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BH4 - GCH1 Question

Discussion in 'Genetic Testing and SNPs' started by Mimi, Jan 24, 2015.

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Do you have a GCH1 mutation?

  1. Yes

    20 vote(s)
    76.9%
  2. No

    6 vote(s)
    23.1%
  1. Valentijn

    Valentijn Senior Member

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    I found this part of the study to be rather odd. First of all, saying that a single SNP can act as a surrogate for an entire haplogroup completely undermines the existence of the haplogroup. If it is a sufficient substitute, they haplogroup shouldn't have been necessary in the first place, as a single SNP in it would have worked just as well.

    The other thing is that D values in the 80's aren't very impressive. That means that their conclusion is wrong approximately 12% of the time.

    And finally, if their single SNP acts a satisfactory substitute for the haplogroup, they should be able to prove that via follow-up research directly correlating their SNP with symptoms (or better yet, actual gene function). Any idea if they've done that yet? It's been 7 years, so I'd think it's looking like a dead end thus far.

    None of this is looking convincing thus far.
     
  2. nandixon

    nandixon Senior Member

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    @Valentijn

    We'll definitely have to disagree on this one.

    The first reference (from 2014) that I gave in post #20 above (ETA: and also post #35) effectively reaffirms how close the relationship is between rs841 and gs224:

    GTP Cyclohydrolase I Gene Polymorphisms Are Associated with Endothelial Dysfunction and Oxidative Stress in Patients with Type 2 Diabetes Mellitus

    There are others as well.

    I'll take a 9 out of 10 linkage disequilibrium predictive value anytime for these purposes. It might be too much work for you, but it might be interesting to do one of your patient versus control charts - using only the patients and controls who have European Y- and mt-haplogroup ancestry, of course, and using only the 4 SNPs (rs841 and the three for gs224) that have been identified as significant in European ethnicities.
     
    Last edited: May 6, 2015
  3. Valentijn

    Valentijn Senior Member

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    The entire concept as a single SNP standing in for a haplogroup is completely invalid. Especially in the case of gs224 where there are three SNPs which are themselves not completely linked. rs841 could only stand in for the haplogroup if all three SNPS were in very strong linkage disequilbrium, in which case there would not be a haplogroup in the first place: there would be a SNP which has a correlation, and some other SNPs linked tightly enough to be said to have or echo the same non-additive impact.

    Of the 58 patients+controls that I have full data for all four SNPs, 39 are homozygous non-CAT haplotypes (GG-TT-CC), with the corresponding rs841 genotype (GG). The 2 remaining people with rs841 GG are non-CAT, but with mixed genotypes (GG-AT-CT).

    1 person has rs841 AA but only a single-CAT haplogroup (CC-AA-CT).

    16 people have rs841 AG. 13 of those have a likely (but uncertain) single-CAT haplogroup (CG-AT-CT), and 1 has a certain but un-linked single-CAT haplogroup (CG-AA-TT). 2 with rs841 AG have no actual CAT haplogroup (CG-AT-CC).

    So rs841 isn't failing to indicate any CAT haplogroups, but in 3 out of 17 instances where rs841 indicates single- or double-CAT, it is wrong. So that's a "false positive" rate of 17.6%. Which is pretty high, as is the 12% rate found in the study. The significant findings regarding the original haplogroup would almost certainly not hold up if the study was replicated using rs841 to look for the same correlations.
     
  4. nandixon

    nandixon Senior Member

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    Rs841 might actually be part of the same haplotype that gs224 comprises, so it doesn't make sense to me to make that argument.

    It also sounds like you're thinking that all the SNPs within a haplotype have to be found to be in absolute perfect linkage disequilibrium in order for the haplotype to be valid or useful for scientific or research purposes, which isn't true (especially not in a gene that has as high a mutation rate as GCH1 does).

    I already mentioned previously, in post #40, how the rs841/gs224 relationship was likely to play out in real life, due an inconsistent LD between rs841 and rs8007267, and I gave a method to deal with the problem:

    Looking at it quickly, using that method, and with the actual examples I gave (in post #40), it appears to yield perfect results for the ability of gs224 to correctly predict rs841 for your 58 patients and controls.

    So I definitely think people (with European ethnicity) can feel comfortable using gs224 to get a very good idea of their status for rs841. And again, this is (potentially) useful just because in some scientific papers rs841 and gs224 aren't mentioned together, and because 23andMe no longer tests for rs841.
     
    Last edited: May 7, 2015
  5. Valentijn

    Valentijn Senior Member

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    If that were the case, it still couldn't substitute for the haplogroup. Each member of the haplogroup should add additional value, or it shouldn't be in the haplogroup at all.
    Actually it's exactly the opposite. If they're in complete linkage disequilibrium, there is no haplogroup in the first place, and the entire association can be correlated to any single SNP in the supposed haplogroup. Haplogroups exist because of the variety of its members. Equating a single SNP with the entire haplogroup is ignoring the principle of variation, and basically negating the entire haplogroup.

    But the original research into gs224 says that the haplogroup is necessary to find the correlation, therefore no single SNP in that haplogroup was sufficient by itself. And a single SNP like rs841 really cannot adequately reflect that small, but important, level of variation. As shown in the research you quoted, and the data which I have, it is a very rough approximation - just as some of the haplogroup members themselves might be in isolation.

    In the process of making that approximation between the haplogroup and a single SNP, they are inevitably entering the realm of statistically insignificance regarding the original correlation between the haplogroup and its phenotypical traits. It's going from a somewhat weak but significant association with the initial haplogroup into the realm of no significant association at all. Trying to make something out of insignificant associations (beyond "hey, lets run a bigger trial") is a really bad idea, and inherently unscientific.
     
  6. shoponl

    shoponl

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    Are oxidized-LDL and Myeloperoxidase (MPO) elevated with NO/ONOO problems? I would love to know conclusively whether high peroxynitrite levels result from gs224 and somewhat-low BH4.

    Life Extension is having their annual blood-test sale now through June 1; the ox-LDL test is $56, the ox-LDL panel (ox-LDL + Myeloperoxidase) is $131. These prices are almost half off their normal prices.

    basic = http://www.lifeextension.com/Vitamins-Supplements/itemLC817472/Oxidized-LDL
    panel = http://www.lifeextension.com/Vitamins-Supplements/itemLC100034/Oxidized-LDL-Panel

    They also offer an advanced panel (including "F-2 Isoprostanes" as an indicator of oxidative stress) but that one is $214. http://www.lifeextension.com/Vitamins-Supplements/itemLC100035/Advanced-Oxidized-LDL-Panel

    Good idea, or waste of money? What do you think?
    Has anyone else here measured their ox-LDL?
     
    nandixon likes this.
  7. nandixon

    nandixon Senior Member

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    @shoponl

    None of those tests will specifically tell you that there's a peroxynitrite problem (whether due to either low BH4, or to a low BH4 to BH2 ratio).

    If a test(s) came back positive, you can say that there's probably an undesirable level of oxidative stress present in your body, but you can't say, specifically, whether that's due to increased peroxynitrite (and you certainly couldn't extrapolate to saying there's a low BH4 problem). It might be, but it might not be.

    If they come back negative, that would mean you're much less likely to have an issue with peroxynitrite. (I'm not sure whether it would rule it out entirely, though.)

    (Note: If a person isn't supplementing any antioxidants when they have a test like these, and they have a negative result, then I think it's going to be unlikely they have ME/CFS, which I think is pretty well known to have high levels of oxidative stress. Oxidative stress is common in many serious diseases, though, so a positive result is not diagnostic for ME/CFS by any means.)

    I haven't seen any commercial testing specifically for BH4 (tetrahydrobiopterin), only for total "biopterin."

    Perhaps you might email one or more of the authors who have done the various GCH1 studies, and see if they have an idea for specific BH4 testing? Good luck!
     
    shoponl likes this.
  8. malk

    malk

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  9. ahmo

    ahmo Senior Member

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    Northcoast NSW, Australia
    nan, what's BH2? Is it related to B2? I just started FMN form of B2 and it's radically shifted my peroxynitrite/oxidative stress issues. As well as histamine/mast cell, and sulfur processing.
     
  10. nandixon

    nandixon Senior Member

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    BH2 is the abbreviation for 7,8-dihydrobiopterin, which is the oxidized form of BH4 (tetrahydrobiopterin). BH2 can be recycled ("reduced") back to BH4 by the enzyme dihydrofolate reductase (DHFR). (This is the same enzyme that the body uses to convert the synthetic/artificial form of folate known as folic acid into a usable form.)

    Note for general information: A very similar molecule is quinoid 6,7-dihydrobiopterin, which should be abbreviated as "q-BH2," but which Amy Yasko unfortunately mistakenly refers to as "BH2" in all of her work. q-BH2 can also be recycled back to BH4, but by a different enzyme - DHPR (not DHFR). The gene for DHPR is QDPR, while the gene for DHFR is DHFR, i.e., the enzyme and gene have the same abbreviation in the latter case.
     
    Last edited: May 21, 2015
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  11. nandixon

    nandixon Senior Member

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    I think @shoponl has already had the neopterin/biopterin test done, if I remember correctly (from the Yahoo GCH1 Discussions forum for "gs224"). That test doesn't give her the information she's looking for, i.e., what her actual BH4 levels are.

    In Antoniades' 2008 paper, they determined actual BH4 by a somewhat complicated series of steps/procedures (described in reference #23 in the quote below), some of which I used to do in the lab myself, and which I doubt very much is available commercially. (Perhaps there may be a simpler process now, though, some 7 years later.):

    GCH1 Haplotype Determines Vascular and Plasma Biopterin Availability in Coronary Artery Disease
     
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  12. Helopilot52

    Helopilot52

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    Hello. I'm a long time lurker and new Member. I realize this thread is a bit old but wanted to start somewhere on my questions about BH-4.

    After suffering from anxiety for a number of years, I've discovered a number of gene mutations using 23andme.com and other test. I've found that I'm CBS699T +, MAO-A + and A1298C + among others, including high amounts of heavy metals, including aluminum. Recent testing showed that I have very depleted BH-4 levels, elevated phenylalanine with low dopamine and serotonin, low GABA, etc. Needless to say, I have a ND/MD developed protocol in place to deal for the anxiety which has helped some, but felt BH-4 supplementation might provide additional support.

    After two months of pleading with both my MD and ND, I finally got a prescription for BH-4.....! I (I strongly suspect the MD or ND did not have a clue what BH-4 is/does). I live in Washington State, just south of Seattle and found a compounding pharmacy that can formulate BH-4 in 2.5MG individual dosages, so I'll be on the BH-4 bandwagon hopefully on Monday June 4. What I'm hoping to learn is the following:

    1) Does BH-4 help with depression and or anxiety?
    2) Dosage amounts that worked for you (I'm a 59 yr. old male, 6'1", 206 #)
    3) Side effects? Some sites complain about "detox" symptoms (?) when starting BH-4 or the need to reduce any SSRI type drugs before starting BH4. (Fortunately I've been off SSRI's for a number of years).
    4) Does BH-4 poop out after a period of time?

    Any other pearls of wisdom would be appreciated. Oh and my Compounding Pharmacy says that their formula has never required refrigeration and that most compouders have switched to non-refrigration formula.

    Thanks again for your help.
     
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  13. Gondwanaland

    Gondwanaland Senior Member

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  14. Stacey1121

    Stacey1121

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    I have several mutations of GCH1


    Now I am the new member and am very curious as to how the BH4 worked for you.

    I have never been diagnosed with PKU but I have several GCH1 mutations (no PAH mutations on 23&me) and extremely elevated phenylalanine (1263+ μmol/L) with very low tyrosine. I am working on Freddd's methylation protocol but there seems to be something underlying it and the elevated phenylalanine might be a big clue. I have 7 hetro and 2 homo GCH1 mutations so that would fit in the puzzle. I am also C677 hetro with a handful of other hetro methylation mutations.

    The methylfolate, B12, l-carnitine fumarate, potassium and r-lipoic acid along with the other nutrients have been helpful. Tyrosine has helped. But it is as if I am building my castle on shaking ground. The high doses of methylfolate (I am up to 20 mg) help but as soon as it wears off (in just a couple of hours - I take 5 mg 4x/day) this underlying foggy fatigue comes right back. Did you have swelling with elevated phenylalanine? Have you modified to a low protein diet? I have a call with my doc in a couple of days but I was hoping to get some feedback re the BH4. My doc is fairly open minded so I am hoping she will be open to letting me give it a try with all of the data we have collected.

    I have always had symptoms of high phenylalanine for as long as I can remember - although I can't say for sure that its always been high as I have never been tested for it until now. But it sure would make a lot of sense. Really appreciate any specifics you might be able to offer. Thanks so much!
     
    Last edited by a moderator: Dec 28, 2015
  15. nandixon

    nandixon Senior Member

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    Can you list the nine GCH1 SNPs you mentioned (by their "rs" numbers and your results for each)?

    With phenylalanine that high, it seems somewhat more likely (just from an odds standpoint) that you might have a mutation on the PAH (phenylalanine hydroxylase) gene that 23andMe is not picking up. I would think you might want to see a PKU specialist, who may want to have that gene fully sequenced. That sort of specialist might be your easiest route to obtaining (large amounts of) BH4 - and getting your insurance to pay for it - if that is determined to be a suitable/necessary treatment for you.
     
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  16. Stacey1121

    Stacey1121

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    Here are my GCHI snp's
    GCH1 rs10131232 A AA +/+
    GCH1 rs11158026 T CT +/-
    GCH1 rs2878169 T GT +/-
    GCH1 rs3783641 A AT +/-
    GCH1 rs3783642 C CC +/+
    GCH1 rs4411417 C CT +/-
    GCH1 rs7147286 A AG +/-
    GCH1 rs752688 T CT +/-
    GCH1 rs8017210 A AG +/-
    These last two supposedly affect upcycling of BH4 as well but are not GCH1
    QDPR rs1031326 T CT +/-
    QDPR rs3796809 A AG +/-

    Here is other info I read last night I thought was interesting...link is at the end....


    In the 1970s, it was discovered that not all HPA was PKU. Some forms of HPA were caused by disorders of synthesis and recycling of the cofactor (tetrahydrobiopterin, or BH4) involved in the Phe hydroxylation reaction. During the 1980s, the human PAH gene was mapped and cloned, and the first pathogenic variants identified. In the 1990s, in vitro expression analysis was being used to study the effects of different PAH alleles on enzyme function and the crystal structure of PAH was elucidated.
    HPA is treatable. Affected individuals can lead normal lives. Continuous efforts are made to improve the taste and convenience of the current synthetic dietary supplements [Rohr et al 2001]. Research to improve the current treatment with restrictive phenylalanine diets, supplemented by medical formula, is ongoing (see Management, Therapies Under Investigation).

    Differential Diagnosis

    Hyperphenylalaninemia (HPA) may also result from the impaired synthesis or recycling of tetrahydrobiopterin (BH4), the cofactor in the phenylalanine, tyrosine, and tryptophan hydroxylation reactions. All of the HPAs caused by BH4 deficiency are inherited in an autosomal recessive manner. They account for approximately 2% of individuals with HPA. BH4 is also involved in catecholamine, serotonin, and nitric oxide biosynthesis (seewww.biopku.org).

    Defects in BH4 synthesis result from guanosine triphosphate cyclohydrolase (GTPCH) deficiency caused by mutation of GCH1 or from 6-pyruvoyl tetrahydrobiopterin synthase (PTPS) deficiency caused by mutation of PTS.
    Impaired recycling of BH4 is caused by dihydropteridine reductase (DHPR) deficiency caused by mutation of QDPR or by pterin-4 acarbinolamine dehydratase (PCBD) deficiency caused by mutation of PCBD1.
    Blau et al [2001] and Scriver & Kaufman [2001] emphasize that all neonates with persistent hyperphenylalaninemia must be screened for the BH4 deficiencies. The following tests are best performed in specialized centers. Prenatal diagnosis is possible for all forms of BH4 deficiencies. The following screening tests are essential:

    Analysis of pterins in the urine which will aid in diagnosis of PTPS deficiency, DHPR, and pyruvate carboxylase deficiency (PCD). The pterin profile may be normal for DHPR, and enzyme activity should be measured in erythrocytes or from a dried blood spot.
    Other disorders of BH4 metabolism including dopamine-responsive dystonia (DRD) and sepiapterin reductase (SR) deficiency do not have elevations in phenylalanine but abnormalities in pterins can be detected in cerebrospinal fluid (CSF).
    Some centers will also perform a loading test with BH4 on babies with a positive newborn screen. This test can rapidly differentiate BH4 deficiencies from PKU.
    If screening test results show abnormal pterins, the following confirmatory tests are recommended:

    Analysis of folates and neurotransmitter metabolites in CSF. This analysis will help to determine the severity of the BH4 defect [Blau 2006].
    Enzyme activity measurements
    Molecular genetic testing. Mutation testing for each of the genes involved in BH4 synthesis and recycling is available for confirmatory diagnosis and is useful for prenatal diagnosis. The genotype/phenotype relationships have not been determined and the above biochemical tests are still important for prognostic classification.
    The typical (severe) forms of GTPCH, PTPS, and DHPR deficiency have the following variable, but common, findings: intellectual disability, convulsions, disturbance of tone and posture, drowsiness, irritability, abnormal movements, recurrent hyperthermia without infections, hypersalivation, and swallowing difficulties. Microcephaly is common in PTPS and DHPR deficiencies. Plasma phenylalanine concentrations can vary from slightly above normal (>120 μmol/L) to as high as 2500 μmol/L. Mild forms of BH4 deficiency have no clinical signs.

    PCD deficiency, sometimes referred to as 'primapterinuria' is associated with benign transient hyperphenylalaninemia.

    In principle, BH4 deficiencies are treatable. Treatment requires the normalization of BH4 availability and of blood Phe concentration and restoration of the BH4-dependent hydroxylation of tyrosine and tryptophan. This is achieved by BH4 supplementation along with dietary modification, neurotransmitter precursor replacement therapy, and supplements of folinic acid in DHPR deficiency. The treatment should be initiated early and probably continued for life [Blau et al 2001,Ponzone et al 2006].

    More information on the BH4 deficiencies can be found atwww.biopku.org.

    http://www.ncbi.nlm.nih.gov/books/NBK1504/

    I do think a full PAH test would be great. How was your response to the BH4 btw?
     
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  17. nandixon

    nandixon Senior Member

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    I don't believe that any of those particular SNP results should be capable of causing the high phenylalanine you're seeing.

    Using small amounts of BH4, between 2.5 and 25mg, made me feel worse. I haven't tried very large amounts of BH4, though, which might have a different effect.

    Note: My understanding is that it doesn't work to supplement intermediate-size amounts of BH4 directly since this results in large amounts of oxidized BH4 (i.e., BH2), and the ratio of BH4 to BH2 is actually as important as the absolute amount of BH4. This is because the enzyme DHFR, which recycles BH2, becomes impaired at lower BH4:BH2 ratios. However, this problem can potentially be overcome by using VERY large amounts of BH4, i.e., several hundred milligrams daily, and this is actually what is often done to treat PKU disorders. The problem is that BH4 is currently very expensive and difficult to obtain without a PKU or related diagnosis.
     
    Last edited: Dec 28, 2015
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  18. Stacey1121

    Stacey1121

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    You seem very knowledgeable - are you a practitioner or researcher? I am an artist so this whole situation is really unfamiliar ground, but I will do whatever it takes to feel better. The reason I ask is that my doctor(s) did not mention the high phenylalanine as a potential problem - its has been me pouring over my tests (blood and genetic) looking for clues because I want to feel better. I also have extremely high unexplained selenium count - and I take nothing at all with selenium in it - its 1.26mcg and the range is .25-.75. I believe the answer (or one of the big answers) as to why I have had such fatigue and body pain has to do with the phenylalanine and selenium counts. Elevations in either can make you very fatigued and foggy - and I have both. The body pain is getting better with the methylation but I know there is something else going on. Side note, I would welcome a well versed referral to consult with if you know of anyone.

    Also, thanks for the info above about BH4. I will request a PAH test tomorrow and keep searching :)
     
  19. nandixon

    nandixon Senior Member

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    @Stacey1121

    I'm a former researcher (medicinal chemistry). You may need to see a PKU specialist, who will likely be a pediatric doctor, to sort things out. He or she should be able to tell if doing the sequencing of PAH (or any other genes) is worthwhile. A regular doctor wouldn't normally order that sort of genetic testing, I don't think.

    I'm not familiar with elevated selenium issues - unless you've been eating a lot of Brazil nuts.
     
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  20. malk

    malk

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    off-topic:
    Do you have a link to the side effects (fatigue) of selenium? Never heard of that. Very interessting.

    I do also have elevated levels (levels increased after starting treatment), as others described it too in another forum.

    The only exlpanation I heared: Selenium bounds with mercury and this compound stays within the cells. When starting successfull detox regime (incl. methylation etc.) the selenium starts to pour out of the cells and is shown in urine tests.

    This makes sense in my case because I had amalgam fillings and I have elevated mercury levels while detox (DMSA).

    But I would be courious if there are other connections/ explanations.

    Btw. I do supplement selenium because it is in my all-in one capsule but nevertheless the selenium was not high before detox. Others do not suplement selenium at all (as you described too) and still have elevated levels since various treatments.
     

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