercial competitor whose methods are as transparent as ours. Innovation and Speed – All of our software is designed for your standard desktop or laptop, without needing access to the cloud. We eliminate the need to work with a biostatics core and our clients can get results very often within hours and minutes. We provide the ability to leverage insights quicker. Golden Helix is a decisive edge in today’s competitive environment. Customizations – We offer an expandable and customizable platform via scripting, and many scripts can be downloaded on our website at no additional cost. We are also species agnostic, working with data from all diploid organisms: humans, plants and animals.
• #9: VarSeq is a power, flexible, and scalable variant annotation filtering and interpretation engine. This commercial grade software designed for your local hardware so it is a desktop application that is installed locally on your computer and is very simple to use (as well see). VarSeq also has GenomeBrowse built in which allows rich visualization of for a variety of data types built right into the software. In addition with Varseq we have the ability to build and run repeatable workflows within the GUI and on the command line with our VSpipeline add-on tool. So once you have developed a workflow in VarSeq, and decided the type of filtering you would like to do, what annotation sources you would like, you can save that workflow as a template which can really streamline the analysis time.
• #10: So now that we’ve got a better understanding of the company overall and the value that VarSeq provides, let’s discuss our objectives for todays webcast. First and foremost, we’re going to explore some basics of a trio analysis. Keep in mind todays project is a simple representation of a workflow, and you would likely develop a more elaborate workflow in your own projects. Nevertheless, we’ll cover how to access all the necessary algorithms and how to integrate them into a filter chain. The goal will then be to isolate some interesting variants through our filter chain and classify these variants in VSClinical. From VS Clinical we will touch on how to utilize the ACMG guideline tool to classify our variants, and finally produce a clinical report. Along the way, we will also cover some simple tips/tricks that you may find helpful when building your trio project. Before we jump into VarSeq, let’s discuss the project a bit more, and also cover the value that VSClinical provides to variant analysis.
• #11: Since the focus of this presentation is to demonstrate an alternative approach to Trio analysis, and eventually include the VSClinical tool, it is worth briefly mentioning the background of where this project came from. As some of you might know, we ship example projects with VarSeq; one being the Example YRI Exome Trio project. In this project, we imported the YRI trio with the daughter proband. The workflow (i.e. filter chain) aims to capture such variants as de novos, dominant hets, and compound hets among others. We will quickly reference this project after the presentation, to gain some perspective on different approaches of building a filter chain.
• #12: With VarSeq we have the ability to run workflows for Whole Exome and Genome data, Trio analysis, Tumor-Normal Workflows, and gene panel analysis as well. VarSeq comes with several default work flows that are built into the system. We call these default workflows templates. These templates are just a set of annotations sources and filter options that are available for any particular workflow. As you can see here are some examples of our default workflows. Trio Analysis Workflow – for Whole Exome trio analysis, that looking for 6 different inheritance patterns simultaneously within that project. It looks for things like De Novo Candidates, Compound heterozygous candidates and there are options for customizing those as well. Gene Panel Workflows available – some are specific to Hereditary Panels and some more specific cancer panels including a Tumor/normal pair analysis that’s available as well. They key to these workflows is that these are just a way with getting started with the software so if you are not familiar with how we filter/annotate and set things up, you can start with these and then customize your particular workflow for your needs. We do ship a couple of example projects that are available from the file menu. These example projects have data already loaded into them and the majority of analysis is already complete. There’s also a nice html document that walks you through what's been done in that project.
• #13: VaqSeq is a filtering and annotation engine so of course that means we are only as good as the annotations that we have available. Varseq is backed by an extensive list of publically curated data sources – this is where you are going to see the common variant frequency catalogs from 1k genomes, ExAC, NHLBI the exome variant server data. These are all free for use for anybody within the software. If there is a public source of data that we don’t currently support that you would like added to the software, all you have to do is let us know and we can look into getting that supported. With annotation sources and workflows in VarSeq, Any template created is going use a specific version of these annotations sources and lock it down and this is not something that we overwrite automatically. For example, you create a workflow and annotate your variants of interest using ClinVar annotation source. ClinVar releases updated information on a monthly schedule and we do try to stick to that monthly schedule. So once a month there is a new version of ClinVar that becomes available. if your workflow is specifying that you were using a (Octobers), then anytime you run that workflow, the (Octobers) version of ClinVar will be used. But we will provide a notification within the software that tells you when a new version of ClinVAR becomes available. So with just a few clicks, you can updating your workflow with a newer version of ClinVar. We are not going to do it for you because we are cognizant of anybody that needs to validate those sources and so your workflows don’t get overwritten. We also have a couple of annotation sources that we consider secure or cloud based annotations because these are not freely available for public use. There is a fee associated with these annotation sources to get access to them, but your sales representative can talk more about these when he talks licensing and getting everyone set up. So these sources are 1.) MedGenome OncoMD which is specific cancer database which provides drug targeting information as well as supporting clinical trial information specifically involving cancer mutations. 2.) OMIM information is available on the cloud source. We have their genes, phenotypes, and variants information. This information is a part of our reporting functionality and is used to fill in a lot of information regarding particular variants. We will explore this when we discuss reporting in VarSeq. 3.) CADD functional prediction scoring tools as well. If you have your own custom annotation source that you would like made available in the software, we have tools available that can basically take any delimited text file, a VCF file, or BED file just a wide variety of other sources that can be taken into the software using our convert wizard and converted into a source that can be used as an annotation source or filtering option within the software.
• #14: Lets discuss the goal we are trying to accomplish with the VS Clinical tool. We are looking for a simple way to leverage all the available evidence for a variant and score it for the potential impact it has on a disorder. The available evidence can be categorized into groups which includes considerations for gene function/phenotype association, variant segregation in a family, results of running variants through prediction algorithms or their presence in databases, but also utilizing previous discoveries you yourself have made for any variant. This collection of evidence is then linked to 33 criteria that the user can efficiently assess in a streamlined effort. These criteria then are aggregated and provide the basis for the final classifications of pathogenic, likely pathogenic, uncertain significance, likely benign or benign. When considering the amount of available evidence and requirements for eventually classifying variants, this process can become complex and difficult to master. Fortunately, VSclincal is a solution to this complexity for a number of reasons.
• #15: VS Clinical provides a means of simplifying not only the process of scoring and classifying variants, but also provides both a simple yet sophisticated means of presenting all evidence and criteria visually. As you work through the classification process, you will be presented with questions you are left to answer to connect the evidence to the best criteria, but VS also computes recommended answers while providing all the supporting evidence for each recommendation. One major concern for the process of answering these evidence-based questions and selecting the appropriate criteria is the repeatability of the resulting classification. A possible overlooked issue when manually going through the guideline process is the impact of fatigue. Thorough work at the beginning of the day may become a bit more loose as the afternoon rolls around. VSClinical removes all the variability and is easy to learn! Through VarSeqs intuitive workflow, any user not well versed in the ACMG guidelines can be brought up to speed rapidly and produce consistent results. Moreover, we have even implemented a capability of having multiple users make individual blind evaluations, which then can be compared to determine if multiple evaluations have consistent classifications. It is also worth mentioning that despite the application of a simplified workflow, the user can really deepdive into any of the supporting evidence to further clarify the variants impact/classification. Another advantage of getting new users up-to-speed is that the high workload of assessing variants is streamlined and less users can do more. On top of this, our team works to incorporate new developments to the guideline process so you don’t have to. This is a simple interpretation into the immense power behind the VS clinical ACMG workflow, and we have a number of other resources/publications that can shed more light on the content, but for now let’s focus on looking at an example workflow.
• #16: Incorporating VSClinical into a pre-existing project is incredibly simple. We can establish a filter chain that will allow us to rapidly narrow our search to a small set of variants we need to classify. From the small set of variants, the ACMG auto classifier will work to automatically score variants for 18 of the 33 criteria using previous findings in other samples but also 7 specific annotation sources. Any variants left to score that aren’t obvious are then passed through VS Clinicals intuitive ACMG guidelines so that the user can finalize the path to classification. Once classification is complete, the user then has access to generate custom clinical reports of their findings, all within the VS software.
• #17: Once we have annotated and filtered our variants of interest in VarSeq, We can create custom clinical reports based on our findings. For our reporting functionality, this is another option where we use the word “template” quite a bit. The templates in our reporting functionality are just a list of options for us to actually automatically create a given report. We have a basic example of a clinical report here (Point to example of report on slide) The template is really just a set of instructions, whether its sample level information, lab type information, how that variant is reported in the template. The template just tells VarSeq how to render that report. Please keep in mind, all of our templates can be customized, so if you need additional sample level fields or specific information regarding your variants reported in a different way, all of that can be changed to your particular needs. So the one I will show you today is just one example of a customizable report. VarSeq comes with three report templates. 1.) Hereditary workflows 2.) Cancer gene panels AND 3.) Exome Trio analysis. The take home message here is that the reports are configurable (point to slide). We can customize the sample level input, variant level inputs, and just overall customization. We’ll show you some examples once we get into that.