Establishing a stable, repeatable platform for measuring changes in sperm DNA methylation

Mohammad Abbasi,Peer Sangngern,Andrew D Smith,Harish Swaminathan,Amanda Douglas,Alan Horsager,Douglas T Carrell,Philip J Uren

doi:10.1186/s13148-018-0551-7

Abstract

BackgroundSeveral independent research groups have shown that alterations in human sperm methylation profiles correlate with decreased fecundity and an increased risk of poor embryo development. Moving these initial findings from the lab into a clinical setting where they can be used to measure male infertility though requires a platform that is stable and robust against batch effects that can occur between sample runs. Operating parameters must be established, performance characteristics determined, and guidelines set to ensure repeatability and accuracy. The standard for technical validation of a lab developed test (LDT) in the USA comes from the Clinical Laboratory Improvement Amendments (CLIA). However, CLIA was introduced in 1988, before the advent of genome-wide profiling and associated computational analysis. This, coupled with its intentionally general nature, makes its interpretation for epigenetic assays non-trivial.ResultsHere, we present an interpretation of the CLIA technical validation requirements for profiling DNA methylation and calling aberrant methylation using the Illumina Infinium platform (e.g., the 450HM and MethylationEPIC). We describe an experimental design to meet these requirements, the experimental results obtained, and the operating parameters established.ConclusionsThe CLIA guidelines, although not intended for high-throughput assays, can be interpreted in a way that is consistent with modern epigenetic assays. Based on such an interoperation, Illumina’s Infinium platform is quite amenable to usage in a clinical setting for diagnostic work.

Highlights

Several independent research groups have shown that alterations in human sperm methylation profiles correlate with decreased fecundity and an increased risk of poor embryo development
Interpretation of Clinical Laboratory Improvement Amendments (CLIA) for genome-wide sperm Deoxyribonucleic acid (DNA) methylation levels The Clinical Laboratory Improvements Amendment (CLIA) is a regulatory instrument that applies to facilities within the USA that perform laboratory tests on human tissue intended for use in diagnosis, health assessment or disease treatment
The Center for Medicare and Medicaid Services (CMS) oversees CLIA and certifies laboratories that are in compliance

Summary

Introduction

Several independent research groups have shown that alterations in human sperm methylation profiles correlate with decreased fecundity and an increased risk of poor embryo development. Moving these initial findings from the lab into a clinical setting where they can be used to measure male infertility though requires a platform that is stable and robust against batch effects that can occur between sample runs. The rapid drop in costs to profile genomic information has resulted in the development of numerous molecular laboratory-developed tests (LDTs), in cancer and reproductive health [1,2,3,4,5]. A molecular assay for measuring sperm quality would significantly improve our understanding of men’s reproductive health and allow for more targeted treatment of the infertile couple

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