Trypanosoma Brucei

Author: angela spalatro
Date: 24/02/2010


Sleeping Sickness


The sleeping sickness Human African trypanosomiasis (also known as TAU) is a tropical disease spread only in the African equatorial areas, inside the belt between the 15th north and the 20th south parallel . It's one of the worst African health problems, with AIDS and malaria. This disease is called "Sleeping Sickness" because its terminal stage is defined by apathy, sleepiness and cachexia. During the last phase of this stage, the patient is no longer able to stand up and eat.
This stage causes neuropsychiatric problems, convulsions and serious sleep disorders; sometimes these symptoms may lead to the coma. This sickness results deadly without the appropriate treatments.


The pathogen of TAU is the Trypanosoma brucei flagellate.

Life Cycle


1) The Occidental TAU, caused by the subspecies Tripanosoma brucei gambiense, is found in Occidental and Central Africa, in an area delimitated by Senegal at north, Angola at south and Victoria Lake at east. It's also known as Chronic Sleeping Sickness, because it causes a chronic condition that can extend in a passive phase for months or years before symptoms emerge, and it's transmitted by a fly of the Glossina family, palpalis species.

2) The Oriental TAU is caused by the subspecies Tripanosoma brucei rhodesiense and is spread in the limited area of southern and eastern Africa. It's the acute form of the disease (also known as Subacute Sleeping Sickness) and emerges in a few weeks or months, in some cases 6-7 days are enough; it's transmitted by the Glossina morsitans fly.

This kind of flies are commonly known as "tsé-tsé" fly. When it stings an already infected subject, it becomes infected too and so it can transmit the disease to other subjects.
Every year over 40000 new cases are estimated but over 300000 is a more reliable year valuation; this disease kills almost 66000 people all over the world (OMS data for 2003).
The low interest by the international community about this disease still causes the use of medicines from the second half of the 19th century for the treatments, although their significant secondary consequences and their low therapeutic efficacy, Pentamide, Suramin and Melarsporol for example. In particular, being a toxic organic compound of arsenic, the melarsoprol is a highly dangerous treatment which quickly reacts with sulfhydryl group compounds.

The Therapeutic Approach Problem

The main problem of this disease is that any kind of vaccine is ineffective; the parasite evolved a defensive mechanism against the host immune defense system: the parasite cell capsule is covered with a single kind of protein; when the immune defense system reacts with it (or a vaccine against this antigenic component is synthesized), the parasite changes the protein cell wall so the defense system becomes unable to recognize it. It's the same old "pathogen survival" story: be more clever than the immune defense system!
In this "trypanosoma case", this "escape process" is called "capsule change" and is based on a genetic recombination mechanism similar to the salmonella's one (synthesis of the flagellin genes by the Hin Recombination), but through a different process called "Converting Non-Reciprocal Genetic": the genetic information is moved from a passive to an active cell part. Acting this way, it's possible to codify different capsule glycoproteins in order to evade the immune defense system.
This process recurs thousands times and illustrates the disease case history: the patient has temperature which disappears every time the immune defense system recognizes and responds to the antigen, then the temperature comes out again when the parasite begins the capsule change in order to stop the defense system.

Knowing before Fighting Back the Enemy

What can we try to stop in its metabolism? Why can't we try to inhibit its replication, grown or elusion mechanisms instead of focusing on the pathogen effect?

Trypanothione. The glutathione moieties are shown in black and the polyamine linker in red.

In 1985, Alan Fairlamb discovered a unique thiol compound present in these parasites, and named it trypanothione. This thiol metabolite is quite different from its human equivalent, glutathione. Trypanothione allows the parasites to fend off free radicals and other toxic oxidants produced by the immune system of the infected patient, and was shown to be vital for parasite survival and virulence.
It's unique to the Kinetoplastida and not found in other parasitic protozoa such as Entamoeba histolytica. Trypanothione-dependent enzymes include reductases, peroxidases, glyoxalases and transferases.
Trypanothione-disulfide reductase (TryR, Oxidoreductase class) was the first trypanothione-dependent enzyme to be discovered. It's an NADPH-dependent flavoenzyme that reduces trypanothione disulfide and catalyzes the following reaction:

trypanothione + NADP+ ⇄ trypanothione disulfide + NADPH + H+.

This is the main mechanism used by the parasite to defend itself from the action of the immune defense system and shows why attacking it "from outside" is useless.
Now the question is: how can the parasite synthesize the trypanothione? What can we do to make it vulnerable to the immune defense system action?
A parasite which is quickly growing up (high rate of DNA propagation) needs large amounts of spermine and spermidine for the DNA compact folding, but also to synthesize the trypanothione for its survival.
Where are spermine and spermidine (and so the trypanothione) come from?

As shown in the picture, the cause is the ornithine amino acid from which spermine and spermidine polyamine are synthtisized, essential for the parasite survival.
To remark from the synthesize reaction:

1) the ornithine decarboxylase (ODC) enzyme is used as needed step inside the process,
2) the pyridoxal phosphate cofactor is an essential element for the amino acid decarboxylation reactions and for the enzyme action.

Is stopping this enzyme enough to fight back the parasite?
It’s not so easy, because the enzyme, present and active inside its cells, is at the same time essential for the host cell proliferation: to inhibit it means to inhibit the cell turnover of the healthy organism.
Actually, the ornithine decarboxylase enzyme turnover inside the mammal cells is extremely fast, so it’s possible to quickly replace it when the enzyme has been destroyed. It’s not the same inside the trypanosoma cells and the destroyed enzyme can’t be replaced.
For this reason, the medicines based on this biochemistry explanation result selective on the parasite and innocuous for the host organism!

New Selective Therapies

1) DFMO (difluoromethylornithine)

It’s a suicide enzyme inhibitor for the ornithine decarboxylase which catalyze the ornithine conversion to putrescine (1,4-diaminobutane), which makes the spermine and spermidine polyamine creation happen.
The cysteine 360 is alkylated by the eflornithine which stops the putrescine synthesis. Also the human ODC can be inhibited, but its turnover is very fast so the eflornithine can’t cause dangerous secondary consequences. Further, the eflornithine can be administered either by intravenous injection (but with fast elimination rate), either by oral sedation (circumscribed use because of the low absorption level).
The medicine weakly binds to the plasmatic proteins and breaks the blood-brain barrier, so it’s useful during the initial and final phase of the trypanosomiasis. It’s effective only on the Tripanosoma brucei gambiense.

2) Nifurtimox

Just like the metronidazole, it should reduce and produce ROS; it also is an inhibitor for the trypanothione reductase (a protective enzyme present only inside trypanosomas) causing the trypanothione synthesis inhibition.

What about the good old Melarsoprol?

It derives from the arsenic and causes a lot of secondary consequences, The most dramatic being a reactive encephalopathy (encephalopathic syndrome) which can be fatal (3% to 10%). An increase of resistance to the drug has been observed in several foci particularly in central Africa.
Recently Fairlamb proposed a new inhibition action mechanism of the trypanothione reductase that consists in a reaction with the sh group of trypanothione cisthein. The synergic action of melarsoprol and eflornithine, resulting from a sequential block of the trypanothione synthesis, confirms this theory.

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