PGx Analysis in VarSeq: A User’s Perspective

Since our release of the PGx capabilities in VarSeq, we’ve had a few months to gather some insights from various use cases. Some users approach PGx workflows by means of array genotyping or what seems to be a growing trend of adding the star allele calling to the existing NGS pipeline for whole genome data. Luckily, both approaches are supported with the VarSeq software platform. The genotyping method being used will also dictate what the scope of the tertiary analysis will be. For example, are your PGx reports a standalone pipeline or would your lab’s goal be to handle a dual-purpose workflow and report on PGx + Diagnostic findings.

The purpose of this webcast is to:

Discuss and demonstrate the approaches with array and NGS genotyping methods for star allele calling to prep for downstream analysis.
Following genotyping, explore alternative tertiary workflow concepts in VarSeq to handle PGx reporting.

Moreover, we will include insights users will need to consider when validating their PGx workflow for all possible star alleles and options you have for automating your PGx analysis for large number of samples. Please join us for a session dedicated to the application of star allele genotyping and subsequent PGx workflows in our VarSeq software.

1. PGx Analysis in VarSeq : A User’s Perspective June 19, 2024 Presented by Solomon Reinman, Technical Field Application Scientist and Darby Kammeraad, Director of Customer Success

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3. PGx Analysis in VarSeq : A User’s Perspective June 19, 2024 Presented by Solomon Reinman, Technical Field Application Scientist and Darby Kammeraad, Director of Customer Success

4. Today’s Presenters 4 PGx Analysis in VarSeq : A User’s Perspective Solomon Reinman Technical Field Application Scientist Darby Kammeraad Director of Customer Success

5. NIH Grant Funding Acknowledgments 5 Research reported in this publication was supported by the National Institute Of General Medical Sciences of the National Institutes of Health under: Award Number R43GM128485-01 Award Number R43GM128485-02 Award Number 2R44 GM125432-01 Award Number 2R44 GM125432-02 Montana SMIR/STTR Matching Funds Program Grant Agreement Number 19-51-RCSBIR-005 NIH SBIR Grant 1R43HG013456-01  PI is Dr. Andreas Scherer, CEO of 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.

6. Golden Helix at-a-Glace 6 Company Snapshot: Leading SaaS provider of tertiary genomic analysis solutions for NGS labs Golden Helix is a SaaS bioinformatics solution provider specializing in next-gen sequencing (“NGS”) data analysis  The Company’s software enables automated workflows and variant analysis for gene panels, exomes, and whole genomes  Key Clinical Applications Prenatal testing Hereditary disease testing Reproductive testing Oncology Marquee Global Clients Golden Helix’s solutions allow clients to increase throughput, ensure consistent quality, maximize revenue, and save time  1998 Company Founded Bozeman, Montana Headquarters Recognitions Government Research Pharmaceuticals Agrigenomics Testing Labs Translational Labs Human Genetics Research Hospitals Academia Publications Content & Resources Pharmacogenetics testing

7. NGS Clinical Workflow Golden Helix provides comprehensive data analytics software that scales across gene panels, whole exomes, and whole genomes DNA Extraction in Wet Lab and Sequence Generation Interpretation and Result Reporting Primary Read Processing and Quality Filtering Alignment and Variant Calling Secondary *Golden Helix provides Secondary Analysis through a reseller agreement Tertiary Golden Helix’s software and primary focus Comprehensive secondary and tertiary analysis solutions for primary data aggregated by all commercially available sequencers Type Size Gene Panel Small (100MB) Whole Exome Medium (1GB) Whole Genome Large (100GB) Cancer use case Hereditary use case Process Analysis … and scales across multiple data set sizes for cancer and hereditary use cases Filtering and Annotation Data Warehousing Workflow Automation Golden Helix works with all major sequencers… Medical Device Certification

8. ISO Certification 13485:2016 8 ISO 13485:2016 from TÜV SÜD ISO 13485:2016 is an international standard that specifies requirements for a quality management system (QMS) for organizations involved in the design, development, production, and servicing of medical devices. maintain a quality management system demonstrate sufficient risk management show consistent tracking of customer satisfaction and safety in the market demonstrate continued improvement efforts on the product and system level.  ISO 13485:2016 is designed to objectively document that we are holding ourselves to the highest quality standards as we are providing innovative solutions to hospitals, testing labs, and research institutions globally.

9. Overview 9 Recap on PGx feature release in VarSeq 2.6.0 Recent webcasts: https://www.goldenhelix.com/resources/webcasts/index.html VarSeq 2.6.0: Advancing Pharmacogenomics and Genomic Analysis Introducing VSPGx : Pharmacogenomics Testing in VarSeq Users Perspective with PGx PGx diplotype calling and star alleles: Options for genotyping with NGS or array VarSeq Workflow design options: PGx stand-alone or Diagnostic + PGx reporting Options for report customization Deployment and Validation considerations Automation options Investigating PGx markers

10. Clinical Pharmacogenomic Implementation Consortium (CPIC) 10 CPIC has developed best practice guidelines for pharmacogenomic tests Includes standardized grading of evidence linking genotypes to phenotypes Assigning phenotypes to genotypes and diplotypes Prescribing recommendations based on diplotype and phenotypes CYP2D6 Diplotype Metabolizer Phenotype CYP2D6 *1/*1 Normal metabolizer CYP2D6 *2/*122 Intermediate metabolizer CYP2D6 *3/*3 Poor Metabolizer CYP2D6 *1/*1x2 Ultrarapid metabolizer CYP2D6 *5/*5 Poor metabolizer

11. Secondary options: NGS genotyping 11

12. Secondary options: Conversion of array data 12

13. PGx Variant Detection and Recommendations Algorithm 13 Algorithm Function (In VarSeq ) Diplotype caller Phenotype and drug recommendation annotation Algorithm Requirements: Call variants at all required positions for PGx genes Required annotations Optional Customization Customize annotation sources used by the algorithm Recommendations can also be provided for structural variant data Variant calling (Prior to VarSeq ) 1. Array data converted to vcf for VarSeq import 2. Utilize NGS pipeline (ex. Sentieon ) Call alternate calls along with must call positions GVCF calling hard filtered to PGx markers in import

14. Pharmacogenomic Report 14 What’s in the PGx Report from VarSeq ? Current Patient Medications and Recommendations Gene-Drug Interactions Prescribing Recommendations Phenotype Associations Tested Alleles Word-based reports are fully customizable Reports can be rendered for a single sample or a batch of samples

15. 15 How to deploy both workflows in one template Filter 1: PGx diplotypes Filter 2: Diagnostic workflow driven by sample phenotype PGx + Diagnostics

16. Demo 16 Genotyping: Review of NGS calling and array conversion script examples gVCF subset on VarSeq import Usecase 1: PGx standalone Usecase 2: PGx + Diagnostic analysis

17. 17 Product Demo

18. NIH Grant Funding Acknowledgments 18 Research reported in this publication was supported by the National Institute Of General Medical Sciences of the National Institutes of Health under: Award Number R43GM128485-01 Award Number R43GM128485-02 Award Number 2R44 GM125432-01 Award Number 2R44 GM125432-02 Montana SMIR/STTR Matching Funds Program Grant Agreement Number 19-51-RCSBIR-005 NIH SBIR Grant 1R43HG013456-01  PI is Dr. Andreas Scherer, CEO of 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.

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20. eBook Library 20 Prenatal Genetics – Learn the Following Existing approaches to prenatal WES, along with clinical indications for its use   How VarSeq and VSClinical can be utilized for its use A few interesting cases of variants and their classifications Pharmacogenetics – Learn the Following Foundations of Pharmacogenomics Genetic variability and drug response Pharmacogenomic test reporting nomenclature and terminology The Pharmacogenomic eco-system VSPGx - A pharmacogenomics application

21. Secured CE Mark for EU 21 VarSeq Dx VarSeq Dx is our flagship software, VarSeq , that is CE marked to meet the European In Vitro Diagnostic Regulation (IVDR 2017/746) requirements. VarSeq Dx satisfies the IVDR requirements within the European Economic Area (EEA). Verification Our customers will work with our Field Application Scientist to verify the installation and ensure proper usage of the software. This can be used for ISO QMS software validation documentation. User Certification Tiered certification option to ensure training and proper usage of the various VarSeq Suite modules. User Proficiency Certificate upon successful completion of evaluation.

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