Why do MDMA and 4-FA both release 5-HT? And methamphetamine only releases 1/10 of 5HT

[serotonin2A]

I just recently (thanks to adder) learned that the methylenedioxy ring in MDA is quite out of plane of the benzene ring. What does the indanyl version look like then? Somehow my gut feeling tells that the difference can't be drastic enough to warrant such difference in binding affinity. I would still go with the explanation that the oxygens' electron pairs and/or negative charge have something to do with it.

X-ray crystallographic studies have actually shown that the methylenedioxy ring in MDMA is in the same plane as the benzene ring:

http://onlinelibrary.wiley.com/doi/10.1107/S0108270197012390/abstract

http://www.sciencedirect.com/science/article/pii/S1093326307001908

MM2 energy minimizations on MDA and MDMA confirm that the benzodioxole rings should be in plane.

 

[aced126]

X-ray crystallographic studies have actually shown that the methylenedioxy ring in MDMA is in the same plane as the benzene ring:

http://onlinelibrary.wiley.com/doi/10.1107/S0108270197012390/abstract

http://www.sciencedirect.com/science/article/pii/S1093326307001908

MM2 energy minimizations on MDA and MDMA confirm that the benzodioxole rings should be in plane.

So this suggests the system is aromatic? The molecule doesn't have to necessarily bind in its energy minimised conformation though? There probably wouldn't be a big degree of freedom of movement in the methylenedioxy carbon, however, wouldn't the molecule be able to change it's conformation elsewhere especially if the binding interactions it gains from its unfavourable conformation lower its energy to below that of the energy minimised conformation that it exists in freely?

 

[serotonin2A]

So this suggests the system is aromatic? The molecule doesn't have to necessarily bind in its energy minimised conformation though? There probably wouldn't be a big degree of freedom of movement in the methylenedioxy carbon, however, wouldn't the molecule be able to change it's conformation elsewhere especially if the binding interactions it gains from its unfavourable conformation lower its energy to below that of the energy minimised conformation that it exists in freely?

I don't think it implies it is aromatic -- there just doesn't seem to be a great deal of comformational flexibity in the methylenedioxy bridge. Ligands don't have to bind in their minimal energy conformation, but they often do. In this case, though, it didn't seem like that part of the molecule is likely to change conformation. Can anyone point to some data showing that part of the molecule adopts an out-of-plane conformation?

 

[belligerent drunk]

I don't think it implies it is aromatic -- there just doesn't seem to be a great deal of comformational flexibity in the methylenedioxy bridge. Ligands don't have to bind in their minimal energy conformation, but they often do. In this case, though, it didn't seem like that part of the molecule is likely to change conformation. Can anyone point to some data showing that part of the molecule adopts an out-of-plane conformation?

http://www.sciencedirect.com/science/article/pii/S0009261410003830

It isn't about MDA/MDMA but 1,3-benzodioxole, shouldn't make a difference in this aspect though. I didn't read the whole thing because I don't understand some parts of it, but what I gathered is that the most favorable conformation is somewhere between "puckering" and "flapping" (using terms from the paper) which kind of makes sense to me. The methylenedioxy ring is not conjugated with the other ring as far as I can tell, I'm not sure if oxygens' lone pairs do interact with the aromatic ring though; in any case, there's little stabilization to be found from it being in plane with the other ring in my opinion.

 

[serotonin2A]

http://www.sciencedirect.com/science/article/pii/S0009261410003830

It isn't about MDA/MDMA but 1,3-benzodioxole, shouldn't make a difference in this aspect though. I didn't read the whole thing because I don't understand some parts of it, but what I gathered is that the most favorable conformation is somewhere between "puckering" and "flapping" (using terms from the paper) which kind of makes sense to me. The methylenedioxy ring is not conjugated with the other ring as far as I can tell, I'm not sure if oxygens' lone pairs do interact with the aromatic ring though; in any case, there's little stabilization to be found from it being in plane with the other Unlessring in my opinion.

Unless I'm missing something, didn't the puckering and flapping primarily happen in an excited state? Is there a good reason to think that would be relevant to MDMA bound to the transporter?

 

[belligerent drunk]

Unless I'm missing something, didn't the puckering and flapping primarily happen in an excited state? Is there a good reason to think that would be relevant to MDMA bound to the transporter?

As far as I understood the excited state is more out of plane than the ground state, but the ground state is also out of plane to some degree. I don't know about transporter binding, I was just commenting on the conformations. I will re-read the paper later today, maybe you're right and I misunderstood something.

 

[Zeds(NCO2CH2CH3)2]

Sequence homology varies among gpcrs very widely actually, and although some characteristics between the SAR of 5HT NA and DA receptors and their drugs appear identical, or very similar, they are all not the exact same. Not at all.

On the topic of the 3,4 positions, they primarily recruit a double serine on one of the transmembrane helices or two. That's the simplest answer i can give and the strength of the interaction relies on secondary characteristics of this portion of the ring (van Der walls interactions, pi pi stacking) besides that crucial ser interaction. Go look at crystal strong structures.

They aren't all directly correlative remember.

Zeds

 

[serotonin2A]

As far as I understood the excited state is more out of plane than the ground state, but the ground state is also out of plane to some degree. I don't know about transporter binding, I was just commenting on the conformations. I will re-read the paper later today, maybe you're right and I misunderstood something.

The ground state may be distorted out of plane, but how much distortion are we really talking about? The "twist, state, one of the three distorted states, looks relatively planar. Maybe MDMA has to adopt that conformation before it binds. Contrast those states with the very non-planar form of IAP. It still seems like IAP is more out of plane than MDMA.

 

[belligerent drunk]

Could you show me what IAP looks like? After a quick search I couldn't find anything. I think you're right - the ground state most likely isn't strongly out of plane if at all. In that case it would make sense that IAP doesn't bind well while methylenedioxy, 4-fluoro- and methoxy variants do, because of steric effects. I guess one way to test this hypothesis would be to compare 4-methyl and 4-ethyl or isopropyl amphetamines. What do you think?

 

[aced126]

I thought IAP was a good 5-HT releaser (82nM) but didn't really release DA or NE very well at all.

 

[aced126]

No, it wouldn't. As adder wrote, compare the water solubility of ethanol and ether. Alkoxy groups are widely known to be lipophilic, but are capable of accepting a hydrogen bond.

Adder, I used to assume that a H-bond was involved in the potency of MDMA, PMA, etc but the modeling/docking studies I have seen haven't identified a hydrogen bond donor in the vicinity. That also wouldn't explain the potency of 4-methylamphetamine. That still leaves unanswered the question of why you can't replace both of the oxygens in the MDMA methylenedioxy ring with carbons and retain potency at SERT but that might be a stereochemical issue.

Yeah, ether prefers lipophilic solvents (LogP=1.00) a lot more than ethanol does (logP=-0.59 ). But if you replace the oxygen atom in ether with carbon you get pentane, a compound which prefers liphophilic substituents almost 400 times more than ether (pentane logP=3.59).

http://www.rsc.org/suppdata/nj/b3/b303016d/b303016d.pdf


So if lipophilic substituents were needed at the 4 position, then comparitively the indanyl derivative would have much more favourable interactions than MDA.

 

[aced126]

Adder, I used to assume that a H-bond was involved in the potency of MDMA, PMA, etc but the modeling/docking studies I have seen haven't identified a hydrogen bond donor in the vicinity.

On the topic of the 3,4 positions, they primarily recruit a double serine on one of the transmembrane helices or two. That's the simplest answer i can give and the strength of the interaction relies on secondary characteristics of this portion of the ring (van Der walls interactions, pi pi stacking) besides that crucial ser interaction. Go look at crystal strong structures.

Is there a H bond or not?

http://onlinelibrary.wiley.com/doi/10.1002/minf.201300013/pdf

In this article it shows there being a serine residue in DAT around the phenyl part of the ring, but in SERT there are only lipophilic residues, being leucine, isoleucine and valine. In SERT there is a threonine residue closer to the aliphatic part of the molecule rather than the ring substituents, but the article suggests it could donate a hydrogen bond to the ring substituents.

 

[serotonin2A]

Is there a H bond or not?

http://onlinelibrary.wiley.com/doi/10.1002/minf.201300013/pdf

In this article it shows there being a serine residue in DAT around the phenyl part of the ring, but in SERT there are only lipophilic residues, being leucine, isoleucine and valine. In SERT there is a threonine residue closer to the aliphatic part of the molecule rather than the ring substituents, but the article suggests it could donate a hydrogen bond to the ring substituents.

You have to be a little suspicious of findings like that because if there is a H-bond formed with Thr then they should have been able to find evidence for the interaction. But from the figure, the Thr looks like it was aligned with the opposite side of the phenyl ring. It could be that MDMA adopts a different binding pose and they never got around to docking it. But either way, they never actually presented evidence that a H-bond forms.

Anyone reading this want to take the time to energy minimize MDMA and then dock it?

 

[Rio Fantastic]

All three compounds act in the same basic manner but the big difference is their selectivity. (+)-METH releases DA and NE with EC50 of 10-20 nM, but the concentration required to release 5-HT is almost 2-orders of magnitude higher. So the effects on DA and NE max out before METH can alter 5-HT.

By contrast, MDMA is slighly more potent a releaser of 5-HT compared with DA and NE. So it will tend to release all three with some preference for 5-HT.

4-fluoro-AMPH is closer to METH in it's selectivity, but compared to METH it is much less selective for DA vs 5-HT. So at doses that release DA it will also have moderate effects on 5-HT release.

There is no difference in the structure of VMAT of TAAR1 in different neurons.

No offence intended here. In this subforum, often the topics you are talking are absolutely fascinating, but me certainly and most likely others can be turned away from it by the impenetrable and dense use of technical terms that must be deciphered by the layman before they can grasp the post. Have you considered just putting a brief explanation of some of the terms you use, or at least not relying on acronyms too much? What's an 'EC50'? What's 'nM'? What is VMAT and TAAAR1? Again no offence intended here, and it would be totally understandable if you refused to elaborate, but I figure it'd make this part of the forum more accessible, which I think is key to harm reduction. If people understand the effects of drugs a little better, especially the dangers & risks, the harm they cause will be reduced.

 

[serotonin2A]

Yeah, ether prefers lipophilic solvents (LogP=1.00) a lot more than ethanol does (logP=-0.59 ). But if you replace the oxygen atom in ether with carbon you get pentane, a compound which prefers liphophilic substituents almost 400 times more than ether (pentane logP=3.59).

http://www.rsc.org/suppdata/nj/b3/b303016d/b303016d.pdf


So if lipophilic substituents were needed at the 4 position, then comparitively the indanyl derivative would have much more favourable interactions than MDA.

I actually broight up this issue in some of my earlier posts (see above). Indanyl is very non-planar compared to benzodioxole. The indanyl ring may not be flat enough to bind with high affinity.

But it is important to note that IAP is a potent 5-HT releaser. It just isn't as potent a 5-HT releaser as MDMA. It's action is still consistant with lipophilicity being involved.

 

[aced126]

I actually broight up this issue in some of my earlier posts (see above). Indanyl is very non-planar compared to benzodioxole. The indanyl ring may not be flat enough to bind with high affinity.

But it is important to note that IAP is a potent 5-HT releaser. It just isn't as potent a 5-HT releaser as MDMA. It's action is still consistant with lipophilicity being involved.

Looking at that picture of fenfluramine in SERT, it doesn't look as though the active site is really strict on small geometry changes in IAP. The carbons directly substituted on the benzene would have to be planar, so the maximum possible angle that final carbon #2 can be depressed by is 109.5 degrees. That is the maximum and my gut instinct tells me it is more likely to be depressed an angle of 30-90 degrees. If you do have a source for the angle of the 2 carbon in IAP I'd like to have a look at it. Even if this was so, I think the displacement caused to that #2 carbon would not be a problem for SERT accommodating it (I don't know if the bond lengths in the picture are relatively coherent with each other, but I'd presume they would be). In fact it seems like you could add another 1 and a half benzenes and it would still fit.


On another note, where is the basic nitrogen binding to? I would've thought it would bind to aspartate or something but it appears that either the positive nitrogen is binding with T439 or Y176 in SERT and Y156 in DAT.

 

[aced126]

No offence intended here. In this subforum, often the topics you are talking are absolutely fascinating, but me certainly and most likely others can be turned away from it by the impenetrable and dense use of technical terms that must be deciphered by the layman before they can grasp the post. Have you considered just putting a brief explanation of some of the terms you use, or at least not relying on acronyms too much? What's an 'EC50'? What's 'nM'? What is VMAT and TAAAR1? Again no offence intended here, and it would be totally understandable if you refused to elaborate, but I figure it'd make this part of the forum more accessible, which I think is key to harm reduction. If people understand the effects of drugs a little better, especially the dangers & risks, the harm they cause will be reduced.

To be fair you could look up quite a lot of the stuff you listed on wikipedia. It gets strenuous to keep on saying vesicular monoamine transporter over and over again so it gets abbreviated. EC50 is the concentration of a compound required to produce 50% of the maximal response. nM is concentration of a drug in nanomoles (1 nanomole = 10^-9 moles). VMAT is a transporter protein which transports neurotransmitters into vesicles which eventually fuse with the cell membrane and release the neurotransmitters into the synapse. TAAR1 is a recently discovered protein that is thought to be involved in the reversal of the neurotransmitter transporters on the cell membrane.

https://en.wikipedia.org/wiki/EC50
https://en.wikipedia.org/wiki/Vesicular_monoamine_transporter
https://en.wikipedia.org/wiki/TAAR1#Function

 

[serotonin2A]

Looking at that picture of fenfluramine in SERT, it doesn't look as though the active site is really strict on small geometry changes in IAP. The carbons directly substituted on the benzene would have to be planar, so the maximum possible angle that final carbon #2 can be depressed by is 109.5 degrees. That is the maximum and my gut instinct tells me it is more likely to be depressed an angle of 30-90 degrees. If you do have a source for the angle of the 2 carbon in IAP I'd like to have a look at it. Even if this was so, I think the displacement caused to that #2 carbon would not be a problem for SERT accommodating it (I don't know if the bond lengths in the picture are relatively coherent with each other, but I'd presume they would be). In fact it seems like you could add another 1 and a half benzenes and it would still fit.


On another note, where is the basic nitrogen binding to? I would've thought it would bind to aspartate or something but it appears that either the positive nitrogen is binding with T439 or Y176 in SERT and Y156 in DAT.

Nichols did some SAR work regarding the methylenedioxy bridge of MDMA and is seems like that region of the binding site is sterically hindered. There is a drastic drop in potency when you add steric bulk the the benzodioxole ring:

http://www.ncbi.nlm.nih.gov/pubmed/2623014

Note that illustrations like the figure you are talking about are usually prepared so that they show interactions of interest, with many other features of the binding site omitted for clarity. You won't be able to get a sense of how crowded the binding cavity is from a figure like that. Their illustration makes it look like the binding cavity is huge, but it is likely that many neighboring side-chains were cut-off. Those residues weren't interacting with the CF3 in fenfluramine, but they might sterically hinder a bulkier substituent.

It is common to find that amines form a salt bridge with a highly conserved Asp in GPCRs, but the transporters are not from the GPCR family and have a different structure.

 

[aced126]

Nichols did some SAR work regarding the methylenedioxy bridge of MDMA and is seems like that region of the binding site is sterically hindered. There is a drastic drop in potency when you add steric bulk the the benzodioxole ring:

It is common to find that amines form a salt bridge with a highly conserved Asp in GPCRs, but the transporters are not from the GPCR family and have a different structure.

Ok so in that link about bulkier substituents, would the methyl group sticking out of the methylenedioxy ring in ethylidenedioxy-MDA be flat? Isopropylidenedioxy definitely cannot be, and so if the former statement is true then that is strong evidence that a depression in bond angle will hinder activity. Similarly napthylaminopropane is potent at SERT (3.4nM), and also at DAT (12.6nM) and NET (11.6nM) as well.

This has been made (https://en.wikipedia.org/wiki/DiFMDA) and is inbetween MDA and MDMA for SERT affinity; the fluorine radius is small and so it should not be too sterically bulky, but it should technically be very slightly less lipophilic.

However if the molecules below were found to have greatly reduced SERT affinity, that would confirm the sterical tightness in the binding cleft, even though the area of the molecule in question would be a lot more lipophilic.

So in that diagram what actually is binding to the nitrogen; doesn't seem like the nitrogen is binding to anything negative. I thought that a salt bridge would be a crucial bond seeing as a basic nitrogen is crucial for activity (maybe the nitrogen lone pair is interacting with an electrophile although I suspect if this was the case the bond formed would be too strong to dissociate from the electrophile once the molecule is exposed to the cytosol. Has the figure omitted this binding? If not, why couldn't the nitrogen be replaced with say an ether bridge. If that does somehow get into the neuron, I suspect it would not be able to reverse the transporter and only very slightly release the monoamines from their vesicles; the only activity it would have is maybe slight blockade of reuptake.

What is D98 and D79 in the figure? Is the sodium ion only present in SERT and what is it bound to? Is there any place I can get SERT bound to a ligand without residues and other stuff omitted?

Wouldn't these molecules be of very high affinity to SERT. They are quite lipophilic and are planar.

And what about these 2 aromatics? The nitrogen isn't even basic so it should be more lipophilic than if the lone pair of the nitrogen was localised. The n-demethylated 5 isomer is apparently a thing but I couldn't find any binding data.

 

[aced126]

Another oddity which does not seem to fit with the data. Amphetamine and phenmetrazine are known monoamine releasers, with high selectivity for DA and NE. Para fluorine substitution on amphetamine (4-FA) drastically increases SERT affinity. Yet, meta fluorine substitution doesn't increase SERT affinity as expected, but instead slightly reduces it compared to amphetamine. Both phenmetrazine and 3-F-phenmetrazine are very selective for DA and NE but the former compound (ec50 SERT ~8000nM) experiences a 3-4 fold increase from this substitution (3FPM ec50 SERT = 2500nM). What can explain these observations, especially the amphetamine substitution one. SERT tolerates an oxygen meta to the ring (MDMA, 6-APB). Somehow the fluorine is creating slightly negative interactions in 3-FA where as in 4-FA it seems to be binding strongly. The article does suggest a hydrogen bond between one of the fluorines in fenfluramine and the threonine residue, and although the distance will be shorter in fenfluramine, if this is so surely there should be a weaker H-bond in 3-FA and thus would slightly increase affinity rather than decrease it. Any insights are appreciated.