Genetics

New Marker Additions to GPL-SNP1000 DNA Sequencing Profile

BY: MATTHEW PRATT-HYATT, PHD

The number one goal for The Great Plains Laboratory is to provide the best quality results to our clients.  Our GPL-SNP1000 DNA Sequencing Profile has proven to be a great tool in helping provide personalized healthcare to our clients.  The nine pathways we analyze include: methylation, mental health, oxalate metabolism, drug and environmental metabolism/detoxification, gluten sensitivity, cholesterol metabolism, autism risk genes, and transporter gene.  These are crucial biological pathways, which are at the root of many chronic health conditions.  We are now announcing the addition of nine new markers to our already incredibly comprehensive genetic test:

Dopamine Beta Hydroxylase (DBH)
This is an enzyme that catalyzes the oxidation hydroxylation of dopamine to norepinephrine.  DBH can be inhibited by phenolic compounds including those produced by Clostridium species as well as certain organophosphate herbicides and pesticides.  There are two SNPs that can cause decreased activity of DBH.  These are rs2007153 and rs2283123.  These polymorphisms can lead to an increase in dopamine levels and a deficiency in norepinephrine.  Mental health disorders can result because of the imbalance of dopamine and norepinephrine.  Common symptoms can include depression and anxiety.

Paroxonase 1 (PON1)
This is an important enzyme in the metabolism and elimination of many organophosphorus insecticides (PMID: 13032041) and is located mainly in the liver.  PON1 is important in the reduction of atherosclerosis because of its involvement in the protection of high and low density lipoproteins from oxidation.  Individuals with polymorphisms to PON1 are more susceptible to heart disease (PMID: 8675673).  There are two known polymorphisms that can decrease the activity of PON1 and make the individual more susceptible to pesticide exposure, which are Q192R (rs662) and L55M (rs854560).

Hemochromatosis Protein (HFE)
The hemochromatosis gene HFE (high iron) codes for the HFE protein.   This protein is important for regulating the uptake of circulating iron.  This is done by regulating the interaction between transferrin receptor with transferrin.  SNPs to this gene can cause hemochromatosis, a disorder in which the body loads excess iron, which is autosomal recessive.  This means the patient normally needs two bad copies of the gene in order to exhibit symptoms.  There are three SNPs that can lead to hemochromatosis, rs1800562, rs1800730, and rs1799945.  Patients that are homozygous positive for this SNP should have their iron level measured. 

Vitamin K Epoxide Reductase Complex Subunit 1(VKORC1)
This is an enzyme that is necessary for the reduction of vitamin K 2,3-epoxide to its active form, which is important for clotting.  This enzyme is the primary target for the drug warfarin (Coumadin).  The three SNPs that are associated with warfarin sensitivity are rs9923231 (VKORC1*2), rs9934438, and rs8050894.  These polymorphisms can be used in conjuncture with the genotype of Cyp2C9 in order to accurately dose warfarin.

Tryptophan Hydroxylase 2 (TPH2)
This enzyme catalyzes the first and rate-limiting step in the biosynthesis of serotonin. Mutations to this enzyme have been associated with numerous psychiatric diseases including depression, OCD, bipolar disorder, and suicidal behavior.

Major Histocompatibility Complex DQA1 and DQA8
Patients with SNPs to HLA DQA1 and DQA8 have a higher risk of celiac disease.   The HLA-DQA1 and DQA8 are human leukocyte antigen serotype (also called major histocompatibility complex II). The role of this peptide is to present proteins on the surface of cells for identification purposes. This particular serotype presents proteins belonging to a foreign invader on the cells the macrophages, B cells, and dendritic cells in order to activate the helper T cells of the immune system. Proper presentation is critical for immune system activation against pathogens and may possibly be a mediator of autoimmunity.

UDP Glucosyltransferase 1A1 and 1A8 (UGT1A1 and UGT1A8)
These enzymes are important members of the glucuronidation phase II detoxification pathway.  These enzymes catalyze the addition of a glycosyl group from a nucleotide sugar to a small hydrophobic molecule.  The addition of glycosyl groups results in these molecules becoming more water-soluble and easier to excrete. Some of the target molecules for these enzymes include bilirubin, drugs, hormones, and steroids.

Your Body’s Detoxification Pathways

Welcome back to the GPL-BLOG.  Over the past several weeks we’ve been discussing a lot of the environmental toxins that everyone is exposed to on a daily basis.  These toxins must be processed and detoxified.  Most of this is done in the liver through several different processes that include Cytochrome P450 (P450) biotransformation, glutathione conjugation, enzyme hydrolyzing, sulfation, and glucuronidation.

Detoxification is often referred to as a two stage process (phase 1 and phase 2) of metabolism (Figure 1).  Phase 1 metabolism involves the reduction or hydrolysis of the compound (usually caused by the addition of an oxygen molecule).  The addition of oxygen to a compound is referred to as oxidation.  This process is usually performed by the P450 enzymes.

Figure 1

The P450s are a family of enzymes that are found in numerous tissues throughout the body. However, a majority of these are found in the liver.  The P450s are important for the detoxification of many foreign substances, including environmental toxicants and medications.  The P450s are also important in controlling the levels of different molecules produced in the body such as the synthesis and breakdown of hormones, steroids, and multiple other molecules. 

In humans, 58 different P450s have been discovered.  However, only a subset of these is involved in the degradation of xenobiotics (chemicals that come from outside the body).  These enzymes have different substrates, which are determined by the activity pocket of each enzyme.   In regards to detoxification the most important P450s are Cyp1A2, Cyp2A6, Cyp2C9, Cyp2C19, Cyp2D6, Cyp2E1, and Cyp3A4.  Besides detoxification, these enzymes metabolize a majority of medications (figure 2).

Here are some important detoxification enzymes:

Figure 2

Cyp1A2 is important for the metabolism of polycyclic aromatic hydrocarbons (PAHs), which are found in cigarette smoke.  Other substrates include medications, aflatoxin B1, caffeine, and acetaminophen.  The major polymorphism is Cyp1A2*1K, which results in a decrease of activity.

Cyp2A6 is involved in the metabolism of nicotine.  Cyp2A6 is also involved in the metabolism of medications.  The major polymorphic alleles are Cyp2A6*4 and Cyp2A6*9 (which can have between 35% -70% activity depending on if you have one or two polymorphic copies). 

Cyp2C9 is involved with the metabolism of a large number of medications including NSAIDs, warfarin, and tamoxifen.  There are multiple polymorphisms that affect activity of the enzyme. 

Cyp2C19 is involved with the metabolism of multiple medications.  The most common are diazepam, omeprazole, and sertraline.  Cyp2c19 also metabolizes progesterone.   There are two major variants that result in loss of activity.  These are Cyp2C19*2 and Cyp2C19*3.

Cyp2D6 is involved with the metabolism of about 20% of drugs on the market.  It also metabolizes serotonin and neurosteroids.  There are five different polymorphisms that can lead to decreased activity.  Some of the classes of drugs that are metabolized by Cyp2D6 are antidepressants, SSRIs, opioids, and antipsychotics. 

Cyp2E1 is involved with the detoxification of many industrial pollutants, as well as carcinogens.  Cyp2e1 also metabolizes ethanol to acetaldehyde and acetate.  Cyp2e1 is also responsible for bioactivating a number of carcinogens, including cigarette smoke. 

Cyp3A4 is responsible for metabolizing more compounds than most other P450s.  It is responsible for metabolizing sex hormones, caffeine, statins, SSRIs, antifungals, antidepressants, and many other medications.  Some antibiotics can negatively affect its activity. Also, grapefruit and pomegranate juice have been shown to be potent inhibitors. 

Sulfur transferase is a phase 2 enzyme that adds sulfur groups to compounds in order to make them more water soluble and less reactive.  This process is used on a wide variety of toxic molecules including phenols, amines, acetaminophen, and food dyes.  Many chemicals that are able to become airborne are sulfated.  Patients with autism have been found to have impaired sulfation ability, which will make these individuals more sensitive to toxins.

Glutathione transferase is a phase 2 enzyme that catalyzes the conjugation of glutathione to substrates.  The addition of glutathione to toxins prevents these compounds from interacting with proteins in the body and allows them to be excreted via urine or bile.  There are a wide variety of compounds that are conjugated with glutathione.  A partial list includes pesticides, herbicides, carcinogens, acetaminophen, and mycotoxins.   

Glucuronosyltransferase (UGT) is another phase 2 enzyme that is responsible for the glucuronidation of many different toxic chemicals.  This process involves the addition of a glucuronosyl group to substrate molecules making them more polar and more easily excreted by the kidneys. 

Paraoxonase 1 (PON1) is an enzyme that is able to perform paraoxonase activity on substrates.  This enzyme is able to hydroylse and detoxify many different types of organophophate molecules.  PON1 is one of the major pathways that protects people from these types of compounds.  Mutations to PON1 could lead someone to be more sensitive to pesticides.  Infants do not have a lot of PON1 activity.  PON1 becomes active between birth and seven years of age. 

These are the major pathways that you should be aware of when you are thinking about detoxification.  Please see Table 1 to help you understand which pathway is mostly responsible for detoxifying these common toxicants.  Also see Figure 1 to help you understand what you can do to help support type 1 and type 2 detoxification pathways.  Detoxification of compounds by glutathione can be assisted by the supplementation of additional glutathione.  Next week I will discuss some additional methods to help with detoxification.  

Email gplblog@gpl4u.com if you have any questions about this blog post.

The Importance of Genetic Testing for Mental Health

Two weeks ago I talked about how we are starting to move into the age of personalized medicine.  Our goal at The Great Plains Laboratory is to determine what factors lead to chronic illnesses and what treatments may help.  Previously I discussed how the Organic Acids Test (OAT) and the GPL-SNP1000 DNA Sequencing Profile work well together to determine both the risk for and causes of many chronic illnesses and how useful these two tests are in helping practitioners develop individualized treatments.  Last week I specifically discussed the DNA methylation pathway, of the GPL-SNP1000.  This week I am going to go over the mental health pathway of the this same test.  I will talk about what genes are most important, some specific polymorphisms to be aware of, how the OAT can help diagnosis these disorders, and what treatments seem to work best. 

The two most important genes in this pathway are MAO-A (monoamine oxidase A) and COMT (catechol-o-methyltransferase) (Please see Figure 1). Both enzymes are required for metabolizing neurotransmitters critical to mental health. Mutations to either one can have serious consequences relating to how we think, feel, and interpret the world around us.

Figure 1

MAO-A
Monoamine oxidase A is important for the metabolism (breakdown) of biogenic amines such as the neurotransmitters dopamine, norepinephrine, and serotonin (figure 2).  Mutations to this gene have also been linked to depression, borderline personality disorder, and bipolar disorder. 

 

Figure 2

 

There are two different types of polymorphisms involved with MAO-A.  The first type, which is characterized by rs6323 , causes an increase in activity of the enzyme. Rapid metabolism resulting from this mutation causes depletion in the neurotransmitter. This depletion is directly related to the extent of the enzyme activity which can be determined on the Organic Acid test (OAT) by measuring the production of Homovanillic acid (HVA) and Vanillylmandelic acid (VMA) which are the end products of dopamine and epinephrine metabolism (see figure 1).  Patients who are rapid metabolizers will show an increase of HVA of VMA (Figure 3).  Such patients will have a depletion of these neurotransmitters which leads to multiple neurological diseases including depression. Treatment for these polymorphisms includes supporting the methylation pathway which helps to promote the cofactor required for neurotransmitter synthesis and providing neurotransmitter precursors such as tyrosine, B6 and 5-HTP.

Figure 3

A second type of polymorphism to MAO-A is one that decreases activity (rs72554632,  Gln296X), whichcauses a premature stop codon in the gene..  This polymorphismwill lead to a buildup of neurotransmitters, which can lead to the development of Brunner syndrome.  Brunner syndrome is characterized by a decrease of mental capabilities, increased impulsivity, mood swings, and sleep disorders. These individuals will have a decrease of HVA and VMA metabolites on the OAT. Without the benefit of genetic testing, these results could lead a practitioner to treat with neurotransmitter precursors. Increasing neurotransmitters for individuals with this type of mutation would actually exacerbate the condition. Treatments include decreasing the amine containing foods such as fish, cheese, and fruit, taking progesterone to increase MAO; taking inositol(which reduces 5HT2A serotonin receptor), taking riboflavin (increases MAO activity), or taking ginkgo, which has been shown to decrease aggressiveness in MAO-A deficient patients.

COMT
Catechol-O-methltransferase (COMT) is present in the body in two different forms.  The short form is called soluble catechol-O-methyltransferase (S-COMT).  The longer form is called membrane-bound catechol-O-methyltransferase (MB-COMT).   MB-COMT is mainly present in the nerves of the brain, while S-COMT is located in the liver, kidney, and blood.  In the brain, MB-COMT is responsible for degrading neurotransmitters called catecholamines, which include dopamine, epinephrine, and norepinephrine. Therefore the membrane bound form is believed to have a greater affect on mental health, though both forms may be implicated in disease since both are capable of metabolizing catecholamines.

GPL-SNP1000 analyzes six different SNPs for COMT.   Of these, the most extensively studied is a mutation to rs4680 (Val158Met). Despite the many studies available about this mutation, the scientific community is still at odds about the degree to which this mutation causes disease. What is known is that individuals with this mutation have lower activity of the enzyme.  Mutations to COMT will lead to dopamine and epinephrine not being broken down properly, which can be detected by a decrease in the HVA metabolite in the Organic Acids Test.

Figure 4

Interestingly, some individuals may benefit from increased dopamine while others may benefit from less. Dopamine functions differently depending on the area of the brain it is produced in and there are likely other mutation to this gene may also influence its function and expression. These interactions are still being investigated and are why GPL is looking beyond the most common mutation to help inform results.  Conditions associated with these mutations include OCD, depression, and schizophrenia.  Symptomatic patients may benefit from treatment aimed at promoting the enzyme function so that dopamine and epinephrine can be metabolized. This is a SAM dependant enzyme and methylation support can be helpful in patients with this mutation.

APOE
Apolipoprotein E (APOE) combines with lipids in the body to form lipoproteins.  Lipoproteins are responsible for packaging cholesterol and other lipids and carrying them through the bloodstream.  Like so many genes in the human body, this is not its only function. The ApoE protein is involved with more than was originally thought. In this case, it has a role in immune expression, cognitive function, and telomere regulation.  There are four different gene versions of APOE called e1 (the double mutant), e2 (rs7412), e3, and e4 (rs429358).  The most common version is e3, the most detrimental is e4

 Mutations to APOE can lead to an increased risk of developing Alzheimer’s disease. People who inherit even one copy of the APOE e4 allele have an increased risk of developing the disease. Certain mutations to this protein impair the ability of the body to phosphorylate NMDA (glutamate) receptors causing a reduction in activity (Glutamate receptors have been implicated in a number of neuropsychiatric disorders including depression, biopolar disorder, schizophrenia, and Autism (PMID: 12404584, 21315104). While this gene is being extensively studied for its role in Alzheimer’s disease, vulnerable patients may also be predisposed to other psychiatric disorders. (PMID: 25751510, 15557508). Patients with certain ApoE mutations may develop cognitive symptoms well before onset of Alzheimer’s disease (http://psycnet.apa.org/journals/neu/16/2/254/).  Patients with these polymorphisms should look for early signs of the development of Alzheimer’s disease.  Treatment with estrogen has demonstrated some encouraging results.

DAOA
The D-amino acid oxidase activator (DAOA) protein (also known as G72) is a 153 amino acid protein that localizes in the brain, spinal cord, and testis.  This protein is located in the endoplasmic reticulum and the mitochondria cellular compartments.   DAOA is a modulator of D-amino acid oxidase (DAO) activity.  If DAO is hyper activated it can result in a decrease in the D-serine level and hypo function of the NMDA receptor.  Overexpression of DAOA has been found in schizophrenics and those suffering from bipolar disorder.  Mutations to DAOA have been linked to a higher incidence of schizophrenia and bipolar disorder. 

It is important to remember that our genes influence our mental health and well being but do not necessarily determine the eventual outcome. There are countless factors that play an equal role in how we perceive the world including diet and lifestyle.   I believe that understanding the most common mutations and their function is very helpful in diagnosing, treating, and preventing different neurological disorders.  Using the GPL-SNP1000 test in combination with the OAT can further help determine the extent to which our genes are being expressed metabolically. The combination of these two tests can help bring about the best chance to achieve emotional health and physical well being.

Email gplblog@gpl4u.com if you have any questions about this blog post.

 

Genetic Testing and Organic Acids Testing: A Dynamic Duo of Diagnostics

Today I have two words for you:  Personalized medicine.  What does this phrase mean to you?  When I think of personalized medicine I think of treatments that are custom designed for each individual patient, and I believe this is the ultimate goal for all of us in the field of functional medicine.  To make this happen, we will have to work together as a team - the healthcare practitioner, the lab, and the patient.  If we do so effectively, the result should be better health and improved lives of our patients.  The only way to get there is to design a treatment plan that addresses the underlying cause(s) of our patient’s ailments and not try to just suppress the symptoms. 

When I talk to both practitioners and patients, they often ask “Where do we start?” or “What is your most important test?”, and until recently I would have always said that the Organic Acids Test (OAT) is the obvious place to start.  The reason for this is that the OAT provides more information than any other test.  The OAT gives us a metabolic snapshot of multiple pathways in the body, offering insight into possible underlying causes of symptoms, as well as what kind of nutritional support is needed.  However, now the OAT by itself is no longer the obvious choice.  I am now recommending the OAT + GPL-SNP1000 combo because these two tests, one metabolic and one genetic, work so well together.   Today I would like to share some of the markers in each of these tests that work really well in tandem.    The primary pathways where we see overlap between the two tests are methylation, mental health, detoxification, and oxalate metabolism. 

The first pathway that GPL-SNP1000 covers is the DNA methylation pathway, also called the MTHFR pathway.  This pathway is a process by which carbons are added onto folic acid from amino acid and redistributed onto other compounds throughout the body.  This process is responsible for the formation of methionine, S-Adenosyl methionine (SAMe), and thymidylate monophosphate (dTMP).  These compounds are then used in neurotransmitter metabolism, detoxification, nucleotide synthesis, and multiple other processes.  I can’t say enough about how important neurotransmitter metabolism and detoxification of chemicals are to everyone’s health.  We have so many patients for whom the majority of their symptoms result from the upset of these two processes.  Since the methylation pathway is so important we decided to make it a high priority in our new genetic test.   GPL-SNP1000 looks at 105 different methylation SNPs (single-nucleotide polymorphisms).   Next week I plan on going more in-depth on the methylation pathway and how GPL-SNP1000 can be useful.

So what markers in the OAT are important for patients with MTHFR mutations?  The first one we have is vitamin B12.  We evaluate B12 levels by measuring the amount of methylmalonic acid (see Figure 1).  B12 is an important cofactor for many of these methylation enzymes.  The second important marker is pyridoxic acid, which is a form of vitamin B6. I have counted over 50 enzymes that require B6 in the body.  It is an important cofactor in the methylation pathway.  It is directly involved with the function of CBS enzyme and indirectly involved with MTHFR, BHMT, and SHMT.  Another marker involved with the MTHFR pathway is uracil.  Having an elevated uracil level can be indicative of folate pathway malfunction.

The next pathway that is helpful to analyze in both the OAT and GPL-SNP1000 is the mental health pathway, which involves the synthesis and breakdown of neurotransmitters in the brain.  The combination of measuring the neurotransmitter metabolites and knowing if the enzymes involved are functional will help guide us to the best treatment options.  GPL-SNP 1000 covers 14 different mental health genes, which I will cover next week (I’m trying not to make these blog posts too long).  Three of the best markers in the OAT for measuring neurotransmitter metabolism are homovanillic acid (a dopamine metabolite), vanilymandelic acid (epinephrine/norepinephrine), and 5-HIAA (serotonin, marker).  These markers are the metabolites of the neurotransmitters by the enzymes MAOA and COMT (see Figure 2), the genes for which are analyzed in GPL-SNP1000.  Deficiencies in these enzymes due to faulty SNPs  may cause low neurotransmitter levels, which may also be caused by low amounts of precursors, cofactors, or increased inhibitors which is why information from both the OAT and GPL-SNP1000 is so incredibly useful.   

The third pathway that I will briefly touch on today is the detoxification pathway, and specifically for glutathione (GSH).  Detoxification is so important in today’s industrial, polluted, and toxic world.  Every day we are inundated by hundreds of chemicals.  We are exposed to many through the environment and some by choice (like medications).  Our bodies have to process these chemicals in some way.  GPL-SNP1000 looks at dozens of genes that are important for detoxification.  A good marker in the OAT for how well the body is detoxifying is pyroglutamic acid.  Elevated values of pyroglutamic acid are indicative of glutathione deficiency due to excessive toxic exposure or a genetic issue. 

The final pathway I’m going to discuss today is oxalate metabolism.  Oxalates are crystalline molecules that we absorb from our diet (high oxalate foods) or are produced by an infection, like yeast/fungal overgrowth.  These oxalates can accumulate in the body and cause inflammation.  The symptoms of oxalate accumulation include pain, nephrolithiasis, and neurological symptoms. Oxalates are known to cause/create kidney stones.  Children with autism who exhibit eye-poking behavior have been shown to have a build-up of oxalates behind their eyes, causing tremendous pain, and thus the eye-poking.  GPL-SNP1000 covers five different genes involved with the production and elimination of oxalates.  The OAT has three oxalate markers:  glyceric, glycolic, and oxalic acids. (Figure 3)  In addition, low B6 and increased yeast or fungal markers are associated with increased oxalates. 

I think that is all I’ll cover today.  In the future I will cover the methylation pathway and the neurotransmitter pathway a little more in-depth.  If there is another pathway you want me to cover in greater detail, please let me know.  I want to be a part of your healthcare team as we all work together for the better well-being of our patients.

Email gplblog@gpl4u.com if you have any questions about this blog post.