METHYLOME IMPACTS OF CANNABIS USE IN THE CHRISTCHURCH HEALTH AND DEVELOPMENT STUDY

Background Cannabis has been very much in the news of late, with controversy and debate surrounding legalisation and medicinal uses purposes. However, there is also strong evidence for negative consequences of prolonged cannabis use. Significant contributions in this area have been made by the Christchurch Health and Development Study (CHDS), a longitudinal study of a birth cohort of 1265 children born in 1977 in Christchurch (New Zealand), who have been studied on 23 occasions from birth to the age of 35 (n=987). Methods It is now clear that many environmental factors, including drugs, impact on the pattern of epigenetic marks throughout the genome. We sought to explore the impacts of heavy cannabis use on the methylome in participants of the CHDS. We selected DNA (purified from peripheral blood samples) of 96 CHDS subjects including 49 heavy long-term cannabis smokers (25 of whom also smoked tobacco), and 47 non-smoking, non-cannabis using controls. Epigenome wide analysis of methylation was carried out using Illumina 850 K EPIC arrays. Results Our key observations were that for the cannabis plus tobacco smokers compared with non-smoking controls, the most differentially hypermethylated sites were in the AHRR and F2RL3 genes (adjusted P 4.4×10^-8, 4.9×10^-6 respectively), replicating multiple previous studies on methylomic effects of tobacco. For those sites (~15000) significantly showing greater than 5% differential methylation after multiple comparison adjustment, 99% were hypermethylated. Comparing cannabis only users with non-smoking controls, for those sites (~4500) significantly showing greater than 5% differential methylation after multiple comparison adjustment, 99% were hypomethylated. The differentially methylated sites which were hypermethylated in tobacco smokers but hypomethylated in cannabis only users were most strongly represented in PI3K-Akt and oxytocin KEGG pathways (FDR corrected p values Discussion These data suggest that cannabis has quite different methylome effects to tobacco smoking, and that there may be interaction effects which modify the methylome impacts when both cannabis and tobacco are smoked. We are seeking to validate a proportion of these methylation changes using bisulfite-amplicon sequencing in a wider selection of samples from the CHDS and other cohorts. These analyses may help to identify epigenome impacts of cannabis use, and point to gene regulatory processes that underlie both potentially positive as well as adverse consequences of its use.