CYP2B6-Related Disorders via the CYP2B6 Gene

CYP2B6 is a member of the cytochrome P450 superfamily of enzymes, and like other members of the family, is responsible for catalyzing reactions that lead to the metabolism and bioactivation of drugs and toxins as well as the synthesis of endogenous molecules like cholesterol, steroids, and lipids (Manikandan and Nagini. 2018. PubMed ID: 28124606). Compared to CYP3A4, which accounts for ~40% of the liver CYP content, CYP2B6 accounts for 2-10% (Wang and Tompkins. 2008. PubMed ID: 18781911). CYP2B6 has been classified as a “Very Important Pharmacogene” (https://www.pharmgkb.org/vip/PA166169423) and is responsible for metabolizing ~4% of the top 200 most prescribed drugs (Thorn et al. 2010. PubMed ID: 20648701; Zanger et al. 2008. PubMed ID: 18695978). Polymorphisms in CYP2B6 have been linked to adverse drug events for medications such as efavirenz, methadone, artemisinin, bupropion, cyclophosamide, ketamine, and nevirapine (https://cpicpgx.org/genes-drugs/; Zanger and Klein et al. 2013. PubMed ID: 23467454).

The relationship between CYP2B6 polymorphisms and adverse drug reactions for efavirenz treatment has been well documented (Desta et al. 2019. PubMed ID: 31006110). Efavirenz (brand name: Sustiva) is an antiretroviral medication used in the treatment and prevention of HIV/AIDS and works by blocking the function of HIV reverse transcriptase, preventing virus proliferation. Efavirenz is often administered in combination with other antiretroviral drugs and has proven to be effective in not only treating individuals with HIV/AIDs, but also in prophylactic treatment of individuals who have had a potential HIV exposure through a needle stick or other event. Efavirenz is consumed as an active drug. It is metabolized to an inactive form through a hydroxylation reaction by CYP2B6 and eventually excreted in urine. Other CYP enzymes such as CYP3A4, CYP2A6, and CYP1A2 also play a minor role in efavirenz metabolism.

Individuals that have decreased or absent CYP2B6 metabolism (i.e. poor metabolizer status) and are treated with efavirenz are at risk for increased plasma concentrations and decreased clearance of the drug which can lead neurotoxicity, dizziness, insomnia, drowsiness, impaired concentration, abnormal dreams, psychosis, suicidal thoughts, hallucinations, and depression (King and Aberg. 2008. PubMed ID: 18753940; Desta et al. 2019. PubMed ID: 31006110). Central nervous system toxicity has been observed in 40-60% of individuals taking efavirenz (Apostolova et al. 2015. PubMed ID: 26203180). For most individuals, these effects resolve within a few days to weeks following treatment initiation (King and Aberg. 2008. PubMed ID: 18753940). However, for some people taking efavirenz, these effects are serious enough to impact quality of life and can lead to discontinuation of treatment (Zanger and Klein et al. 2013. PubMed ID: 23467454; Desta et al. 2019. PubMed ID: 31006110). Individuals that have increased CYP2B6 metabolism (i.e. ultra-rapid metabolizer status) and are treated with efavirenz are at risk of lowered plasma concentrations of the drug which can lead to drug resistance and treatment failure (Zanger and Klein et al. 2013. PubMed ID: 23467454).

Besides its role in drug metabolism, variants in CYP2B6 have also been associated with leukemia (Yuan et al. 2011. PubMed ID: 20878158; Yu et al. 2020. PubMed ID: 32119768; Daraki et al. 2016. PubMed ID: 27865701) and Hirschsprung's disease (Xu et al. 2015. PubMed ID: 25424204; Yang et al. 2019. PubMed ID: 31240788). Although clinical and functional studies are limited, it has been hypothesized that defects in CYP2B6 metabolism prevent xenobiotic metabolism, leading to the buildup of environmental pollutants and carcinogens within the body. This buildup can then cause additional DNA damage and chromosomal aberrations that increase the risk for leukemia development or congenital disorders such as Hirschsprung’s disease (Yuan et al. 2011. PubMed ID: 20878158; Xu et al. 2015. PubMed ID: 25424204).

Germline testing of CYP2B6 can help identify variants that may affect an individual’s response to a particular medication metabolized by CYP2B6. This information, along with clinical pharmacogenetics-based dosing guidelines, can be used as by healthcare providers to optimize drug therapies for patients through dosage adjustments or alternative drug recommendations. Clinical dosing guidelines have been established for CYP2B6 and efavirenz (https://cpicpgx.org/guidelines/; Desta et al. 2019. PubMed ID: 31006110). Although evidence exists for adverse reactions between CYP2B6 and other drugs such as methadone (Victorri-Vigneau et al. 2019. PubMed ID: 30907440) and bupropion (Wang et al. 2020. PubMed ID: 32238417), the evidence is considered provisional at this time and clinical dosing guidelines have been not been developed. Pre-emptive pharmacogenetic testing (occurring prior to a patient receiving a prescription) has been shown to improve clinical outcomes with benefits including improved efficacy, improved safety, reduced cost, reduced hospitalization, and improved adherence (Krebs and Milani. 2019. PubMed ID: 31455423).

The CYP2B6 gene is located on chromosome 19 at 19q12-q13.2 within a large cluster of other CYP2 genes known as the CYP2ABFGST cluster (Zanger and Klein. 2013. PubMed ID: 23467454; Hoffman et al. 2001. PubMed ID: 11692077). Is it composed of 9 exons which encode the 491 amino acid protein CYP2B6. CYP2B6 has a closely related pseudogene, CYP2B7P, which is located ~41 kb upstream. Comparison of CYP2B6 and CYP2B7 reveal highly similar sequences that vary at only 61 nucleotides residues throughout their nine exons (https://a.storyblok.com/f/70677/x/df34ae40b5/cyp2b6_variation_v1-1.pdf).  

CYP2B6 is expressed primarily in the liver, comprising ~2-10% of total hepatic CYP content. CYP2B6 is also expressed at much lower levels in tissues with barrier functions such as the skin, kidney, lung, and nasal mucosa. It has been hypothesized that its presence in these additional tissues helps to protect the body from contact with chemicals and environmental pollutants (Zanger and Klein et al. 2013. PubMed ID: 23467454). Expression of CYP2B6 can vary widely between individuals (>100-fold). Much of this variability has been attributed to the ability of the CYP2B6 gene to undergo auto-induction upon exposure to chemical stimuli as a mechanism to enhance metabolism (Zanger and Klein et al. 2013. PubMed ID: 23467454). For instance, long-term dosing with efavirenz has been shown to lead to increased expression CYP2B6 in certain tissues (zu Schwabedissen et al. 2012. PubMed ID: 22588604; Ngaimisi et al. 2010. PubMed ID: 20881953). The magnitude of auto-induction can be influenced by polymorphisms in CYP2B6 with high levels of auto-induction seen in individuals with wild type or intermediate metabolizer phenotypes and little or no auto-induction seen in individuals with poor metabolizer phenotypes (Desta et al. 2019. PubMed ID: 31006110; Ngaimisi et al. 2010. PubMed ID: 20881953). In addition to efavirenz treatment, exposure to other drugs as well as the insect repellent DEET (N,N-diethyl-meta-toluamide) can also auto-induce CYP2B6 expression (Das et al. 2008. PubMed ID: 19326769; Zanger and Klein et al. 2013. PubMed ID: 23467454). Ethnicity, gender, and age differences may also lead to variation in CYP2B6 expression (Zanger and Klein et al. 2013. PubMed ID: 23467454).

CYP2B6 is a highly polymorphic gene with ~45 variants catalogued by the Pharmacogene Variation Consortium (PharmVar) (https://www.pharmvar.org/gene/CYP2B6). The vast majority of these variants are missense and are further grouped into 38 different star (*) alleles (i.e. haplotypes) based on known population frequencies (Kalman et al. 2016. PubMed ID: 26479518; Lang et al. 2001. PubMed ID: 11470993; Klein et al. 2005. PubMed ID: 16272958). Many of the star alleles have known functional impacts on the metabolic activity of CYP2B6 leading to non-functional, decreased, normal, or increased enzyme activity (https://www.pharmvar.org/gene/CYP2B6). CYP2B6 variants are inherited in a co-dominant manner and an individual’s CYP2B6 metabolizer status (phenotype) is determined by assessing both CYP2B6 alleles (genotype). For instance, an individual who is an intermediate metabolizer has one normal function allele and one decreased or non-functional allele, while an individual with a poor metabolizer phenotype has two non-functional alleles. Individuals who are ultrarapid metabolizers have two increased function alleles. For additional CYP2B6 genotype-phenotype definitions see Table 1 in Desta et al. 2019. PubMed ID: 31006110. 

There have been ~950 CYP2B6 variants catalogued in the gnomAD population database with ~96% of these variants occurring at an allele frequency of <0.1%. The most common missense polymorphisms in CYP2B6 are c.516G>T (p.Gln172His) and c.785A>G (p.Lys262Arg) which occur at allele frequencies of 10% - 40% in some populations (https://gnomad.broadinstitute.org/gene/ENSG00000197408?dataset=gnomad_r2_1). Together, these variants cause defective splicing of CYP2B6 and are characterized as the non-functional CYP2B6*6 allele. Another common variant is c.983T>C (p.Ile328Thr) (i.e. CYP2B6*18 allele) which is found at an allele frequency of ~7.0% in the African population and prevents the production of functional protein. Structural variants leading to the formation of CYP2B6-CYP2B7 hybrids (i.e. CYP2B6*29 and CYPB6*30), as well as non-coding variants have also been documented (Martis et al. 2013. PubMed ID: 23164804, Rotger et al. 2007. PubMed ID: 17885627). Multiple splice variants leading to alternative spliced CYP2B6 transcripts have been reported; however, the functional impact of many of these variant have yet to be determined (Lamba et al. 2003. PubMed ID: 14551287). De novo single nucleotide variants and copy number variants within CYP2B6 have not been documented, and the functional and clinical significance of rare variants within CYP2B6 has not been studied.

Unlike humans with only 1 functional copy of CYP2B6, mice and other rodents have multiple functional CYP2B genes which are located in a cluster known as the Cyp2abfgs cluster, a region similar to the human CYP2ABFGST cluster. To develop a humanized mouse model for CYP2B6, the CYP2B mouse cluster (Cyp2abfgs null) was first knocked down and human CYP2B6 was inserted (Li et al. 2018. PubMed ID: 29402634). Cyp2abfgs null mice are fertile, viable, and are absent of physical abnormalities with the exception of an increase in liver weight. When treated with insecticide metabolized by CYP2B, null mice are poor metabolizers and prone to neurotoxicity when compared to wild type mice. Cyp2abfgs null mice harboring human CYP2B6 were viable and displayed not apparent phenotype. Treatment of transgene mice with classic CYP2B6 inducers such as phenobarbital and dexamethason lead to increased hepatic expression of CYP2B6. This mouse model can be used to study CYP2B6 variants and their functional impact on metabolism (Li et al. 2018. PubMed ID: 29402634). 

It is difficult to estimate the clinical sensitivity for CYP2B6 testing. Analytical sensitivity should be high as reported pathogenic variants are detectable by sequencing.

This test is performed using Next-Gen sequencing with additional Sanger sequencing as necessary.

This test provides full coverage of all coding exons of the CYP2B6 gene plus 10 bases of flanking noncoding DNA in all available transcripts along with other non-coding regions in which pathogenic variants have been identified at PreventionGenetics or reported elsewhere. We define coverage as ≥20X NGS reads or Sanger sequencing. PGnome panels typically provide slightly increased coverage over the PGxome equivalent. PGnome sequencing panels have the added benefit of additional analysis and reporting of deep intronic regions (where applicable).

Dependent on the sequencing backbone selected for this testing, discounted reflex testing to any other similar backbone-based test is available (i.e., PGxome panel to whole PGxome; PGnome panel to whole PGnome).