Tremendous technology improvements of the last decades has given mass
spectrometry a more and more expanding role in the study of a wide range of
molecules: from the identification and quantification of small molecular weight
molecules to the structural determination of biomacromolecules. Many are the
fields of application for this technique and the various versions of it.
In the present study three different applications have been explored.
The first application is a pharmacokinetics study of anticancer drug Gemcitabine
and its principal metabolite, where the role of the LC-MS/MS is essential both for
the selectivity of the detection of the small analytes and the sensitivity enhanced
by multi-reaction monitoring experiments. The design of the study involved the
collection of several blood samples at selected times and from patients that
would have met certain eligibility criteria. The ESI demonstrated to be the most
suitable approach and it provided the necessary data to conclude that toxicity of
Gemcitabine did not increase when administered at FDR (Fixed Dose Rate)
infusion in patients with impaired hepatic function.
The second application describes an example of how MS represents a powerful
tool in cancer research, from serum profiling study with high resolution MALDITOF
and bioinformatic analysis, to the identification of potential biomarker
through peak identification. Almost 400 serum sample – homogeneously
distributed between biopsy confirmed ovarian cancer and high risk serum
samples – were analyzed on a high resolution MALDI-TOF instrument after
automated reverse phase magnetic beads separation. The high throughput data
have undergone sophisticated bioinformatic procedures that lead to a list of upand
down-regulated peaks, although identification studies were possible only for
those peaks that showed a good reproducibility. One down-regolated peak has
been identified using the LC-MS/MS technique. The identified peak confirmed a
basic role of fibrinogen in the ovarian cancer; the other four peaks that have been identified as down-regulated showed an absolutely not satisfactory ionization in
electro-spray, therefore further analysis will be performed on these analytes in
order to determinate their amino acidic sequence. The most suitable technique
seems to be MALDI-TOF/TOF mass spectrometry, since the peptides already
showed a good degree of ionization in MALDI.
The third and last study belongs to a quite new field, which is the combination of
immuno precipitation assays with MALDI-TOF (Immuno Precipitation Mass
Spectrometry, IPMS) experiments in order to evaluate the specificity of a series
of monoclonal antibodies to specific antigen. The automated assay that has been
developed provides structural information about the antigen that binds the
monoclonal antibody to be tested and previously conjugated to the surface of
magnetic beads, ideal support for robotic automation. IPMS showed its potential
as a complementary tool of crucial importance in the selection of the monoclonal
antibody for the development of ELISA based assay to be applied in the
screening of a consistent number of human specimens for the clinical validation
of proteins indicated in literature as potential biomarkers.
Mass spectrometry in association with fractionation techniques, such as liquid or
magnetic beads chromatography, is a very flexible tool in the cancer research
field. Further improvement in the instrumentation and in the technology will bring
always more and more results to be confident in