Interview : Dr John Cashman of HBRI in San Diego

helpful links :
CYP
CYP 3A4
DME's
FMO
FMO3With the interviewee's permission, we have managed to compile a question and answer interview from a recent email dialogue about the enzyme Flavin containing monooxygenase isoform 3 (FMO3) with one of the leading experts in this group of enzymes (as well as expertise in many other human enzymes), Dr John Cashman of the Human Biomolecular Research Institute in San Diego. The dialogue was to try and understand more about FMO3 since sub-normal FMO3 is responsible for genetic trimethylaminuria. Dr Cashman has a long history in TMAU/FMO3 research, especially in the genetic field.
Dr John Cashman medical papers on TMAU/FMO3

We thank Dr Cashman for being so helpful to the TMAU community, and for his long association in research that is directly related to genetic TMAU.

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Is FMO3 one of the most widely used enzymes we have ?
Based on the publications in the literature, I think FMO is underrecognized. It is present in adult liver to a great extent, almost 60% of the amount for the major CYP enzyme (CYP3A4) that does most human drug metabolism currently listed in the literature.

Is there a rule of thumb that we can use to know when something in the diet is likely an FMO3 substrate ? (eg. greenfoods, foods with sulfides, etc) It sounds like almost all vegetables, herbs, spices and proteins potentially.
Yes, FMO prefers smaller nucleophilic compounds: Sulfur-, Nitrogen-, Selenium-, and Phosphorous-containing compounds. But not all that contain these heteratom-containing compounds are nucleophilic as the atoms can be in aromatic rings and non-nucleophilic.

Are all (or many) sulfides, amines, and phosphates FMO3 substrates ?
These substrates need to be nucleophilic. Not all sulfides etc are nucleophilic. Phosphines not phosphates, and organo selenium compounds. But the enzyme substrate binding region has some size requirements or limitations.

Do you expect people with FMO3 deficiency could have problems with FMO3 substrates other than TMA ? (including endogenous)
Yes, I think there could be adverse drug reactions as the amines are not N-oxygenated, for example and are not rapidly cleared. There are many drugs that are amines but not enough research has gone into determining if any of these amine drugs are substrates for FMO. My suspicion is that many more than are currently recognized are substrates for FMO.

Is FMO3 the only likely Drug Metabolizing Enzyme (DME) associated with smells ?
I am not sure about this because other DMEs including CYP metabolize amines, sulfides and phosphines.

Can we survive with zero FMO3 function with no obvious problems apart from TMAU?
I am not sure that the experiment has been done in terms of making a FMO3 knock out mouse and looking at the consequences of this. My suspicion is that FMO3 is involved in some important mammalian developmental function that we don't recognize currently.

CYP3A4 seems to be inhibited by a wide range of chemicals which I presume may be constantly updated. We know that indoles inhibit FMO3. Do you think it is likely that more FMO3 inhibitors that are common in the diet are likely to be discovered?
FMO evolved to metabolize plant-derived materials. This was important early on as evolving mammals needed to protect themselves from all the nucleophilic materials in plants. Yes, I suspect there are a few more nucleophiles that avoided this process but I think most are detoxicated (N-oxides and S-oxides are metabolites). The problem may come in as reductases also evolved to retro reduce N-oxides and S-oxides. These may be more efficient and be the source of the problem. Another important issue is that sometimes FMO produces a reactive metabolite that does not inhibit FMO but leaves the FMO enzyme and inhibits CYP.

Endogenous compounds are compounds created in the systemic circulation. Are there any endogenous compounds that are good FMO substrates ? e.g. hormones or neurotransmitters or blood pressure regulators.
We have published that tyramine and phenylethylamine are FMO substrates. We looked at the other major ones and they do not appear to be substrates: Serotonin, Dopamine, Norepinephrine, Histidine apparently are not substrates for FMO3.

Is FMO present in the gut ?
Yes, FMO is in the gut. I think FMO1 and FMO5 more so than FMO3 but this is in a review with Jun Zhang we published years ago.
see: http://www.ncbi.nlm.nih.gov/pubmed/16402899

Looking at the list of inhibitors for CYP 3A4 on wikipedia (eg citrus, echinacea, milk thistle), It looks as if flavonoids are a particular problem for CYP enzymes? I wonder if these compounds could inhibit FMO3 ?
Probably not. But not sure on this one. Most of the CYP (inhibitors ?) in citrus are polycyclic flavonoids. No nucleophilic atoms are present. So it is unlikely non-nucleophiles are inhibitors. There is a report out there that caffeine is a substrate for FMO3. This is wrong. Not an inhibitor either. No nucleophilic centers. There are a number of azoles (ketoconazole) and imidazoles (cimetidine) that inhibit/alternate substrates for FMO3 but these are from nucleophilic groups in the molecule.

It has been hypothesized that if someone has a bad reaction to a drug it is more likely due to a DME enzyme deficiency rather than an allergy. What are your thoughts on this ?
I am not sure of this one. I think most reported adverse drug-drug interactions (DDIs) occur when there is an excess of a drug compared to normal clearance and reasonable therapeutic levels. There is a DOSE-dependence. Often with allergic reactions, dose is not that important and small concentrations can illicit an immune response. Most DDIs reported are related to liver metabolism.

Pepper is a spice that has anecdotally caused bad reactions such as nightmares.
Pepper inhibits gut metabolism and permits things to be more efficiently taken up through the gut. I dont believe this has been examined as a substrate or inhibitor of FMO.

Could there be an FMO connection with intolerance of garlic ?
Lots of people don’t do very well with garlic including me. My father loved it. There are numerous sulfides in garlic. But this is likely complicated because some of these compounds exist as more complex precursor compounds and intolerance may be related to other enzyme systems.

A lot of people on the forums have as one of their many various odors a smell of burning rubber. Could this be a phosphorous compound ?
I think the compound smelled in burning rubber is a sulfur-compound based.

Quite a few in the group feel they have an allergy to penicillin. If this was rather to do with the DME's, could it be FMO3 deficiency ?
I think penicillin is a direct-acting allergen that is probably normally moped up but if someone doesn’t have adequate protein to protect then it reacts with key proteins that illicit an immune response. That is why it may be idiosyncratic. I don’t think it has much to do with DMEs or metabolic bioactivation. However, reaction to penicillin may indicate a more sensitive immune system.

Is smoking bad for someone with a FMO3 deficiency ?
Nicotine is an FMO3 substrate. It is only N-oxygenated about 5% of a dose but it is a detox pathway. Defective FMO3 will cause individuals to be more susceptible to nicotine. It may be that people with defective FMO3 may be more sensitive to the properties of nicotine.

Would it be impossible to follow a ‘low FMO3 substrate’ diet?
Due to the ubiquitous nature of amines and sulfides in the diet, it may be difficult to have a diet free of FMO3 substrates. However, one can try to avoid choline, for example and this may help a great deal. However, it is important to note that choline is essential in development and child-bearing age women need to consume adequate choline if they are contemplating pregnancy.

What do you think the results of this dissertation imply? http://digitalcommons.library.tmc.edu/dissertations/AAI1450285/
I did not read the dissertation but I did read the Abstract. I agree, more work needs to be done especially the role of non-exon mutations of FMO3.

Many people say they smell of many various smells systemically. We wondered if there is a possible FMO3 connection.
Yes, individuals reporting TMAu symptoms report various smells and this has been confirmed by other experts.

We wondered if the following were good FMO3 substrates ?
hydrogen sulfide: probably not
methanethiol: yes
dimethylsulfide: yes
dimethyldisulfide: yes. I believe it is a very good substrate.
ammonia: probably not
mercaptans: Yes, depending on the structure
Any others you wish to mention : Phosphines.

We wondered if the following things mainly in the diet or endogenously are FMO3 substrates ?
amines in diet
sulfurs in diet
phosphorous in the diet
selenium supplement
FMO3 substrates created endogenously
FMO3 substrates created by the gut flora (aside from trimethylamine)
methyls
It is difficult to answer a broad question like this without some specific examples.

There was a recent paper about FMO3 induction. http://www.ncbi.nlm.nih.gov/pubmed/20570689
Do you think it has any potential as a treatment ?
Not 3-MC. This also induces lots of other things and leads to liver cancer induction. I think induction FMO3 is not the real solution to TMAu. And note, in the report, the functional enzyme activity was not "induced" as much as the RNA.

It’s been suggested that Ataluren may be worth trialling for those with FMO3 nonsense mutations http://www.ptcbio.com/3.1.1_genetic_disorders.aspx. Do you think this is an option for investigation and what percentage with TMAU1 have nonsense mutations ?
Very very few individuals with TMAu in our analyses have nonsense mutations. I don’t think this is worth the risk.

A DME expert suggested ingestion of synthetic FMO3 may be theoretically possible but thought the side effects may make it unfeasible. What do you think of this idea ?
We are looking into this. There are many hurdles in this business. I am not sure about side effects of FMO3.

Is there any other ideas you have as potential treatments ?
I have lots of ideas but no money or time to pursue them.

MEBO RESEARCH STAFF

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