Environmental Engineering Reference
In-Depth Information
mainly in MPS I (Hurler) and MPS VII diseases, while for MPS II and MPS VI is
used in conjunction with ERT [
50
]; (4)
substrate reduction therapy (SRT)
involves
the use of certain selective inhibitors, as
L
-cycloserine, to reduce the substrate
synthesis to a level approximately equal to the rate of breakdown. Since this type
of therapy presents significant side effects, its use is limited, however is success-
fully used in the Gaucher disease [
51
]; (5)
gene therapy
which requires in vivo
injection of a gene transfer vector directly into a tissue or into the circulation, or
ex vivo
genetically modification of cells followed by their transplant into an
affected patient in order to create a repository of enzyme that can be secreted into
the circulation [
52
,
53
]. Although up to now have been performed preclinical
studies on animal models of lysosomal storage diseases, and just a few clinical
trials on human with lysosomal storage diseases, gene therapy is a promising
treatment modality for the LSDs.
Taking into account that only 5-6 decades have passed since the discovery of the
lysosome and its correlation with certain diseases already described, it is obvious
the effort on investigating the lysosomal storage diseases and the advanced level
reached on their treatment.
8.4 Applications of Mass Spectrometry to Diagnosis
of Lysosomal Storage Diseases
Considering the advanced level achieved in terms of treatment methods,
presymptomatic diagnosis and early intervention are crucial. As briefly mentioned
above, in the screening of LSDs, several assays are used, as the fluorometric assays
with 4-methylumbelliferone (4-MU) substrates, tandem MS (MS/MS) or liquid
chromatography MS/MS (LC-MS/MS), multiplex enzyme assays, digital
microfluidic fluorometric assays, and immune-quantification assays. Unlike the
fluorometric assay, high-throughput mass spectrometry methods based on either
ESI or matrix-assisted laser desorption/ionization (MALDI), originally developed
to screen newborns for lysosomal storage disorders, are most specific, powerful,
reliable and efficient tools. On one hand, these methods can be used from different
perspectives: for quantification, by measuring in urine or blood samples the sub-
strate concentration, or the enzyme activity, as well as for investigation of protein
expression on cultured cells; on the other hand, the chosen or synthesized substrates
for these assays are structurally closer to the natural substrates, as the artificial
4-MU substrates. The procedure for the measurement of enzyme activities involves
a simple reasoning: based on the fact that enzyme products are specific to each
enzyme reaction, they are quantified against internal standards with known con-
centrations. Since the method was optimized and refined into a standardized
protocol, in 2008 Genzyme Corporation began the production in large quantity
and the distribution of the analyte specific regents (ASR), namely the substrates,
internal standards and products. Due to the complexity of the initial procedures,
