6-Diazo-5-oxo-L-norleucine
Encyclopedia
6-Diazo-5-oxo-L-norleucine (DON) is a glutamine
antagonist
, which was isolated originally from Streptomyces
. It is a non-standard amino acid
. The diazo compound
was characterized in 1956 by Henry W Dion et al., who suggested a possible use in cancer therapy. This antitumoral efficacy was confirmed in different animal models. DON was tested as chemotherapeutic agent in different clinical studies, but was never approved. The last clinical results were published in 2008, though not as DON monotherapy but in combination with a recombinant glutaminase
.
, acetone
or ethanol
, but dissolution in absolute alcohols is difficult. Solutions of at least 50 µM DON in 0.9 % NaCl are lightly yellowish. The crystalline form appears as yellowish greenish needles. The specific rotation is [α]26D +21° (c = 5.4 % in H2O). In phosphate buffer, pH 7 are the ultraviolet absorption maxima at 274nm (E1%1cm. 683) and 244 nm (E1%1cm 376).
of different glutamine utilizing enzymes. Due to its similarity to glutamine it can enter catalytic centres of these enzymes and inhibits them by covalent binding, or more precisely by alkylation
. In the following table gives a survey of DON targets.
synthesis. It could be shown in vitro that DON treatment led to apoptosis
, the programmed cell death. Different pathways were investigated. So it could be shown that the inner mitochondrial membrane was damaged, and single strand DNA breaks were observed. The exact mode of action remains unclear and needs further research.
DON is not approved as pharmaceutical agent, but is tested in combination with a recombinant glutaminase in clinical trials for the treatment of different solid tumors.
Glutamine
Glutamine is one of the 20 amino acids encoded by the standard genetic code. It is not recognized as an essential amino acid but may become conditionally essential in certain situations, including intensive athletic training or certain gastrointestinal disorders...
antagonist
Receptor antagonist
A receptor antagonist is a type of receptor ligand or drug that does not provoke a biological response itself upon binding to a receptor, but blocks or dampens agonist-mediated responses...
, which was isolated originally from Streptomyces
Streptomyces
Streptomyces is the largest genus of Actinobacteria and the type genus of the family Streptomycetaceae. Over 500 species of Streptomyces bacteria have been described. As with the other Actinobacteria, streptomycetes are gram-positive, and have genomes with high guanine and cytosine content...
. It is a non-standard amino acid
Amino acid
Amino acids are molecules containing an amine group, a carboxylic acid group and a side-chain that varies between different amino acids. The key elements of an amino acid are carbon, hydrogen, oxygen, and nitrogen...
. The diazo compound
Diazo
Diazo refers to a type of organic compound called diazo compound that has two linked nitrogen atoms as a terminal functional group. The general formula is R2C=N2. The simplest example of a diazo compound is diazomethane...
was characterized in 1956 by Henry W Dion et al., who suggested a possible use in cancer therapy. This antitumoral efficacy was confirmed in different animal models. DON was tested as chemotherapeutic agent in different clinical studies, but was never approved. The last clinical results were published in 2008, though not as DON monotherapy but in combination with a recombinant glutaminase
Glutaminase
Glutaminase is an amidohydrolase enzyme which generates glutamate from glutamine. Glutaminase has tissue-specific isoenzymes. Importantly, glutaminase is found in glial cells.Glutaminase catalyzes the following reaction:Glutamine + H2O → Glutamate + NH3...
.
Chemistry
DON is a water soluble yellowish powder, which can be dissolved also in aqueous solutions of methanolMethanol
Methanol, also known as methyl alcohol, wood alcohol, wood naphtha or wood spirits, is a chemical with the formula CH3OH . It is the simplest alcohol, and is a light, volatile, colorless, flammable liquid with a distinctive odor very similar to, but slightly sweeter than, ethanol...
, acetone
Acetone
Acetone is the organic compound with the formula 2CO, a colorless, mobile, flammable liquid, the simplest example of the ketones.Acetone is miscible with water and serves as an important solvent in its own right, typically as the solvent of choice for cleaning purposes in the laboratory...
or ethanol
Ethanol
Ethanol, also called ethyl alcohol, pure alcohol, grain alcohol, or drinking alcohol, is a volatile, flammable, colorless liquid. It is a psychoactive drug and one of the oldest recreational drugs. Best known as the type of alcohol found in alcoholic beverages, it is also used in thermometers, as a...
, but dissolution in absolute alcohols is difficult. Solutions of at least 50 µM DON in 0.9 % NaCl are lightly yellowish. The crystalline form appears as yellowish greenish needles. The specific rotation is [α]26D +21° (c = 5.4 % in H2O). In phosphate buffer, pH 7 are the ultraviolet absorption maxima at 274nm (E1%1cm. 683) and 244 nm (E1%1cm 376).
Biochemistry
DON is used as inhibitorEnzyme inhibitor
An enzyme inhibitor is a molecule that binds to enzymes and decreases their activity. Since blocking an enzyme's activity can kill a pathogen or correct a metabolic imbalance, many drugs are enzyme inhibitors. They are also used as herbicides and pesticides...
of different glutamine utilizing enzymes. Due to its similarity to glutamine it can enter catalytic centres of these enzymes and inhibits them by covalent binding, or more precisely by alkylation
Alkylation
Alkylation is the transfer of an alkyl group from one molecule to another. The alkyl group may be transferred as an alkyl carbocation, a free radical, a carbanion or a carbene . Alkylating agents are widely used in chemistry because the alkyl group is probably the most common group encountered in...
. In the following table gives a survey of DON targets.
| Enzyme | Metabolic pathway | References |
|---|---|---|
| Carbamoyl phosphate synthase (CAD) | Pyrimidine-De-Novo-Synthesis Pyrimidine Pyrimidine is a heterocyclic aromatic organic compound similar to benzene and pyridine, containing two nitrogen atoms at positions 1 and 3 of the six-member ring... |
|
| CTP synthase CTP synthase CTP synthase is an enzyme involved in pyrimidine biosynthesis that interconverts UTP and CTP.- Enzyme Structure :... (CTPS) |
Pyrimidine-De-Novo-Synthesis Pyrimidine Pyrimidine is a heterocyclic aromatic organic compound similar to benzene and pyridine, containing two nitrogen atoms at positions 1 and 3 of the six-member ring... |
|
| FGAR amidotransferase | Purine-De-Novo-Synthesis Purine A purine is a heterocyclic aromatic organic compound, consisting of a pyrimidine ring fused to an imidazole ring. Purines, including substituted purines and their tautomers, are the most widely distributed kind of nitrogen-containing heterocycle in nature.... |
|
| Guanosine monophosphate Guanosine monophosphate Guanosine monophosphate, also known as 5'-guanidylic acid or guanylic acid and abbreviated GMP, is a nucleotide that is used as a monomer in RNA. It is an ester of phosphoric acid with the nucleoside guanosine. GMP consists of the phosphate group, the pentose sugar ribose, and the nucleobase... synthetase (GMPS) |
Purine-De-Novo-Synthesis Purine A purine is a heterocyclic aromatic organic compound, consisting of a pyrimidine ring fused to an imidazole ring. Purines, including substituted purines and their tautomers, are the most widely distributed kind of nitrogen-containing heterocycle in nature.... |
|
| PRPP amidotransferase | Purine-De-Novo-Synthesis Purine A purine is a heterocyclic aromatic organic compound, consisting of a pyrimidine ring fused to an imidazole ring. Purines, including substituted purines and their tautomers, are the most widely distributed kind of nitrogen-containing heterocycle in nature.... |
|
| Mitochondrial glutaminase Glutaminase Glutaminase is an amidohydrolase enzyme which generates glutamate from glutamine. Glutaminase has tissue-specific isoenzymes. Importantly, glutaminase is found in glial cells.Glutaminase catalyzes the following reaction:Glutamine + H2O → Glutamate + NH3... |
First step of glutaminolysis Glutaminolysis Glutaminolysis is a series of biochemical reactions by which the amino acid glutamine is degraded to glutamate, aspartate, CO2, pyruvate, lactate, alanine and citrate.- The glutaminolytic pathway:... |
|
| NAD Nicotinamide adenine dinucleotide Nicotinamide adenine dinucleotide, abbreviated NAD, is a coenzyme found in all living cells. The compound is a dinucleotide, since it consists of two nucleotides joined through their phosphate groups. One nucleotide contains an adenine base and the other nicotinamide.In metabolism, NAD is involved... synthase |
Coenzyme of the electron transport chain Electron transport chain An electron transport chain couples electron transfer between an electron donor and an electron acceptor with the transfer of H+ ions across a membrane. The resulting electrochemical proton gradient is used to generate chemical energy in the form of adenosine triphosphate... |
|
| Asparagine Asparagine Asparagine is one of the 20 most common natural amino acids on Earth. It has carboxamide as the side-chain's functional group. It is not an essential amino acid... synthetase |
Amino acid synthesis |
Pharmacology
DON is a cytotoxic inhibitor of many enzymes of nucleotideNucleotide
Nucleotides are molecules that, when joined together, make up the structural units of RNA and DNA. In addition, nucleotides participate in cellular signaling , and are incorporated into important cofactors of enzymatic reactions...
synthesis. It could be shown in vitro that DON treatment led to apoptosis
Apoptosis
Apoptosis is the process of programmed cell death that may occur in multicellular organisms. Biochemical events lead to characteristic cell changes and death. These changes include blebbing, cell shrinkage, nuclear fragmentation, chromatin condensation, and chromosomal DNA fragmentation...
, the programmed cell death. Different pathways were investigated. So it could be shown that the inner mitochondrial membrane was damaged, and single strand DNA breaks were observed. The exact mode of action remains unclear and needs further research.
DON is not approved as pharmaceutical agent, but is tested in combination with a recombinant glutaminase in clinical trials for the treatment of different solid tumors.