Functional characterization of novel rare CYP2A6 variants and potential implications for clinical outcomes

CYP2A6 activity, phenotyped by the nicotine metabolite ratio (NMR), is a predictor of several smoking behaviors, including cessation and smoking‐related disease risk. The heritability of the NMR is 60–80%, yet weighted genetic risk scores (wGRSs) based on common variants explain only 30–35%. Rare variants (minor allele frequency <1%) are hypothesized to explain some of this missing heritability. We present two targeted sequencing studies where rare protein‐coding variants are functionally characterized in vivo, in silico, and in vitro to examine this hypothesis. In a smoking cessation trial, 1687 individuals were sequenced; characterization measures included the in vivo NMR, in vitro protein expression, and metabolic activity measured from recombinant proteins. In a human liver bank, 312 human liver samples were sequenced; measures included RNA expression, protein expression, and metabolic activity from extracted liver tissue. In total, 38 of 47 rare coding variants identified were novel; characterizations ranged from gain‐of‐function to loss‐of‐function. On a population level, the portion of NMR variation explained by the rare coding variants was small (~1%). However, upon incorporation, the accuracy of the wGRS was improved for individuals with rare protein‐coding variants (i.e., the residuals were reduced), and approximately one‐third of these individuals (12/39) were re‐assigned from normal to slow metabolizer status. Rare coding variants can alter an individual’s CYP2A6 activity; their integration into wGRSs through precise functional characterization is necessary to accurately assess clinical outcomes and achieve precision medicine for all. Investigation into noncoding variants is warranted to further explain the missing heritability in the NMR.

Study Highlights

• WHAT IS THE CURRENT KNOWLEDGE ON THE TOPIC?

Common CYP2A6 variants (minor allele frequency >1%) explain 30–35% of the variation in CYP2A6 activity, despite high heritability estimates (60–80%) in the CYP2A6 activity biomarker measure. One hypothesis is that rare coding variants (minor allele frequency <1%) may explain a portion of the missing heritability from pharmacogenes, including CYP2A6.

• WHAT QUESTION DID THIS STUDY ADDRESS?

What is the relative contribution of rare coding variants in explaining variation in CYP2A6 activity? How necessary is the incorporation of rare coding variants in predicting individual metabolic status, and consequent tailoring of treatment?

• WHAT DOES THIS STUDY ADD TO OUR KNOWLEDGE?

Rare coding variants may explain only a small fraction of the variation on a population level; however, their role may be important on an individual level, altering the predicted metabolic status in a third of the individuals with these rare coding variants.

• HOW MIGHT THIS CHANGE CLINICAL PHARMACOLOGY OR TRANSLATIONAL SCIENCE?

Evaluating rare coding variants in pharmacogenes, such as CYP2A6, will be valuable in enhancing the investigation of CYP2A6’s influence on tobacco addiction and disease pathogenesis, by providing a more accurate reflection of the phenotypic metabolic status through improved genetic assessments.     

Secondary structure of the Tetrahymena ribosomal RNA intervening sequence: structural homology with fungal mitochondrial intervening sequences.

Splicing of the ribosomal RNA precursor of Tetrahymena is an autocatalytic reaction, requiring no enzyme or other protein in vitro. The structure of the intervening sequence (IVS) appears to direct the cleavage/ligation reactions involved in pre-rRNA splicing and IVS cyclization. We have probed this structure by treating the linear excised IVS RNA under nondenaturing conditions with various single- and double-strand-specific nucleases and then mapping the cleavage sites by using sequencing gel electrophoresis. A computer program was then used to predict the lowest-free-energy secondary structure consistent with the nuclease cleavage data. The resulting structure is appealing in that the ends of the IVS are in proximity; thus, the IVS can help align the adjacent coding regions (exons) for ligation, and IVS cyclization can occur. The Tetrahymena IVS has several sequences in common with those of fungal mitochondrial mRNA and rRNA IVSs, sequences that by genetic analysis are known to be important cis-acting elements for splicing of the mitochondrial RNAs. In the predicted structure of the Tetrahymena IVS, these sequences interact in a pairwise manner similar to that postulated for the mitochondrial IVSs. These findings suggest a common origin of some nuclear and mitochondrial introns and common elements in the mechanism of their splicing.     

New discoveries and directions for medical, dental and dental hygiene research: low temperature atmospheric pressure plasma

Abstract:  The study of plasma integrates physics, chemistry, biology, and engineering, and has recently engaged medicine and dental hygiene in research efforts. The study of plasma holds promise for a myriad of applications ranging from lasers and electronics, hazardous waste management, decontamination, sterilization and disinfection of foods, soil, water, instruments, to medical uses in wound healing and treating certain types of tumours and cancers. Plasma represents a new state-of-the-art sterilization and disinfection treatment for certain oral and enviornmental pathogens, heat-sensitive materials, contaminated medical waste, hard and soft surfaces, and ventilation systems may assist health care facilities in the management of various health concerns. The role that Low Temperature Atmospheric Pressure Plasma (LTAPP) could play in the inactivation of pathogenic microorganisms might prove to be a new, faster, noncorrosive, more economical alternative, as well as support green healthcare.