| dc.description.abstracteng | Nuclear magnetic resonance spectroscopy (NMR spectroscopy) and its related technique,
magnetic resonance imaging (MRI), enable scientists and doctors to make statements about the
atomic structure of a sample and to produce anatomical images of the human body. The effect of
magnetic resonance is used to manipulate magnetically active atomic nuclei. By applying an
external, constant magnetic field in the NMR spectrometer or the MRI machine, it is possible to
align the atomic nuclei either parallel or antiparallel to the external magnetic field, like small
magnets. This produces the so-called thermal polarization, which describes the sum of the
magnetization of a sample. It is roughly as if all the atomic nuclei, which we imagine as small
magnets, had formed a large magnet together. The thermal polarization thus describes the
strength of this entire magnet. With additional, short radio frequency (RF) pulses, it is possible to
trigger the magnetic resonance and set the thermal polarization in motion. This is often done on
hydrogen nuclei (1H), but investigations on so-called heteronuclei such as 13C are also possible.
However, a major challenge of NMR and MRI is their sensitivity. Due to the aforementioned
parallel or antiparallel alignment of the atomic nuclei, the magnetization vectors of a large part of
the atomic nuclei cancel each other out. Only a fraction actually contributes to the detectable
signal. This leads to the fact that in MRI, the signals of hydrogen nuclei in water and fat are
detected for the most part, since they occur in high concentrations in the body. By developing
special measurement programmes, the RF pulse sequences, it is nevertheless possible to produce
anatomically high-resolution images. However, for special applications, such as the improved
imaging of cancer, contrast agents are required.
In this dissertation, contrast agents were investigated that achieve NMR or MRI signal
enhancement with the help of hyperpolarization. Hyperpolarization refers to a state of increased
magnetization of a sample compared to its thermal polarization. One of the methods of producing
hyperpolarization is para-hydrogen induced polarization (PHIP). Here, para-hydrogen is used, an
isomer of hydrogen, and its polarization is transferred to another target nucelus with pulse
sequences. In the case of PHIP side-arm hydrogenation (PHIP-SAH), a modified molecule is used
for this, which has a so-called side arm to which para-hydrogen is chemically added. After the
pulse sequence and polarization transfer, this side arm is split off and the target molecule is
present in its native form, with a strongly increased signal for NMR or MRI. One target molecule
of particular interest is pyruvate, which plays a central role in human metabolism. In many types
of cancer, for example, the metabolic conversion is greatly increased and altered compared to
healthy tissue. By applying hyperpolarized pyruvate, it is then possible to better visualize cancer
during an MRI examination, as well as to make additional statements about its metabolic activity.
This could also be used for early detection or to assess the success of therapy. Other areas of
application are neurodegeneration and cardiovascular diseases.
There are already initial findings from clinical studies in humans that demonstrate the potential
of hyperpolarized pyruvate. However, the pyruvate for these studies was generated using a
different technique, Dynamic Nuclear Polarization (DNP). PHIP-SAH has several advantages over
this technique. For example, the production of one dose of hyperpolarized pyruvate can be done
much quicker, and with simpler and cheaper equipment. However, PHIP-SAH was developed later
and is therefore not yet in clinical trials.
This thesis demonstrates the process of biological application of hyperpolarized pyruvate using
PHIP-SAH. First, the method was optimized so that physiologically compatible application
solutions could be produced. These were then examined in a first application in cell culture
samples (in vitro). Here, HEK298T cells (human embryonic kidney cells) were used, which
contain a protein that occurs in neurodegenerative diseases in larger aggregates, either strongly
increased or not at all. In addition, the cells were treated with a metabolic inhibitor. After
examining these cells with hyperpolarized pyruvate, we were able to show the differences in the
metabolic conversion of pyruvate of the different cell groups. This served as a feasibility study
that we are able to visualize metabolic differences with PHIP-SAH pyruvate.
In a second project, the hyperpolarized pyruvate was now tested in vivo. For this purpose, three
mice each were examined that had human-derived pancreatic or colon tumors (xenografts).
Pancreatic cancer is considered one of the most aggressive cancers and has a poor prognosis
because it is often diagnosed at an advanced stage. Colorectal cancer has a higher survival rate,
but due to its frequency it still causes a high number of deaths. For both types of cancer, therefore,
an additional examination technique would be helpful to assist in early detection and therapy
monitoring. After injection of hyperpolarized pyruvate, it has been possible to perform time-
resolved, localized 13C spectroscopy and thus to visualize the pyruvate metabolism of the tumors.
The metabolic conversion of pyruvate to its metabolites lactate and alanine was significantly
higher in the pancreatic tumors than in the colon tumors. This made it possible to differentiate
between these two types of cancer. Our study represents the first investigation of pancreatic and
colorectal cancer in vivo with PHIP-SAH pyruvate and serves as a starting point for further
research. In future, studies will be carried out with a larger number of mice and pulse sequences
that allow better resolution and localization of the 13C signal will be used.
In a third project, the method transfer from the previous preclinical experimental setup to a
clinically used MRI device was started. The challenges here lie in the technical implementation.
Hyperpolarization can now no longer be performed on high-field NMR spectrometers, but must
be carried out with portable polarizers. Two different polarizers were tested for
hyperpolarization in this project. In addition, different pulse sequences suitable for 13C and
hyperpolarized pyruvate were obtained and optimized for this purpose. First experiments were
carried out on thermally polarized standard samples, phantoms as well as with a mouse. These
developments are the starting point for further studies on mice, which will lead to studies on large
animals and finally to clinical application. | de |