Prevention and treatment of some congenital defects of neurological development

J. LEJEUNE (*)

Prenatal diagnosis and surgical treatment of congenital malformations. p105-111. Edited by : Antonio Pachi, Alessandro Calistri, Giovanni Astrei. (*) Transcription from tape recording.


Sommaire

I am supposed to give you in thirty minutes an idea about what we can do to fight against the most peculiar human diseases, because only humans really feel and suffer from it, and also the most inhuman, because it prevents the affected persons from using the most important quality which is due to our genetic patrimony that is the full power of thinking.

It seems as arduous to speak about mental retardation, generally, because it seems to be only a symptom, because a terrible area of catastrophies can produce it: from infection, to distructions, to haemorrages, to hydraulic compression during hydrocephaly and every possibility of distruction of the brain, has as a consequence a mental deficiency, no matter whether it is due to an accident, to a bus, to a virus, to a gene, or a chromosome.

Nervertheless it happens if we try to look, generally speaking, at how our brain is built and functioning, that eventually we can put some kind of order among this terrific area of catastrophies. As you probably remember, when Blaise Pascal, for the first time, was able to build a computing machine, he demonstrated that same day that it is possible to include some logic if matter is given a proper form. Indeed I'm not going to tell you that the machines that we are building nowadays, the little computer that you can have in your pocket, are thinking, because to think that machine do think would be thinking mechanically, but they teach us that all the engines we have manufactured in order of simulating part of the property of our own mind answer to three pre-requisites: first a logical network pre-established, entirely logical by construction; second that there is an insulating system somewhere so that transmission inside the machine can be made without spreading of the signals, and just sending one signal from one point to another point. And third the components of the machine have to follow the basic law of logic, the law of "yes" or "no". And all the gates that the electronicians are using are very similar to the well-known "porte" of Alfred De Musset "II faut qu'une porte soit ouverte ou fermee" (A door must be either shut or open).

It's true in engineering if the door remains half open no reasoning is any longer possible and curiously if we take those three simple inferences from the technology of the pseudothinking machine, we can put together most of the data we have on the causes of mental retardation.

It is possible that the building of the network has not been entirely satisfactory, like for example in trisomy 13. In arhinencefalia the rhinencephalus has not been built. And if we were a man repairing computers we would see that the whole rack of the electronics is not there and we would understand immediately why the computer has not the full power: that is the same thing in neurology. But it happens even that the whole network is not built at all and specially in anencephalia which is a particular case of neural tube defect. If you remember that embriologically the first neural crest has suddenly made a little half-tube and healing at the middle and it continues like a zeep to the end, to the tail and to the head, and if this does not close at the tail you have spina-bifida, whereas if it does not close at the head you have no head whatsoever.

And this is very instructive, that due to the recent discovery of Smittles and confirmed by the statistics of Florence; it seems that the prevention, I mean the true prevention of the diseases, not the prevention of the birth of the babies, can be achieved by very simple trick which is giving to the mother a lot of vitamins especially of folic acid. I would not venture very much on that because I have been told by our President that someone else will speak about it, but just it is interesting to note at the beginning that folic acid has something to do with the building of the brain. I'll come back to that in a moment.

The second thing is that to answer the distance transmission we are obliged to have insulating substances around the wiring and indeed we have myelinisation of all the little filaments inside our brain and around the nerves and this myelinisation is very interesting because we know again from embriology, an early development of neurology of the new-born, that the steps in which part of the brain get finally myelinisated are just the same as the occurrence of new neurological development in the development of the child; if one part of the brain has not finished myelinisation the function related to it does not function yet.

Finally the synapses are in fact the gates of the electronicians I was talking about, but you remember that those gates are very much more complicated than those of the electronicians. You remember that at the end of an axon there is a little bottom which hooks itself on the membrane of the next neuron but this does not hook directly, there is an interstice between and in that cleft the cell releases some chemical mediators which will come on the sensitive membrane of the next cell, and immediately open ion channels which will engulf ions, one by one, exactly like a little devil of Maxwell. Our brain is counting particles one by one much better than any Geiger counter and in that way sends the message from one cell to another one. Now, if we put together those three things we will come to a very simple constatation: to build the brain you have to, beside building special neurons and nervous fibres inside, you have to make those insulating substances which essentially are phospholipids. It means that for each molecule of insulating substance you need to use three methyl group, that is broadly speaking three mono-carbons groups.

Now, if we want to build the security-key, the chemical mediators which transmit the signal from one cell to another cell, again you have to use three methyl group if you want to build acetylcholine. But curiously even if you want to inhibit one of those keys that you have already used once in one synapse you have again to methylate it with catecolamine methylate. And then it happens that if we want to build a brain the greatest demand on raw material will be on the smallest stone in which we build molecules which is monocarbon. The reason it is not only used to make the key of security which makes functioning the synapses, not only the myelinisation, the insulating substance to wrap around the nerves but also to build the cell themselves, because to manufacture the D.N.A. or R.N.A. you have to manifacture purine and pyrimidines, and to make one molecule of purine you need two monocarbons and also to make a molecule of timine you need one monocarbon.

Altogether very simply: those molecules, of monocarbons, are surely the most important to build a brain over all the other ordinary supplies. Hence I would propose to you the general hypothesis, which I know is purely a hypothesis that part and I would say a great part of mental retardation is possibly related to a trouble of the metabolism of the monocarbons. Now I had many reasons to say that because nature has built in our own brain a very peculiar pump and this pump is able to take selectively folic acid, I have already spoken about it, out of the serum, and in our brain the amount of these vitamins is always four times what it is in any other part of the body, and that pump is so efficient that even in a severe carence of this vitamin the depletion becomes very severe in the liver, in the bone marrow ( and then you get agranulocitic anemia) and that before the level in the brain is touched. I cannot believe that it has been made just by chance, just to be interesting for a biochemist, and the other thing which is very simple and known many years ago, that any time you have an anemic syndrome, due to folic acid deficiency, to B12 deficiency, you have very severe neuro-anemic symptons and they can range from neuritis to very severe psychiatric illness. That cannot be by chance either.

The aminoacids can give one atom of carbon which will be later carried by folic acid on the reduced form which is called tetra-hydropholate. Various types of monocarbons are carried by folic acid.

Several synthesis are made with them; there are reactions coming from 5-10 methylen, from 5-formile-tetrahydropholate and from 5-10 methylen-tetrahydropholate and all these three are used to build purines and that is the limiting step to produce D.N.A. and R.N.A. basic stones, which have to be pre-sent in the cells so that the cells can reproduce its genetic message.

But here there is a change in the possibility, here we are dealing with R.N.A. and D.N.A. precursors, so to speak, and here the transfer of a methyl allows through the omo-cystine-methionine system to build myeline, to build chemical mediators, and even to build another one which is carnitine, which is a very important trimethylate molecule.

The result is that, if monocarbons are the most tiny stones which are used in the building, and it is the functioning of our brain, then we could have diseases at different parts of this pathways, for example if it is on the precursors it will be just a strike of the mining industries, raw material is not provided and then the brain is not built. And it is known that different blocks from Hystadine, from aminolevulinate, produce mental deficiency.

Now, if you look at the transfer, folic acid is in fact the transportation system from the raw material to the local factories which are building the nucleus and the neurons. Now, if there is a strike of the transportation corporation either because you don't have enough folic acid, or you don't have enough B12 vitamin, then the raw material will not be transported, and we know very severe mental deficiency and neuropatic diseases which are related to blockage inside the folic acid metabolism or inside the B12 metabolism.

Now the blockage can be later, it can be on the omocystine-methionin system, and you know that omocystinuric children are feeble-minded, but it can be also in the recuperation system. Betine is a recuperation from used acetylcholine, or from used choline, and this recuperation, if it is not good, causes a lost of the monocarbons and again we know diseases in which this is blocked and it gives very severe mental retardation.

Now, this is a general hypothesis which fits with most of the genetic diseases known to produce mental retardation by one biochemical step block. What about chromosomes?

You have heard this morning that there is a special form of mental deficiency in which children look quite normal, but have a very broad spectrum of mental retardation; some of them are very severely affected, some of them reach an i.q. of 70 or 80, most of them are around 40 to 50. Their fades are not specific, only you can remark when they are very small babies, that their skin around the fingers is very floppy. It's very characteristic, but a very minor symptom, that is there, not always, but very often; it is only when they are grown up, after puberty, that they will develop big testicles if they are males, and this is one of the most typical sexlinked mental retardation.

If you look at the chromosomes, you will recognize by the banding pattern that one of the chromosomes, in these cases, has something wrong at its end, it's a gap observed on the X chromosome.

Now, the interesting phenomenon is that all the people of the male sex who do show that fragility are mentally retarded, but not all the cells do show it, only a percentage of them, between 5 to 10 percent. It happens that because it is a sex linked disease, the mothers of the affected children or the affected boys are carrier of this chromosome, but generally they do rather well with it. Some of them are border-line but most of them are normally minded. But if you look at their chromosomes carefully, you will see that around one or two per cent of their cells do show this fragility at this point of the X chromosome.

Now it was demonstrated, that if folic acid was added to the culture medium those affected people who have this fragility were apparently cured in the sense that if folic acid was added to the growing medium this fragility was not detected.

Well, we took over this remarkable achievement and because of all I have said about monocarbon metabolism I tried the precursors, that is aminoacid which produces a raw material which is carried by folic acid, and again, if you add extra-precursors to the culture, you do cure the fragility. The contrary was obvious we should put the transportation system in strike: it is very easy, we have aminoptiorine, methotrexate, which are antifolic drug, because they block the idrofolate reductase (they are used against cancer), and we can block, we can have a strike inside the cells of the transportation, and that immediately increases the frequency of the gap. Then we demonstrated that with antitransportation we increase the fragility but that we decrease it with a precursor, the raw material, and with an excess of transporters; then obviously it is a disease related to some to one of the stepsin the metabolism of the monocarbons.

Now, where it is, we do not know yet, but because we have demonstrated that in vitro, and because the precursors are non-toxic, and because folic acid is non-toxic, we could use it freely on the children. And the result was extremely interesting, because after few months of treatment with one milligram per chilogram of body weight of folic acid per os all of the treated ones showed an almost disappearance of the fragility of their X-chromosome. That was the first demonstration. The second was, and it was expected, I would say it was hoped but not really expected, there was a remarkable improvement in the mental functioning of part of those children. Those of them who were not only mentally retarded but were having behavioural troubles proved to be much easier to manage, and parents, educators, observers, psychometricians agreed that something had happened.

There was the first time in the history of medicine that we were confronted with disease, in which we know part of the biochemistry, in which we know a molecular abnormality on one particular chromosome and in which a very simple application of one vitamin was together alleviating the chromosomal symptoms and the mental symptoms.

Now, you have to know where we are. We are not very far. We have treated around forty patients actually but I have data only on 22 which have received and taken the treatment for more than six months. The results are that on those 22, five of them have really improved in a way no person would have expected: parents, educators or else. Six have improved but not very impressively, eleven have not changed, but no-one has deteriorated. Then we are sure that we are not doing any harm and that some of them do respond very surprisingly.

Now, the most curious fact is that we treated also the mothers or the healthy sisters, but only those of them who had some mental difficulty, and the result was much more brilliant because those women or those girls said themselves that they feel much better, they walk much better, they are able to go to the school, two of them are at the High School, and the mothers are better in their job and obviously they feel something and they refuse to stop.

For the moment it could seem that I am over enthusiastic and that I am describing improvements, without giving i.q. data, without giving hardcore numbers; it just because it is not possible, because we have no good measure of behavioural troubles and of improvement of them, but we had, we come across, of a terrible counter-demonstration. It was a year ago, it was a child, one year and half old, he was diagnosed because he was very floppy; he was treated for two months, one milligram folic acid per kilogram of body weight, At first, before the treatment he could not sit at all, he was one year and half, he could not sit at all, the eye was very dull, very empty, he was not talking, and not interested in the surrounding. After two months he was entirely changed, he was sitting himself, remaining seated for the whole day, began to crawl on the floor and was smiling, responding, playing and we were really very enthusiastic. Two months later he came back to the consultation, he had received correct treatment and he was again terribly floppy, terribly disinterested and every improvement was entirely lost. And because we are between doctors here, I can make a confession, I was so much distressed by this terrible break-down that day I forgot about to take the blood of the child to see what were happening to the chromosomal gap. And I ordered to continue the treatment but I was wondering whether we were not intoxicating him. And during three days it was a very difficult life in the laboratory. But at the third day, my daughter-in-law who was finishing her medical studies and had come to my lab for six months, just to look at chromosome and to look at human genetics, she asked me about: "Maybe he has received something else?", and in fact he had. He had been treated by his family doctor with Bactrim, which contains Trimethoprim and Trimethoprim is a potent inhibitor of the hydrofolate reductase.

We would never have dreamed to try to make the control of our treatment using an antibacteric which can block the use of the folic acid and in fact, solved about it, I would have said: you should never do it, but it had been done and then we did a systematic study and stop on that and we showed on more than 97 patients non-affected by fragile X that Trimethoprim does not produce fragility in people who have not fragility, but we demonstrated also in 40 patients affect by the fragile X syndrome, that if you use the antibacteric Trimethoprim in vitro, you multiply the frequency of the gap by four or five: in children having ten percent you go up to forty percent. That proves that it is not impossible to understand some of the basis of some of the mental retardation. For this little story I would just tell you that the same day we published the first results on the use of folic acid in vivo, another paper was published in another journal, it was the first case of a fragile X child recognized in utero and killed by abortion. It is not a coincidence. It is an apologue of the actual stage of affairs. In front of a disease we can often get so desperate that after doing the diagnosis we discard the affected pretending that we are discarding the disease. In other ways once the diagnosis is made, we have to find why this particular disease does produce what it does, and we have to fight against the disease and not against the patient.