Abstract THE CHARACTERIZATION OF CHIMERIC CHAPERONE FLAGRP170 AS A NOVEL RADIOPROTECTANT By Tyler Nguyen, M.S. A thesis submitted in partial fulfillment of the requirements for the degree of Master of Science at Virginia Commonwealth University. Virginia Commonwealth University, 2017 Major Director: Dr. Xiang-Yang (Shawn) Wang, Ph.D., Professor, Department of Human and Molecular Genetics Radiation therapy (RT) is restricted by toxic effects on adjacent normal tissue, which limits RT efficacy in cancer treatment. Damage to normal tissue, such as radiosensitive intestine and bone marrow compartments, results in acute radiation damage. To reduce normal tissue injury in the setting of RT, we examine the potential radioprotectant, Flagrp170, a chimeric protein. Flagrp170 is comprised of glucose-regulated protein-170 (Grp170) and a NF-κB activating sequence derived from flagellin. We show that Flagrp170 can protect normal tissues post irradiation, indicated by TUNEL and clonogenic assays. However, treatment with Flagrp170 does not influence tumor response to RT. Studies indicate that Flagrp170 activates the transcription factor NF-κB, a strong pro-survival signal. In addition, Flagrp170 can induce production of radioprotective cytokines as well. Data suggests that Flagrp170 has potential as a novel radioprotectant in the setting of RT. The combination of Flagrp170 therapy and RT may lead to improved treatment outcomes.

Craton cores far from plate boundaries have traditionally been viewed as stable features that experience minimal vertical motion over 100-1000Ma time scales. Here we show that the Fennoscandian Shield in southeastern Sweden experienced several episodes of burial and exhumation from similar to 1800Ma to the present. Apatite, titanite, and zircon (U-Th)/He ages from surface samples and drill cores constrain the long-term, low-temperature history of the Laxemar region. Single grain titanite and zircon (U-Th)/He ages are negatively correlated (104-838Ma for zircon and 160-945Ma for titanite) with effective uranium (eU=U+0.235xTh), a measurement proportional to radiation damage. Apatite ages are 102-258Ma and are positively correlated with eU. These correlations are interpreted with damage-diffusivity models, and the modeled zircon He age-eU correlations constrain multiple episodes of heating and cooling from 1800Ma to the present, which we interpret in the context of foreland basin systems related to the Neoproterozoic Sveconorwegian and Paleozoic Caledonian orogens. Inverse time-temperature models constrain an average burial temperature of similar to 217 degrees C during the Sveconorwegian, achieved between 944Ma and 851Ma, and similar to 154 degrees C during the Caledonian, achieved between 366Ma and 224Ma. Subsequent cooling to near-surface temperatures in both cases could be related to long-term exhumation caused by either postorogenic collapse or mantle dynamics related to the final assembly of Rodinia and Pangaea. Our titanite He age-eU correlations cannot currently be interpreted in the same fashion; however, this study represents one of the first examples of a damage-diffusivity relationship in this system, which deserves further research attention.

This thesis presents work carried out using data from the LHCb experiment during the first three years of data taking, 2010 - 2012. A study of the effects of radiation damage on the silicon sensors of the LHCb Vertex Locator is performed, with an emphasis on the implications for the long term performance of the detector. Following three years of operation the sensors have received a maximum delivered neutron equivalent fluence of approximately 1.6E12 per square centimeter, leading to a number of radiation induced effects. In particular the change in charge collection efficiency and signal/noise with fluence is compared to theoretical expectations, and the current trends are extrapolated to the fluences expected at the end of the LHCb detector lifetime. The development of an unexpected effect due to the structure of the routing lines in the sensors is described in detail. Searches for lepton flavour and baryon number violating decays of the tau lepton using the 2011 LHCb dataset are described. Observation of any lepton flavour or baryon number violation would be an unambiguous sign of new physics, whilst setting improved limits helps to constrain a number of Beyond the Standard Model theories. First LHCb limits are set on the branching fractions of the decays tau- to mu- mu+ mu-, tau- to anti-proton mu+ mu- and tau- to proton mu- mu-, with these results also representing the first limits on lepton flavour violating tau decays at a hadron collider. The limit on tau- to mu- mu+ mu- is expected to approach the world's best result from Belle in the coming years whilst the tau- to anti-proton mu+ mu- and tau- to proton mu- mu- results constitute the first limits on the branching fractions of these decays. The future prospects for these measurements with further data are briefly described.

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When the thymine base is oxidized, the resulting cation may deprotonate reversibly at N3, or irreversibly at >C5-CH3. In all thymine derivatives studied so far in the solid state, there is always a significant concentration of a radical formed by net H-abstraction from the >C5-CH 3. DFT calculations on this allyl-like radical are in good agreement with the experimental results for both the isotropic and anisotropic hyperfine couplings. There is a tendency for the thymine cation to deprotonate at N3 in solution. Calculations on the N3 deprotonated thymine cation yield two structures, one planar radical with an unusually large N1-C2 bond length, and one nonplanar radical with the N3 more than 25° out of the molecular plane. Calculations show that the structure with the lowest energy is the allyl-like radical.

In 2027 the Large Hadron Collider (LHC) at CERN will enter a high luminosity phase, deliver- ing 3000 fb 1 over the course of ten years. The High Luminosity LHC (HL-LHC) will increase the instantaneous luminosity delivered by a factor of 5 compared to the current operation pe- riod. This will impose significant technical challenges on all aspects of the ATLAS detector but particularly the Inner Detector, trigger, and data acquisition systems. In addition, many of the components of the Inner Detector are reaching the end of their designed lifetime and will need to be exchanged. As such, the Inner Detector will be entirely replaced by an all silicon tracker, known as the Inner Tracker (ITk). The layout of the Pixel and strip detectors will be optimised for the upgrade and will extend their forward coverage. To reduce the per-pixel hit rate and explore novel techniques for deal- ing with the conditions in HL-LHC, an inter-experiment collaboration called RD53 has been formed. RD53 is tasked with producing a front-end readout chip to be used as part of hybrid Pixel detectors that can deal with the high multiplicity environment in the HL-LHC. A silicon sensor, which makes up the other half of the hybrid Pixel detector, must also be designed to cope with the high fluences in HL-LHC. Significant damage will be caused by non- ionising energy loss in the sensor over its lifetime. This damage must be incorporated into the detector simulation both to predict the detector performance at specific conditions and to understand the e↵ects of radiation damage on data taking. The implementation of radiation damage in the ATLAS simulation framework is discussed in this thesis. Collisions produced by the HL-LHC also presents a challenge for the current track reconstruc- tion software. High luminosity is obtained, in part, by increasing the number of interactions per bunch crossing, which in turn increases the time taken for track reconstruction. Various ap- proaches to circumvent the strain on projected resources are being explored, including porting existing algorithms to parallel architectures. A popular algorithm used in track reconstruction, the Kalman filter, has been implemented in a neuromorphic architecture: IBM’s TrueNorth. The limits of using such an architecture for tracking, as well as how its performance compares to a non-spiking Kalman filter implementation, are explored in this thesis.

This review contains the results of EPR/ENDOR experiments on DNA constituents in the solid-state. Most of the results presented involve single crystals of the DNA bases, nucleosides and nucleotides. The emphasis is on low-temperature ENDOR results. Typical experiments involve irradiations at or near helium temperatures in attempts to determine the primary radiation induced oxidation and reduction products. The use of the ENDOR technique allows one to determine the protonation state of the initial products. Subsequent warming of the sample facilitates a study of the reactions that the primary products undergo. A summary of the results is provided to show the relevance the study of model compounds has in understanding the radiation chemistry of DNA.

Detailed EPR and ENDOR experiments on the cocrystalline complex of 1-methyluracil:9-Ethyladenine (MUEA) have revealed that the major radiation-induced products observed at 10 K on MU are: MUEA1, a radical formed by net hydrogen abstraction from the N1-CH3 methyl group, MUEA2, the MU radical anion, and MUEA3, the C5 H-addition radical. The following four products were observed on the adenine moiety at 10 K, MUEA4, the N3 protonated adenine anion, MUEA5, the native adenine cation, MUEA6, the amino deprotonated adenine cation, and MUEA7, the C8 H-addition radical formed by net H-addition to C8 of the adenine base. The geometries, energetics, and hyperfine properties of all possible radicals of MU and EA, the native anions and cations, as well as radicals formed via net hydrogen atom abstraction (deprotonated cations) or addition (protonated anions) were investigated theoretically. All systems were optimized using the hybrid Hartree-Fock-density functional theory functional B3LYP, in conjunction with the 6-31G(d,p) basis set of Pople and co-workers. Calculations of the anisotropic hyperfine couplings for all the radicals observed in MUEA are presented and are shown to compare favorably with the experimentally measured hyperfine couplings. The calculated ionizations potentials indicate that EA would be the preferred oxidation site. In MUEA, both the adenine cation and its N4-deprotonated derivative were observed. The calculated electron affinities indicate that MU would be the preferred reduction site. In MUEA radical, MUEA2 is a uracil reduction product, however the protonation state of this radical could not be determined experimentally. Calculations suggest that MUEA2 is actually the C4=O protonated anion.

Detailed electron paramagnetic resonance and electron nuclear double resonance experiments on the co-crystalline complex of 1-methylthymine:9-methyladenine (MTMA) have revealed that the major radiation induced products at low temperatures (10 K) are MTMA1, a radical formed by net hydrogen abstraction from the C5 methyl group on thymine, and MTMA2, a radical formed by net hydrogen abstraction of the N1 methyl group on thymine. The following four minor products were also observed: MTMA3, the C4-OH protonated anion of thymine, MTMA4, the C6 H-addition product of thymine, and MTMA5 and MTMA6, radicals formed by net H-addition to C2 and C8 of the adenine base. The geometries, energetics and hyperfine properties of all possible radicals of MT and MA, the primary anions and cations, as well as radicals formed via net hydrogen atom abstraction (deprotonated cations) or addition (protonated anions) were investigated theoretically. All systems were optimized using the hybrid Hartree-Fock density functional theory functional B3LYP, in conjunction with the 6-31G(d,p) basis set of Pople and co-workers. Calculations of the anisotropic hyperfine couplings for all the radicals observed in MTMA are presented, and are shown to compare favorably with the experimentally measured hyperfine couplings. The calculated ionizations potentials indicate that MA would be the preferred oxidation site. However, in MTMA neither the adenine cation nor its N4-deprotonated derivative were observed. The adenine cation in MTMA is not stabilized by deprotonation, and is thus likely subject to recombination. The calculated electron affinities indicate that MT would be the preferred reduction site. Reduction of MT is believed to result in protonation of the anion at C4=O. The calculated hyperfine couplings for the MT anion are very similar to those of the C4-OH protonated anion, and therefore, the theoretical calculations are not useful in deciding the actual protonation state of this reduction product.

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