Page 105 - Hematology_ Basic Principles and Practice ( PDFDrive )

76 Part I Molecular and Cellular Basis of Hematology

Depending on the cell type and specific state (growth, hypoxia, Most of the regulatory pathways that are associated with nucleo-
fasting, and so on) intracellular amino acids are used in anabolic or tide synthesis and degradation are strictly controlled by regulatory
catabolic pathways. components of the cell cycle machinery. The amount of intracellular
Most of the regulation of amino acid metabolism is achieved nucleotides has to reach certain levels in order for the cell to proceed
through substrate fluxes affecting specific enzyme kinetics. However, through the S-phase checkpoint. In addition, several of the key cell
there are two major regulatory pathways that involve amino acid cycle regulators, including the c-myc oncogene (which is translocated
sensing mechanisms and metabolic control. (1) General control in certain myelomas), directly increase the expression of most of the
nonrepressed 2 (GCN2) is a protein kinase that senses amino acid key enzymes associated with nucleotide synthesis.
deficiency through direct binding to uncharged tRNA. GCN2
controls the transcription factor ATF4, affecting different enzymes
of amino acid metabolism. (2) mTOR is a protein kinase activated Nucleotide Synthesis
in response to increased amino acid concentrations (particularly
branch chain amino acids). mTOR controls many aspects involved in There are two pathways for the synthesis of nucleotides, salvage and
protein synthesis, inhibition of protein degradation, and amino acid de novo. The salvage pathway uses free bases via a reaction with phos-
biosynthetic enzymes. The high asparagine requirement of certain phoribosyl pyrophosphate (PRPP) and generation of nucleotides.
acute lymphoblastic leukemias has resulted in the use of asparaginase De novo pathways synthesize pyrimidines and purine nucleotides
to deplete circulating levels of asparagine. Limited amounts of aspara- from amino acids, carbon dioxide, folate derivatives, and PRPP.
gine result in activation of GCN2 in leukemic cells, and reduce their Importantly, both salvage and de novo pathways depend on PRPP,
proliferation and viability rates. which is produced from ATP and ribose-5-phosphate (generated in
the pentose phosphate pathway) by PRPP synthetase, an enzyme
that is inhibited by metabolic markers of low-energy AMP, ADP, and
Biosynthesis of Nonessential Amino Acids GDP to avoid nucleotide synthesis in these conditions. In general,
PRPP levels are low in postmitotic cells but high in proliferating
Nonessential amino acids are synthesized by most of the cells, includ- cells. Folate is essential in nucleotide biosynthesis, and lack of folate
ing hematopoietic lineages. Nonessential amino acids are mainly in the diet can lead to anemia due to inhibition of proliferation of
synthesized from glucose (alanine, arginine [from the urea cycle in red blood cell precursors.
hepatic cells], asparagine, aspartate, cysteine, glutamate, glutamine,
glycine, proline, and serine), except for tyrosine, which is synthesized
from phenylalanine. The rest of the nine amino acids are essential Nucleotide Degradation
and the body needs to obtain these from the diet. Serine, glycine,
and cysteine are synthesized from glycolytic intermediates. Serine Nucleotidases and nucleosidases initially participate in purine nucleo-
synthesis has recently been found to be increased and necessary in tide degradation. For example, adenosine is deaminated to produce
stem cells. For some hematopoietic cells, the synthesis of cysteine and inosine, which, after ribose is removed, generates hypoxantine,
glycine is of elevated importance owing to their use in the synthesis of which is used by xanthine oxidase to form uric acid. Immune cells
the tripeptide glutathione. Aspartate and asparagines are synthesized have potent nucleotide salvage pathways, and a lack of adenosine
by transamination of oxaloacetate by glutamate and amide transfer deaminase causes severe combined immune deficiency (SCID)
from glutamine, respectively. Glutamate, glutamine, proline, and syndrome. SCID is associated with a large accumulation of dATP
arginine are formed from the TCA cycle intermediate α-ketoglutarate. in immune cells, which, through a negative-feedback mechanism on
ribonucleotide reductase, blocks production of dNTPs and results in
a failure to replicate DNA.
Amino Acid Catabolism

Two central reactions in amino acid catabolism are the generation of Introduction to Metabolomics
ammonia through transamination (catalyzed by amino transferases)
and oxidative deamination (catayzed by glutamate dehydrogenase) in Analytical measurements of blood metabolites such as glucose, urea,
which the α-amino group of the different amino acids is transferred and cholesterol is part of clinical biochemistry to track diseases.
to α-ketoglutarate to form glutamate, which undergoes the release Along these lines and facing the new era of personalized medicine
of free NH 3 . Free ammonium is added to glutamate to generate emerges metabolomics, which evaluates metabolism with a com-
glutamine, which is then exported into the circulation to the liver, prehensive and quantitative analysis of all metabolites, as well as
where it then enters the urea cycle. The urea cycle only occurs in the its impact on cell biology, and aims to discover novel biomarkers
liver and has two purposes: (1) to get rid of free ammonium; and or targets for therapy. Recent technical innovations in mass spec-
(2) to supply arginine. Interestingly, one of the enzymes of the urea trometry and nuclear magnetic resonance (NMR) have allowed the
cycle, arginase (which converts arginine to ornithine) is expressed in measurement of many metabolites simultaneously. These advances,
immune cells. Myeloid cell arginase depletes arginine and suppresses in combination with metabolite flux analysis with isotopic tracers,
T-cell immune response, and is an important mechanism of inflam- have provided new information on many metabolic processes. The
mation associated with immunosuppression. Arginase is viewed as use of metabolomics also offers a tool to identify metabolic enzymes
a promising strategy in the treatment of cancer and autoimmunity. as drug targets, as they are poised for inhibition with small-molecule
Arginine is also essential for the differentiation and proliferation of drugs and possess allosteric sites that can be utilized to alter catalytic
erythrocytes. activity.
A major effort in metabolomics has been the identification of
biomarkers for diseases and therapeutic targets. As an example,
Nucleotide Metabolism metabolomics was used to analyze plasma from diabetic patients
showing increases in branch chain amino acids before hyperglycemia.
Nucleotides are involved in a diverse array of cellular functions includ- Another example comes from the combination of genome-wide
+
+
+
ing (1) energy metabolism (ATP, NAD , NADP , and FAD and sequencing analysis and metabolomics: sequence analysis of acute
their corresponding reduced forms); (2) units of nucleic acids (NTPs myeloid leukemias was able to identify IDH1 or IDH2 mutations
are substrates for RNA and DNA polymerases); and (3) physiologic in 20% of patients. Metabolomics analysis revealed accumulation
mediators such as adenosine, ADP (which is critical in platelet aggre- of a noncanonical metabolite, 2-hydroxyglutarate, which promotes
gation), cAMP and cGMP (second messenger molecules), and GTP the tumorigenic process (see following discussion). In general, there
(which participates in signal transduction via GTP-binding proteins). are two different metabolomic approaches: targeted, which measures