Dissertations / Theses: 'Preclinical models of addiction'

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Fouyssac, Maxime. "Behavioural and cellular basis of the vulnerability to develop compulsive heroin seeking habits." Thesis, University of Cambridge, 2017. https://www.repository.cam.ac.uk/handle/1810/271886.

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Addiction is a chronic relapsing disorder for which there is no effective treatment. This may reflect our lack of understanding of the psychological and neural mechanisms that support the transition, in vulnerable individuals, from recreational drug use to compulsive drug seeking habits. Over the last decade clinical and preclinical studies have begun to shed light on the psychological and neural basis of the individual vulnerability to cocaine addiction, but despite the epidemic in opiates addiction in the USA and incremental opioid drug abuse and addiction in the UK, heroin addiction has hitherto been under-investigated. Using a novel preclinical model of compulsive heroin seeking behaviour in which some rats self-administering heroin persist in responding under a second-order schedule of reinforcement despite punishment (Chapter 3), the experiments in this thesis investigated the psychological, behavioural, neural and cellular mechanisms involved in the vulnerability to develop compulsive heroin seeking. Chapter 4 aimed to identify behavioural traits, such as anxiety, stress reactivity or decision making, that predict an increased vulnerability to develop compulsive heroin seeking. Chapter 5 aimed to characterise the neural and cellular correlates of heroin seeking habits, and compulsivity. Based on the combination of hotspot analysis, quantitative PCR, RNAscope and western-blot analyses, the data presented demonstrate that compulsive habits are associated with a differential pattern of cellular plasticity within corticostriatal networks, and are preceded by diverse cellular adaptations, especially in the striatum, in vulnerable individuals. Finally, chapter 6 further investigated the cellular specificity of the observed adaptations in experiments that revealed exposure to heroin and cocaine, triggers a downregulation of the dopamine transporter preferentially in astrocytes, and not in neurons as previously thought. The results presented in this thesis offer new insights into the neural and cellular basis of the vulnerability to develop compulsive heroin seeking, a key feature of opioid addiction.

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Zhang, Ting. "DEVELOPMENT AND PRECLINICAL EVALUATION OF LONG-LASTING COCAINE HYDROLASES FOR COCAINE OVERDOSE AND COCAINE USE DISORDER TREATMENT." UKnowledge, 2018. https://uknowledge.uky.edu/pharmacy_etds/93.

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Cocaine is a plant-based illicit drug commonly involved in substance use disorder. Although cocaine overdose and cocaine use disorders cause adverse health consequences to individuals and the economic burden on their family and society, there are no FDA (Food and Drug Administration) approved medications for treatment. Recently, it has been recognized that delivery of cocaine hydrolase (CocH) is a promising therapeutic strategy. Human butyrylcholinesterase (hBChE), the primary enzyme involved in cocaine metabolism in human, have advantages over other candidates for the development of CocH. Previous studies in our laboratory have designed and characterized hBChE mutants that have ~4,000-fold improved catalytic efficiency against naturally occurring (-)-cocaine as compared to the wild-type hBChE. Besides the catalytic efficiency, the biological half-life is another essential factor that influences the desired therapeutic value in the long-term treatment of cocaine use disorder. In order to provide prolonged effects to reduce administration frequency in clinical use, efforts have been made to increase the retention time of CocHs in blood circulation by fusing CocHs with other thermostable proteins or their mutants, including human serum albumin (Albu) or the Fc region of the human IgG (Fc). In this dissertation, we demonstrated the clinical potential and the benefits of long-lasting CocHs for cocaine overdose treatment. We used rodent models to show the ability of AlbuCocH1 to block or reverse manifestations of toxic effects of cocaine. In addition, a concomitant LC-MS/MS-based analysis was conducted to investigate the pharmacokinetic profile of a lethal dose of cocaine with the presence of AlbuCocH1. These experimental data demonstrated AlbuCocH1 as an effective cocaine detoxification agent by accelerating the metabolism of cocaine. In order to examine the potential therapeutic value of Fc-fused CocHs in the treatment of cocaine use disorder, we conducted a series of behavioral experiments in rats to evaluate the effectiveness and duration of Fc-fused CocHs in blocking or attenuating cocaine-induced psychostimulant and discriminative stimulus effects. In addition, the intravenous self-administration model was used to investigate the long-term effectiveness of Fc-fused CocHs in blocking or attenuating the reinforcing effects of cocaine. It has been shown that a single dose of E30-6-Fc (3 mg/kg) was able to effectively alter the cocaine dose-response curve and attenuate the reinforcing efficacy of cocaine for at least a month in both male and female rats. In summary, AlbuCocH1 (TV-1380), which failed to meet the primary efficacy endpoint in clinical trials for facilitating abstinence in cocaine-dependent subjects with a weekly dosing schedule (due to the short biological half-life), is more suitable to be developed as a cocaine detoxification agent. On the contrary, the newly designed Fc-fused CocH (e.g. CocH3-Fc, E30-6-Fc) with higher catalytic efficiency and longer biological half-life will be beneficial for long-term abstinence management in cocaine-dependent individuals.

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Delgado, San Martin Juan A. "Mathematical models for preclinical heterogeneous cancers." Thesis, University of Aberdeen, 2016. http://digitool.abdn.ac.uk:80/webclient/DeliveryManager?pid=230139.

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Cancer is a deadly, complex disease with 14 million new cases diagnosed every year and the endeavour to develop a cure is a global multidisciplinary effort. The complexity of cancer and the resulting vast volume of data derived from its research necessitates a robust and cutting-edge system of mathematical and statistical modelling. This thesis proposes novel mathematical models of quantification and modelling applied to heterogeneous preclinical cancers, focusing on the translation of animal studies into patients with particular emphasis on tumour stroma. The first section of this thesis (quantification) will present different techniques of extracting and quantifying data from bioanalytical assays. The overall aim will be to present and discuss potential methods of obtaining data regarding tumour volume, stromal morphology, stromal heterogeneity, and oxygen distribution. Firstly, a 3D scanning technique will be discusses. This technique aims to assess tumour volume in mice more precisely than the current favoured method (callipers) and record any cutaneous symptoms as well, with the potential to revolutionise tumour growth analysis. Secondly, a series of image processing methods will be presented which, when applied to tumour histopathology, demonstrate that tumour stromal morphology and its microenvironment play a key role in tumour physiology. Lastly, it will be demonstrated through the integration of in-vitro data from various sources that oxygen and nutrient distribution in tumours is very irregular, creating metabolic niches with distinct physiologies within a single tumour. Tumour volume, oxygen, and stroma are the three aspects central to the successful modelling of tumour drug responses over time. The second section of this thesis (modelling) will feature a mathematical oxygen-driven model - utilising 38 cell lines and 5 patient-derived animal models - that aims to demonstrate the relationship between homogeneous oxygen distribution and preclinical tumour growth. Finally, all concepts discussed will be merged into a computational tumour-stroma model. This cellular automaton (stochastic) model will demonstrate that tumour stroma plays a key role in tumour growth and has both positive (at a molecular level) and negative (at both a molecular and tissue level) effects on cancers. This thesis contains a useful set of algorithms to help visualise, quantify, and understand tissue phenomena in cancer physiology, as well as providing a series of platforms to predict tumour outcome in the preclinical setting with clinical relevance.

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Chaffee, Beth K. "Preclinical Modeling of Musculoskeletal Cancer." The Ohio State University, 2013. http://rave.ohiolink.edu/etdc/view?acc_num=osu1376844544.

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Henkel, Jan. "Bone tissue engineering in two preclinical ovine animal models." Thesis, Queensland University of Technology, 2017. https://eprints.qut.edu.au/109909/1/Jan_Henkel_Thesis.pdf.

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This PhD-research was focused on the development and evaluation of innovative scaffold-based bone tissue engineering concepts for the treatment of large volume bone defects, which still represent a major challenge in orthopaedic and reconstructive surgery. Two different types of bone tissue engineering constructs were investigated and successfully applied to regenerate critically-sized segmental bone defects in ovine animal models. The results outlined in the PhD thesis represent a significant contribution to potential future clinical translations of bone tissue engineering concepts from bench to bedside.

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Ernst, Agnes Stefanie. "Molecular analysis of preclinical models for mental and metabolic disorders." Thesis, University of Cambridge, 2012. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.610813.

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Wetzel, Hanna N. "Preclinical Development of the Anti-cocaine Monoclonal Antibody h2E2 for the Treatment of Cocaine Addiction." University of Cincinnati / OhioLINK, 2017. http://rave.ohiolink.edu/etdc/view?acc_num=ucin1529333855259163.

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Creedon, Helen. "Use of genetically engineered mouse models in preclinical drug development." Thesis, University of Edinburgh, 2015. http://hdl.handle.net/1842/15911.

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The paucity of well validated preclinical models is frequently cited as a contributing factor to the high attrition rates seen in clinical oncological trials. There remains a critical need to develop models which are accurately able to recapitulate the features of human disease. The aims of this study were to use genetically engineered mouse models (GEMMs) to explore the efficacy of novel treatment strategies in HER2 positive breast cancer and to further develop the model to facilitate the study of mechanisms underpinning drug resistance. Using the BLG--HER2KI-PTEN+/- model, we demonstrated that Src plays an important role in the early stages of tumour development. Chemopreventative treatment with dasatinib delayed tumour inititation (p= 0.046, Wilcoxon signed rank test) and prolonged overall survival (OS) (p=0.06, Wilcoxon signed rank test). Dasatinib treatment also induced squamous metaplasia in 66% of drug treated tumours. We used 2 cell lines derived from this model to further explore dasatinib’s mechanism of action and demonstrated reduced proliferation, migration and invasion following in vitro treatment. Due to the prolonged tumour latency and the low metastatic rate seen in this model, further studies were undertaken with the MMTV-NIC model. This model also allowed us to study the impact of PTEN loss on therapeutic response. We validated this model by treating a cohort of MMTV-NIC PTEN+/- mice with paclitaxel and demonstrated prolonged OS (p=0.035, Gehan Breslow Wilcoxon test). AZD8931 is an equipotent signalling inhibitor of HER2, HER3 and EGFR. We observed heterogeneity in tumour response but overall AZD8931 treatment prolonged OS in both MMTV-NIC PTEN FL/+ and MMTV-NIC PTEN+/- models. PTEN loss was associated with reduced sensitivity to AZD8931 and failure to suppress Src activity, suggesting these may be suitable predictive biomarkers of AZD8931 response. To facilitate further studies exploring resistance, we transplanted MMTV-NIC PTEN+/- fragments into syngeneic mice and generated 3 tumours with acquired resistance to AZD8931. These tumours displayed differing resistance strategies; 1 tumour continued to express HER2 whilst the remaining 2 underwent EMT and lost HER2 expression reflecting to a very limited degree some of the heterogeneity of resistance strategies seen in human disease. To further explore resistance to HER2 targeting tyrosine kinase inhibitors, we generated a panel of human cell lines with acquired resistance to AZD8931 and lapatinib. Western blotting demonstrated loss of HER2, HER3 and PTEN in all resistant lines. Acquisition of resistance was associated with a marked change in phenotype and western blotting confirmed all lines had undergone EMT. We used a combination of RPPA and mass spectrometry to further characterise the AZD8931 resistant lines and identified multiple potential novel proteins involved in the resistant phenotype, including several implicated in EMT. In conclusion, when coupled with appropriate in vitro techniques, the MMTV-NIC model is a valuable tool for selection of emerging drugs to carry forward into clinical trials of HER2 positive breast cancer.

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Precazzini, Francesca. "Zebrafish models of uveal and cutaneous melanoma for preclinical studies." Doctoral thesis, Università degli studi di Trento, 2019. http://hdl.handle.net/11572/245749.

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Uveal melanoma (UM) is the most common primary cancer of the eye and its prognosis is strongly influenced by the occurrence of metastases, which are both rapidly developing and mostly fatal. The most frequent driver mutations occur in a small number of genes including GNAQ, GNA11, BAP1, CYSLTR2 and SF3B1. Due to a lack of suitable animal models, the mechanism through which mutations in these genes cause or cooperate in UM initiation and progression is still largely unknown. We aimed at generating transgenic strains expressing the human mutant proteins in zebrafish uveal melanocytes, using the kita promoter. We used the binary Gal4/UAS system to express the mutant genes mentioned above. Moreover, we performed xenotransplantation experiments with uveal melanoma human and zebrafish cell lines in optically-clear, immunocompromised, zebrafish larvae. Transplanted fish developed melanoma near the site of transplantation in two weeks and showed metastatic growth within one month of age. This approach could be used for short-term assays in larvae, and be further developed for long-term uveal melanoma studies. In parallel, we performed a chemical screen using a transgenic model previously generated in our laboratory, where oncogenic RAS is expressed under the kita promoter. As adults transgenic kita:RAS develop cutaneous melanoma with high frequency and uveal melanoma with a much lower percentage. Larvae showed an increased number of melanocytes already at 3 days post fertilization (dpf) as the earliest evidence of abnormal melanocyte growth. Using this model we performed a chemical screen based on automated detection of a reduction of melanocytes number caused by any of the 1280 FDA or EMA approved drugs of the Prestwick library. The analysis showed that 55 molecules were able to reduce by 60% or more the number of melanocytes per embryo. We identified clotrimazole, as the best candidate. The molecule is an azole derivative acting on the energetic metabolism of melanoma cells. We further tested two compounds for each of the 5 pharmacological classes, and a farnesyltransferase inhibitor (lonafarnib), that inhibits an essential post-translational modification of HRAS and suppresses the hyperpigmentation phenotype. Combinations of clotrimazole and lonafarnib showed the most promising results in zebrafish embryos, allowing a dose reduction of both drugs. We performed validation of these observations in the metastatic human melanoma cell line A375M, and in normal human epithelial melanocytes (NHEM) as control cells, in order to investigate the mechanism of action of clotrimazole in blocking the proliferation of transformed melanocytes. Viability assay and analysis of energy metabolism in clotrimazole treated cells show that this drug specifically affects melanoma cells in vitro and transformed melanocytes in vivo, having no effects on NHEM or wild type larvae. Similar effects were observed with another hit of the antifungal class, miconazole. Furthermore, we show that the effects of clotrimazole are mediated by the inhibition of hexokinase activity and suggest further testing of clotrimazole in combinatorial treatments. In conclusion, this thesis investigated different possibilities of modeling the rare cancer uveal melanoma in zebrafish, using both transgenic and transplantation approaches, and developed a pipeline for a high-throughput, semi-automated chemical screen in a zebrafish melanoma that identified clotrimazole and miconazole as targeting a metabolic vulnerability in melanoma cells.

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Samtani, Grace, and Grace Samtani. "Evaluation of Drug "X" in Preclinical Models of Parkinson's Disease." Thesis, The University of Arizona, 2017. http://hdl.handle.net/10150/625144.

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Parkinson's disease (PD) is a hypokinetic, age-related movement disorder associated with chronic, progressive degeneration of dopaminergic neurons, with cell bodies located in the substantia nigra pars compacta (SNpc) and axon terminals in the striatum. Striatal depletion of the neurotransmitter dopamine (DA) gives rise to the cardinal PD motor symptoms. We utilized a 6-hydroxydopamine (6-OHDA) animal PD model to test the application of a preclinical drug candidate, drug "X", in ameliorating and/or preventing advanced parkinsonism in two studies. First, a neurorestoration study tested the efficacy of drug "X" in restoring motor functionality to animals in which the 6-OHDA lesion had fully developed. Second, a neuroprotection study assessed the effectiveness of drug "X" in preventing the initial development of the 6-OHDA lesion and the onset of motor impairments. Behavioral data indicate that there are no significant differences between the control and drug "X" groups in both studies, suggesting that drug "X" does not improve established severe PD motor deficits nor prevent their initial development. However, we are analyzing brain tissue harvested to 1) verify the extent of the lesion with stereology in the SNpc, 2) evaluate striatal DA levels, and 3) investigate any neuroprotective effects of drug "X" on nigral DAergic neurons.

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Brebner, Karen. "The neurobiology and preclinical evaluation of GABA[subscript B] agonists for the treatment of cocaine addiction." Thesis, National Library of Canada = Bibliothèque nationale du Canada, 2001. http://www.collectionscanada.ca/obj/s4/f2/dsk3/ftp04/NQ60952.pdf.

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Venalis, Paulius. "The performance of antifibrotic agents in preclinical models of systemic sclerosis." Doctoral thesis, Lithuanian Academic Libraries Network (LABT), 2010. http://vddb.laba.lt/obj/LT-eLABa-0001:E.02~2010~D_20101001_150819-57019.

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Systemic sclerosis (SSc) – is one of the most complicated and fatal systemic diseases, and the lack of effective therapy is very evident. The tyrosine kinase inhibitor imatinib mesylate (IM) was shown to inhibit TGF-β and PDGF signaling pathways and prevent the development of dermal fibrosis upon challenge with bleomycin in murine model of SSc. The aim of therapy is not only to stop disease progression, but even induce regression of preexisting fibrosis. On other hand, blocking TGF-β and PDGF signaling in angiogenesis might worsen the vascular manifestations of SSc. We found important to evaluate effectiveness of IM for the treatment of pre-established tissue fibrosis and to exclude that the anti-fibrotic effects of IM are complicated by inhibitory effects on endothelial cell functions. Aim of the study: assess the effect of IM on the process of fibrosis and endothelium in experimental models of systemic sclerosis and cell cultures. Objectives of the study: assess the effectiveness of IM on murine models of established fibrosis; evaluate if IM has an effect on basal functions of endothelial cells; assess effect of IM on the process of angiogenesis. We have shown that IM exerts potent antifibrotic effects in two different models of SSc. Imatinib was effective for prevention of fibrosis and for treatment of established dermal fibrosis. We’ve demonstrated that IM does not inhibit major functions of endothelial cells. Thus, IM might not augment further the preexisting vascular... [to full text]
Sisteminė sklerozė (SSc) – viena sunkiausių ir fatališkiausių autoimuninių sisteminių reumatinių ligų, o bazinių vaistų stygius, šiai ligai gydyti, itin didelis. Į onkologinę klinikinę praktiką įdiegtas tirozinkinazių inhibitorius – imatinibo mezilatas(IM). IM blokuoja TGF-β ir PDGF intraląstelinio signalo perdavimą ir taip sąlygoja fibrozės prevenciją SSc pelių modelyje. Mums buvo svarbu išsiaiškinti, ar imatinibas gali turėti įtakos ne tik prevencijai, bet ir susiformavusiai fibrozei. Be to TGF-β ir PDGF blokavimas angiogenezėje, galėtų riboti daug žadančio fibrozės inhibitoriaus IM naudojimą gydant SSc. Darbo tikslas: įvertinti imatinibo mezilato poveikį fibrozės procesui ir endoteliui sisteminės sklerozės eksperimentiniuose modeliuose ir ląstelių kultūrose. Darbo uždaviniai: įvertinti imatinibo efektyvumą neuždegiminiame SSc modelyje ir patikrinti imatinibo mezilato efektyvumą uždegiminiame suformuotos fibrozės modelyje; ištirti, ar terapinės imatinibo mezilato koncentracijos daro neigiamą poveikį gyvybinėms endotelio funkcijoms; įvertinti imatinibo mezilato poveikį angiogenezės etapams. Mūsų gauti duomenys rodo, kad: IM ne tik sustabdė bet ir paskatino jau egzistuojančios (bleomicino sukeltos) odos firbrozės regresiją; IM ryškiai sumažino poodžio ir odos storį, bei normalizavo miofibroblastų skaičių Tsk-1 pelėse; IM neturėjo poveikio endotelio ląstelių bazinėms funkcijoms; IM neturėjo neigiamo poveikio angiogenezės etapams.

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O'Reilly, Jamie Alexander. "Characterising mismatch negativity biomarker signatures in preclinical models relevant to schizophrenia." Thesis, University of Strathclyde, 2017. http://digitool.lib.strath.ac.uk:80/R/?func=dbin-jump-full&object_id=28635.

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Mismatch negativity (MMN) has been hailed as a 'break-through biomarker in predicting psychosis onset' (Naatanen 2015). This is because deficits have been found in clinical populations diagnosed with psychotic syndromes such as schizophrenia. MMN is an auditory evoked potential (AEP) difference waveform produced by subtracting standard from deviant stimuli AEPs elicited by an oddball paradigm; purportedly arising from any discriminable change in auditory stimulation. Despite nearly four decades of basic research into MMN the underlying mechanisms are not fully understood. Although popular theories suggest that it reflects a sensory-memory trace disruption and/or differential adaptation of responses to standard and deviant/oddball stimuli, there remains considerable debate over the neural mechanism and its interpretation. Nevertheless, associations made between N-methyl-d-aspartate (NMDA) receptors in schizophrenia and findings showing that NMDA receptor antagonists (e.g. ketamine) induce MMN deficits in healthy volunteers suggests abnormal MMNs share common traits and support its use as a biomarker from an electrophysiological perspective. However, this is still speculative and there is great impetus on developing reliable preclinical models of MMN in order to examine the underpinning neurophysiology and therefore its reliance on NMDA receptors as a test of pathology in schizophrenia. A question this thesis aims to address is whether a mismatch response (MMR) exists in rodents which is analogous to the human MMN, and whether its modification by NMDA receptor antagonists or as a result of schizophrenia-related genetic modification sheds light on its utility as a biomarker in disease models of schizophrenia. This thesis describes three experiments performed using mitogen activated protein kinase kinase 7 heterozygous (Map2k7+/-) mice and their wild-type littermates, incorporating NMDA receptor antagonism with ketamine (10 mg/kg i.p.). The MAP2K7 gene is associated with schizophrenia and codes for a post-synaptic intracellular signalling enzyme which is activated following glutamatergic excitation, for instance via NMDA receptors. The MMR to stimuli duration, frequency and intensity changes in oddball paradigms are characterised in urethane-anaesthetised and conscious animals, followed by an examination of laminar auditory cortex activity in response to these physical changes. Data recorded throughout this series of experiments includes cortical electroencephalography (EEG), video footage, and intra-cortical spiking information. These data were then analysed using various time, frequency and time-frequency domain techniques; although mainly focussing on the event-related potential (ERP) approach. Recordings demonstrated substantial differences in the AEP waveform evoked from urethane-anaesthetised and conscious animals, with the latter displaying considerably more dynamic responses, although onset and offset of auditory stimuli induced comparable waveform features in both states. Effects of varying physical properties of stimuli in oddball and control paradigms have been identified as key determinants of the AEP and correspondingly the MMR difference waveform amplitudes. The finding that NMDA receptor disruption in conscious animals by ketamine acutely diminishes a specific AEP feature (≈20-50 ms post stimulus onset) which may impact the resulting MMR tentatively links this study in mice with findings from humans noted above. Ketamine was also found to enhance animal movement and increase EEG spectral power in the 50-70 Hz (gamma-band) frequency range, observed for approximately 10 minutes following drug administration. Both anaesthetised and conscious cohorts of Map2k7+/- mice displayed a significantly enhanced onset response (≈0-20 ms) in the AEP. Interestingly, ketamine did not appear to have a differential effect on Map2k7+/- mice compared with the wild-type group, suggesting that NMDA receptor-mediated neurotransmission is unimpaired in this genetic model relevant to schizophrenia. Overall, the findings suggest that the MMR in mice is fundamentally influenced by the physical properties of stimuli employed; ketamine causes an acute, specific alteration to the AEP in conscious mice in addition to other electrophysiological and behavioural changes; and Map2k7 gene disruption causes a specific and replicable change in AEP amplitude. Overall this study indicates that mouse models are useful for exploring the effects of different pharmacological and genetic manipulations on the auditory evoked response; however, MMN data in clinical cohorts still needs to be interpreted with care. In order to address whether the rodent MMR is analogous to human MMN, it would be necessary to probe how influencing factors revealed in the rodent studies impact on the human response. Whilst the rodent MMR and human MMN show some degree of translation, their potential as schizophrenia biomarkers requires further characterisation and validation.

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Ottaviani, Daniela. "In-Depth Characterization of Human Retinoblastoma Subtype 2 and Preclinical Models." Thesis, Université Paris-Saclay (ComUE), 2019. http://www.theses.fr/2019SACLS001.

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Le rétinoblastome, un cancer pédiatrique de la rétine en développement, est la tumeur intraoculaire la plus fréquente chez l’enfant et représente environ 4 % de tous les cancers infantiles. Bien qu'il s'agisse d'une maladie rare, l'hôpital Curie (centre de référence pour le rétinoblastome en France) accueille environ 50 à 60 nouveaux patients chaque année. Notre groupe a précédemment caractérisé deux sous-types de rétinoblastomes. Les tumeurs de type « cone-like » ou sous-type 1 sont plutôt différenciées et homogènes, présentent une surexpression des gènes liés aux cellules cônes (photorécepteurs) de la rétine, sont diagnostiquées cliniquement plus tôt et regroupent la majorité des formes héréditaires et bilatérales. Les tumeurs « mixed-type » ou sous-type 2, présentent une hétérogénéité intra-tumorale et une surexpression des gènes liés aux cellules des cônes et des cellules ganglionnaires de la rétine, sont enrichies en patients unilatéraux qui sont diagnostiqués cliniquement à des âges plus avancés. Nous avons caractérisé le paysage moléculaire et génomique de 102 rétinoblastomes provenant de trois institutions : l'Institut Curie (France), l'Hôpital Garrahan (Argentine) et l'Hôpital Sant Joan de Déu (Espagne). Le développement d'une signature de méthylation par pyroséquençage pour la classification des échantillons nous a permis d'élargir nos échantillons classés, d'une première série de 72 à notre dernière série de 102 tumeurs. L'analyse du paysage mutationnel de notre série a révélé que les tumeurs du sous-type 2 avaient plus de mutations somatiques par échantillon que les tumeurs du sous-type 1. De plus les gènes BCOR et ARID1A étaient les deux seuls gènes mutés de manière récurrente, et identifiés uniquement dans le sous-type 2. En divisant notre cohorte de tumeurs en sous-type 1 et 2, la distribution des mutations le long de RB1 était significativement différente. Par ailleurs, nous avons identifié une région de la protéine RB1 (dans le Domaine A) enrichie en mutations provenant des tumeurs du sous-type 2, avec très peu de mutations du sous-type 1. En plus, nous avons caractérisé deux événements récurrents de fusion chromosomique perturbant le gène DACH1. Les tumeurs de sous-type 2 sont caractérisées par une surexpression de TFF1, non exprimée dans la rétine normale. L'analyse par immunohistochimie de TFF1 dans des tumeurs localement invasives provenant de l'hôpital Garrahan a révélé la présence de cellules TFF1+ envahissant la région rétrolaminaire du nerf optique. Nous avons exploré un possible rôle oncogène de TFF1 dans le rétinoblastome lié à la survie cellulaire, à la migration cellulaire et à l'invasion cellulaire, qui n'a finalement pas été mis en évidence in vitro. Le sous-type moléculaire 2 regroupe les tumeurs MYCN amplifiées et les tumeurs avec une activation de la voie de signalisation MYC et des gènes cibles de MYC. L'utilisation de JQ1 et OTX015 (inhibiteurs des protéines BET) a fortement réduit la viabilité in vitro de lignées cellulaires de rétinoblastomes représentatives du sous-type 2, avec une régulation négative significative du gène et de la protéine MYC/MYCN. Nos résultats préliminaires suggèrent une nouvelle piste thérapeutique par l'inhibition des protéines BET dans le rétinoblastome. Les modèles précliniques largement utilisés dans la recherche sur le rétinoblastome n'ont pas été caractérisés ou classés au niveau moléculaire. Nous avons utilisé la même approche que pour la classification des tumeurs primaires et avons constaté que la plupart des modèles cellulaires et PDX étudiés étaient classés dans le sous-type moléculaire 2 et partageaient des caractéristiques moléculaires, génomiques et protéiques trouvés dans les tumeurs primaires de ce sous-type moléculaire. En conclusion, nous avons pu caractériser de façon plus approfondie le sous-type 2 des rétinoblastomes, qui semble présenter un phénotype plus agressif et qui est le sous-type représenté dans les modèles précliniques analysés
Retinoblastoma (RB) is a rare pediatric cancer of the developing retina that represents the most common intraocular tumor in children, and accounts for about 4% of all childhood cancers. Although being a rare disease, the Curie Hospital (the referral center for retinoblastoma in France) treats about 50-60 new patients each year. Our group has previously characterized two retinoblastoma subtypes. The cone-like or subtype 1 tumors rather differentiated and homogenous, presenting an overexpression of genes related to cone photoreceptor retinal cells, clinically diagnosed earlier and grouping the majority of hereditary and bilateral forms. The mixed-type or subtype 2 tumors, displaying an intra-tumoral heterogeneity and showing overexpression of genes related to cone and retinal ganglion cells, are enriched in unilateral patients clinically diagnosed at older ages. The general goal of my thesis was to extend the molecular characterization of these subtype 2 retinoblastomas. We characterized the molecular and genomic landscape of retinoblastoma in a series of 102 primary tumors, integrating samples from three institutions: the Curie Institute (France), the Garrahan Hospital (Argentina) and Sant Joan de Déu Hospital (Spain). The development of a pyrosequencing-based tool for sample classification allowed us to enlarge our classed samples, from an initial series of 72, to our final series of 102 tumors. Analysis of the mutational landscape in our series revealed that tumors from the subtype 2 had significantly more somatic mutations per sample than tumors from the subtype 1. Besides RB1 gene, BCOR and ARID1A where the only two recurrently mutated genes, and identified only in the subtype 2. Distribution of mutations alongside the RB1 gene has so far been analyzed in terms of a single group of retinoblastomas. When splitting our cohort in subtype 1 and subtype 2 tumors, the distribution of mutations was significantly different. Besides, we identified a region of the RB1 protein (in Domain A) enriched in mutations from tumors of the subtype 2, and devoid of mutations of the subtype 1. Besides somatic mutations, we characterized two recurrent chromosomal fusion events disrupting DACH1. Subtype 2 tumors are characterized by an overexpression of TFF1, not expressed in the normal retina. Immunohistochemical analysis of TFF1 in locally invasive tumors coming from the Garrahan Hospital revealed the presence of TFF1+ cells invading the retrolaminar region of the optic nerve. We then explored a possible oncogenic role of TFF1 in retinoblastoma related to cell survival, cell migration and cell invasion, which was not fully uncovered. Molecular subtype 2 regroups the MYCN amplified tumors and tumors with MYC signaling pathway activation and upregulation of hallmark MYC target genes. The use of JQ1 and OTX015 (BET bromodomains inhibitors) strongly reduced the viability in vitro of retinoblastoma cell lines representatives of the subtype 2, together with a significant MYC/MYCN gene and protein downregulation. We provided preliminary results to explore a new therapeutic avenue of BET protein inhibition in retinoblastoma. Preclinical models widely used in retinoblastoma research has not been characterized or classified at the molecular level. We have used the same approach as for primary human tumor’s classification, and found that most cellular and PDX models studied classed in the molecular subtype 2 and shared many of the molecular, genomic and protein characteristics found in primary tumors of this molecular subtype. Taken together, we have performed a deeper characterization of subtype 2 retinoblastomas, which seems to represent a more aggressive phenotype, and is the represented subtype in the preclinical models analyzed

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15

George, Courtney M. "Medulloblastoma: New animal models, preclinical drug testing, and characterising immune infiltrates." Thesis, Edith Cowan University, Research Online, Perth, Western Australia, 2022. https://ro.ecu.edu.au/theses/2575.

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Medulloblastoma is the most common malignant brain tumour in children. The current treatment for medulloblastoma consists of surgery, radiation, and chemotherapy. Although these therapies have their merits, the outcome for some patients, particularly those with MYC amplified tumours, is poor, and the damaging nature of these therapies results in morbidities that significantly impact on a patient’s quality of life. To improve outcome and reduce adverse side effects, several strategies have been employed, including expanding the repertoire of accurate disease models, the development of novel therapies and the initiation of clinical trials, and an improvement in the understanding of the disease pathogenesis. The purpose of this thesis was to contribute to the repertoire of animal models, assist in identifying novel therapeutic approaches to treatment, and to advance the knowledge of the medulloblastoma immune microenvironment. We did so by aiming to (1) develop more accurate, immune-competent animal models of MYC- or NMYC-amplified medulloblastoma, (2) testing a novel treatment that combines conventional chemotherapies with a cell cycle checkpoint kinase inhibitor, and (3) investigating the effects of clinically used chemotherapies on immune cell populations in the brains of medulloblastoma-bearing mice. Currently, the limited availability of preclinical mouse models that accurately represent subgroup-specific medulloblastoma has hindered the development of therapies that succeed in the clinical setting. In addition, the distinct lack of immunologically competent models has prevented the advancement of immunotherapy drugs in treating medulloblastoma. The models of medulloblastoma developed here were designed to aid in preclinical studies, and to contribute specifically to the repertoire of immune competent mouse models for studies to clarify the role of the immune system in medulloblastoma. Here, we utilised mutated human variants of CMYC, NMYC, and P53, to transform mouse neural stem cells into tumour forming cells. Tumours were established in C57Bl/6 mice and characterised by histology and RNAseq analysis. Whilst these animal models could not be confirmed to accurately represent Group 3 or Group 4 medulloblastomas, this study demonstrated that mouse neural stem cells could be transformed using human genes and could generate brain tumours within immune competent hosts. As we were unable to conclusively define the exact nature of the tumours that were produced here, existing mouse models were used for subsequent chapters in this thesis. Prior to implementing new therapeutic agents into clinical trial, these are evaluated in a pre-clinical setting. The data presented in chapter two contributed to a larger scale pre-clinical testing pipeline that led to the identification and evaluation of multiple kinase inhibitors for the treatment of medulloblastoma. Here, I examined the combination of the cytotoxic agent gemcitabine (GEM) with the cell cycle checkpoint kinase inhibitor (LY2606368, prexasertib) as a novel approach in the treatment of Group 3 medulloblastoma. Combination GEM/LY2606368 treatment improved the survival of mice with aggressive medulloblastoma. Using immunoblotting and flow cytometry I showed that mechanistically LY2606368 enhanced GEM-induced cytotoxicity by impairing DNA damage response pathway activity, which promoted the accumulation of DNA damage leading to increased apoptosis. Together, these data formed part of the preclinical evidence that supported the establishment of the SJ-ELiOT clinical trial, targeted towards improving outcomes for patients who experience recurrent or relapsed disease following standard-of-care therapy. There is evidence to suggest that inhibiting the DNA damage response pathway can stimulate the immune system, and aid in tumour elimination. This provides strong rationale for implementing immunotherapies for patients who are predicted to have a poor response to conventional treatments. Unfortunately, to date all clinical trials investigating current popular immune-based therapies have failed in medulloblastoma, likely due to a poor understanding of the immune microenvironment in this disease. This presented an opportunity to improve our understanding of the effects of treatment on the immune system in brain. Here I characterised the immune cell populations in the brains of mice with Group 3 medulloblastoma treated with vehicle or two clinically-used chemotherapies – cyclophosphamide (CPA) and GEM. I revealed that CPA and GEM differentially alter immune cells within the brain in a manner similar to that observed outside of the central nervous system. I also demonstrate that the lack of an adaptive immune system (using mice deficient in Rag1) does not influence the anti-cancer effects of these drugs. This information provides a rationale for exploring alternative avenues when considering the use of cancer-targeting immunotherapies in combination with conventional medulloblastoma treatments. Collectively, these studies demonstrate the complicated nature of modelling high-risk medulloblastoma in the lab. I have improved upon current preclinical tools for medulloblastoma by the development of accurate immune competent models and advanced our knowledge of disease pathogenesis by elucidating the way medulloblastomas interact with the immune system in the brain. Furthermore, I highlight the translational value of preclinical models in the evaluation of new drug combinations ahead of clinical trial. Improving on the current tools available and accurately evaluating new therapies will ideally lead to improved clinical outcomes for patients with medulloblastoma.

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Crouch, Barry. "Cognitive dysfunction in schizophrenia : novel models and behavioural methods for preclinical research." Thesis, University of Aberdeen, 2015. http://digitool.abdn.ac.uk:80/webclient/DeliveryManager?pid=229384.

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Thirtamara, Rajamani Keerthi Krishnan. "Animal Models of Drug Addiction and Autism Spectrum Disorders." The Ohio State University, 2013. http://rave.ohiolink.edu/etdc/view?acc_num=osu1386011455.

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García, Parra Jetzabel 1983. "PARP1 expression in breast cancer and effects of its inhibition in preclinical models." Doctoral thesis, Universitat Pompeu Fabra, 2012. http://hdl.handle.net/10803/84173.

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Breast cancer is the main cause of cancer death in women. Improved treatments, prevention programs and earlier detection are reducing the rate of death; however, there is still a high percentage of mortality by this cancer. Identification of novel targets to predict response to specific treatments is a key goal for personalizing breast cancer therapy and to improve survival. Few years ago, PARP inhibitors appeared as a promising therapy, particularly in BRCA-mutated cancers. However, there was a clear need to conduct further preclinical and translational work to improve the rational development of PARP inhibition in breast cancer. In this work we described PARP1 expression in breast tumour samples and characterized the effects of its inhibition in preclinical models. We found that nuclear PARP1 protein overexpression was associated with malignant transformation and poor prognosis in breast cancer. PARP1 overexpression was more common in triple negative subtype, but was also detectable in small subsets of estrogen receptor positive and HER2 positive breast cancers. In preclinical models, PARP1 played distinct roles in different molecular subtypes of breast cancer. Moreover, we described that olaparib (novel PARP inhibitor) had antitumour effects in different breast cancer subtypes, and its combination with trastuzumab (anti-HER2 antibody) enhanced the antitumour effects of this therapy.
El càncer de mama és la principal causa de mort per càncer en dones. La millora dels tractaments i la detecció precoç estan reduint la taxa de mort, però segueix sent elevada. Identificar noves dianes per predir la resposta a tractaments és clau per millorar les teràpies contra aquest càncer i la supervivència. Els inhibidors de PARP van aparèixer com una teràpia prometedora, particularment en càncers BRCA-mutants, però, cal dur a terme més estudis preclínics i translacionals per fomentar un desenvolupament racional d’aquesta teràpia en càncer de mama. Aquest treball descriu l’expressió de PARP1 en mostres de tumors mamaris i caracteritza els efectes de la seva inhibició a models preclínics. Vam observar que la sobreexpressió nuclear de la proteïna PARP1 fou associada amb: la transformació maligna; mal pronòstic en càncer de mama; i fou més freqüent al subtipus triple-negatiu, però també es va detectar en un subgrup de càncers de mama receptors d’estrogen positius i HER2 positius. En models preclínics, PARP1 va exercir rols diferents als diferents subtipus de càncer de mama. Per altra banda, vam descriure que olaparib (inhibidor de PARP) té efectes antitumorals en els diversos subtipus, i combinat amb trastuzumab (anticòs anti-HER2) potencia els efectes antitumorals d’aquesta teràpia.

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Cantelli, Erika <1981&gt. "Preclinical neuroblastoma models for a pharmacological study of a new MYCN oncogene inhibitor." Doctoral thesis, Alma Mater Studiorum - Università di Bologna, 2011. http://amsdottorato.unibo.it/4068/.

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Background. Neuroblastoma is the most deadly solid tumor of childhood. In the 25% of cases it is associated with MYCN amplification (MA), resulting in the disregulation of several genes involved in cancer progression, chemotherapy resistance and poor prognosis causing the disregulation of several genes involved in cancer progression and chemotherapy resistance and resulting in a poor prognosis. Moreover, in this contest, therapy-related p53 mutations are frequently found in relapsed cases conferring an even stronger aggressiveness. For this reason, the actual therapy requires new antitumor molecules. Therefore, rapid, accurate, and reproducible preclinical models are needed to evaluate the evolution of the different subtypes and the efficacy of new pharmacological strategies. Procedures. We report the real-time tumorigenesis of MA Neuroblastoma mouse models: transgenic TH-MYCN mice and orthotopic xenograft models with either p53wt or p53mut, by non-invasive micro PET and bioluminescent imaging, respectively. Characterization of MYCN amplification and expression was performed on every collected sample. We tested the efficacy of a new MYCN inhibitor in vitro and in vivo. Results. MicroPET in TH-MYCN mice permitted the identification of Neuroblastoma at an early stage and offered a sensitive method to follow metabolic progression of tumors. The MA orthotopic model harboring multitherapy-related p53 mutations showed a shorter latency and progression and a stronger aggressiveness respect to the p53wt model. The presence of MA and overexpression was confirmed in each model and we saw a better survival in the TH-MYCN homozigous mice treated with the inhibitor. Conclusions. The mouse models obtained show characteristics of non-invasiveness, rapidity and sensitivity that make them suitable for the in vivo preclinical study of MA-NB. In particular, our firstly reported p53mut BLI xenograft orthotopic mouse model offers the possibility to evaluate the role of multitherapy-related p53 mutations and to validate new p53 independent therapies for this highly aggressive Neuroblastoma subtype. Moreover, we have shown potential clinical suitability of an antigene strategy through its cellular and molecular activity, ability to specifically inhibit transcription and in vivo efficacy with no evidence of toxicity.

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Jawad, Dhafer Sahib. "Chemopreventive effect of resveratrol in preclinical colorectal cancer models with different genetic drivers." Thesis, University of Leicester, 2017. http://hdl.handle.net/2381/40308.

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Previous reports indicate resveratrol can modulate many processes and targets in colorectal cancer (CRC) cells and rodent models but it is uncertain whether these effects translate to humans in vivo. The lack of relevant experimental models, particularly premalignant colorectal cells, the major targets for prevention, is a significant obstacle to translation of effective preventive agents to the clinic. Development of improved 3-dimensional organoid models with discrete mutational drivers (Apc, Apc+Kras and Braf) representing different subtypes of early CRC is an aim of this thesis. A further aim is to evaluate resveratrol across these models and examine in greater detail the ability of resveratrol to interfere with the development of cancers driven by mutant BrafV600E. Organoid cultures using intestinal cells isolated from genetically engineered mice and human cancer/adenoma tissue were successfully established and conditions optimised to allow long-term maintenance. Notably, the organoids derived from adenomas arising in Villin-Cre/BrafV600E mice constitute a novel preclinical model of CRC. After first demonstrating that resveratrol had anti-proliferative activity across a panel of human CRC cell lines with different mutation profiles it was tested in the organoids at clinically-achievable concentrations for effects on autophagy, senescence, apoptosis and stem cells. Administration of a high-fat diet to Villin-Cre/BrafV600E mice enhanced Braf-induced cryptal hyperplasia in a short-term study and this was significantly reduced by resveratrol. In a follow-up survival study, high dose resveratrol greatly prolonged the lifespan of Villin-Cre/BrafV600E mice on high-fat diet. However, resveratrol had no effect on the survival of mice given standard diet. Overall, these findings suggest a specific interaction between mutant Braf expressed in mouse intestine and high-fat, and that the effects are blocked by high dose resveratrol. Future studies are needed to investigate underlying mechanisms. Results suggest resveratrol may have value in the prevention of colorectal adenomas/cancers with different genetic alterations, including mutant BrafV600E.

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Gronbach, Leonie [Verfasser]. "Organotypic head and neck cancer models for advanced preclinical drug testing / Leonie Gronbach." Berlin : Freie Universität Berlin, 2021. http://d-nb.info/1230407316/34.

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Bertalan, Gergely [Verfasser]. "Quantitative tissue characterization by mechanical parameters in preclinical, small-animal models / Gergely Bertalan." Berlin : Medizinische Fakultät Charité - Universitätsmedizin Berlin, 2020. http://d-nb.info/1218076739/34.

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Marston, Gemma. "Anatomical and vascular imaging with high frequency ultrasound in preclinical models of colorectal cancer." Thesis, University of Leeds, 2012. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.598038.

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Colorectal cancer (CRC) is one of the most common malignancies in the western world and is the third most common cause of cancer related deaths in the UK. It has a good 5 year survival rate if the cancer is diagnosed at an early stage - 93% for Dukes' stage A; however, for late stage metastatic disease, this falls to only 7%. Improvements in treatment for metastatic disease, detection and screening are required to combat these low survival rates. Preclinical models, and in particular mouse models, are invaluable for studying the mechanisms which initiate tumour development and progression. Many models of CRC exist, including models of hereditary and sporadic CRC, including human CRC cell xenografts which support rapid tumour growth, allowing for relatively fast, short term studies and investigations into novel therapeutics. The ApcMin/+ mouse model recapitulates the early stages of human CRC development, with many polyps developing throughout the small intestine and colon. This project investigated the use of non-invasive imaging with high frequency ultrasound to provide anatomical information on the abdomen, and specifically the colon, of C57BI/6J mice, and the reproducibility of a protocol to measure the colon wall thickness in vivo. Once established, these techniques were then used to detect polyps in aged (170±34 days) and 90 day old ApcMin/+. mice, showing that this technique could be used with sensitivity and specificity of 48% and 100%, respectively. This project also investigated the feasibility of using contrast enhanced high frequency ultrasound to monitor the growth of human CRC cell xenografts and qualitatively and quantitatively assess their longitudinal vascular development in vivo. These data were then used as the basis of an intervention study, which incorporated the vascular disruptive agent Combretastatin A-4 into the CE-HFUS imaging protocols in order to assess the immediate vascular kinetic impact of CA4. This showed that CE-HFUS can detect and quantitate changes in tumour vascular kinetics in vivo following treatment with a known agent, suggesting that it may be useful in preclinical drug development. The work in this Thesis has demonstrated that C57BI/6 mouse colon wall thickness can be accurately determined in vivo, in an operator independent manner. Following on from the characterisation of colon wall thickness in wild type mice; this Thesis has shown for the first time that it is possible to identify colonic polyps in ApcMin/+ mice in vivo in both 90 day old and older age mice. Work in this Thesis has also characterised i for the first time with CE HFUS the blood flow kinetics and vascular perfusion of HCT116 CRC xenograft tumours in a longitudinal study. Building on this work, this Thesis has shown for the first time disruption in blood flow kinetics to HCT116 CRC xenograft tumours after exposure to CA4.

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Hvoslef-Eide, Martha. "Characterising transgenic APP mutation mouse models of amyloid pathology for use in preclinical immunotherapy." Thesis, Cardiff University, 2012. http://orca.cf.ac.uk/43321/.

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No disease-modifying compounds are available to halt disease progression in Alzheimer’s disease (AD). Immunotherapy offers promising possibilities for the manipulation of Aβ levels which the amyloid cascade hypothesis proposes as the causative factor in AD. However, antiAβ antibodies have caused inflammation in vivo. An alternative antibody (2B3) which targets the β-secretase cleavage site of the amyloid precursor protein (APP) from which Aβ is cleaved has been shown to downregulate Aβ in human cell lines. The approach is thought unlikely to cause inflammation as the immune system is not relied upon for Aβ clearance. It was hypothesised that the administration of 2B3 to aged transgenic APP mutation mice would lower Aβ levels through the inhibition of Aβ production, with an associated lowering of cognitive deficits. Two murine APP mutation models [London APP(V717I) and Indiana PDAPP(V717F)] were characterised in order to identify cognitive deficits against which the ability of 2B3 to reduce deficits could be assessed. APP(V717I) mice were assessed in the marble burying task, the elevated plus maze and a foraging task assessing spatial working memory at 3, 6 and 19 months of age. The radial arm water maze was carried out at 10 months, before the T-maze non-matching to position task was administered at 11 months. Object recognition memory was assessed at 18 months. Similarly, PDAPP mice were assessed using the marble burying and elevated plus maze (9.5months), the T-maze (9-10 months), the foraging task (11 and 14 months) and the object recognition task (12 months). Whilst aged transgenic PDAPP mice displayed disrupted spatial working memory, no evidence of agerelated cognitive decline was observed in APP(V717I) transgenic mice despite increases in Aβ pathology with age. 2B3 did not alter Aβ levels or spatial working memory in PDAPP(V717F) mice in a pilot study, whilst the in vitro downregulation of Aβ was successfully replicated in primary murine neurons. The findings indicate that transgenic models of neurodegenerative disease require thorough characterisation to optimise their use in pre-clinical research. Furthermore, the use of alternative immunotherapy in the treatment of AD remains a promising, but early stage avenue of study.

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Bonnycastle, Katherine. "Synaptic vesicle recycling in preclinical models of intellectual disability, autism spectrum disorder and epilepsy." Thesis, University of Edinburgh, 2018. http://hdl.handle.net/1842/31344.

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The development of the central nervous system is dysregulated in neurodevelopmental disorders such as intellectual disability, autism spectrum disorder, and epilepsy. These three disorders have different clinical features, yet there is high comorbidity between them. They can be difficult to study due to their highly complex aetiologies, however there are various monogenic diseases that can cause all of them, including SYNGAP1 haploinsufficiency where the synaptic guanosine triphosphatase (GTPase)-activating protein (SYNGAP) protein levels are highly reduced; Fragile X syndrome where the fragile X mental retardation protein (FMRP) is no longer translated; and DNM1 epileptic encephalopathy where mutations in the Dynamin1 gene alter the protein function. These monogenic conditions are synaptopathies as the proteins affected play important roles in synapse stability and neurotransmission. Because of the high comorbidity between these disorders, it is hypothesised that there may be a common mechanism underlying them. We hypothesise that a deficit in presynaptic vesicle recycling may be part of a common mechanism underlying intellectual disability, autism spectrum disorder, and epilepsy especially in SYNGAP1 haploinsufficiency, Fragile X syndrome, and DNM1 epileptic encephalopathy. Using various fluorescent presynaptic activity reporters including synaptic pHluorins, tetramethylrhodamine dextran and calcium dyes to compare presynaptic activity in in vitro models of these monogenic conditions, we found differences in synaptic vesicle (SV) endocytosis in the genetically altered conditions compared to wildtype controls. We observed various SV endocytosis defects in clathrin-mediated endocytosis (CME) or activity-dependent bulk endocytosis (ADBE) in our models. We observed enhanced CME in SynGAP1 KO mouse hippocampal neurons. This enhanced SV endocytosis was accompanied by decreased SV cargo on the plasma membrane. Rat SynGAP1 KO hippocampal neurons did not display enhanced SV endocytosis, nor did neurons with the GTPase-activating (GAP) domain of SynGAP deleted. This was perhaps due to the altered time course of development between these rodent species. In mouse and rat models of Fragile X syndrome, CME was not altered compared to wildtype controls. However, in a rat model, we observed fewer nerve terminals undergoing ADBE which is the dominant SV endocytosis mode during elevated neuronal activity. De novo epileptic encephalopathy-associated mutations in DNM1 had differential effects on SV recycling through both CME and ADBE. Mouse hippocampal neurons overexpressing Dyn1R237W, Dyn1I289F and Dyn1H396D all showed less CME compared to overexpression of Dyn1WT. Moreover, fewer nerve terminals overexpressing Dyn1H396D were found to undergo ADBE. We also found that a large-conductance potassium (BK) channel opener can accelerate clathrin-mediated endocytosis and thus may be able to rescue the impaired SV endocytosis caused by these mutants. Although there is not yet a common underlying pathway at the presynaptic level between these conditions, SV recycling dysfunction is present across all of these models. Furthermore, we propose an axis of pathophysiology model where optimal SV endocytosis is required for optimised neural performance. We propose that either decreased or increased SV endocytosis can lead to the synaptic dysfunction observed in these models.

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Smolinski, Justin Bruce. "Dietary Chemoprevention Studies in Preclinical Models of Prostate Cancer: Bioactive Lipids and Vitamin D." The Ohio State University, 2010. http://rave.ohiolink.edu/etdc/view?acc_num=osu1282069758.

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Özgür, Günes Yasemin. "Preclinical gene therapy using recombinant AAV vectors in mouse models of two human diseases." Thesis, université Paris-Saclay, 2022. http://www.theses.fr/2022UPASL092.

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Nous avons obtenu une preuve de concept pour la thérapie génique de 2 maladies génétiques perte-de-fonction.L'acrodysostose est une maladie osseuse et rénale causée par des mutations de la sous-unité régulatrice de la protéine kinase A (PRKAR1A). Nous avons testé les effets d'un rAAV9-CAG-humanPRKR1A dans un modèle murin KI. hPRKAR1A a été exprimée dans les chondrocytes de la plaque de croissance et les cellules tubulaires rénales. L'architecture des chondrocytes et la longueur du squelette ont été améliorées.L'AMN est une axonopathie tardive de la moelle épinière causée par des mutations du gène ABCD1.Nous avons construit et développé un vecteur, rAAV9- MAG-humanABCD1-HA (hABCD1), et testé ses effets dans un modèle murin KO de la maladie.L'expression de hABCD1-HA a été observée dans les oligodendrocytes (OL) et les astrocytes (A). Nous avons développé un vecteur ciblant les oligodendrocytes (OL), rAAV9-MAG-humanABCD1-HA. Les déficits neurologiques ont été prévenus avec un suivi de 2 ans.C26:0-lysoPC (VLCFA) était diminué dans la moelle épinière des souris traitées.Chez le primate, l'injection intrathécale du vecteur rAAV9-MAG a induit un niveau élevé d'expression de hABCD1-HA dans les OL et les A de la moelle épinière et du cervelet. Le ciblage OL n'avait pas été obtenu auparavant chez les primates avec d'autres vecteurs ou promoteurs, ce qui ouvre la porte à l'application humaine de notre vecteur dans diverses maladies du système nerveux central (SNC)
We have obtained proof-of concept for the gene therapy of two diseases.Acrodysostosis is a bone and kidney disease caused by loss-of-function mutations in the regulatory subunit of protein kinase A (PRKAR1A). We tested the effects of a rAAV9-CAG-humanPRKR1A in a knock-in mouse model. hPRKAR1A expression was found in growth plate chondrocytes, and kidney tubular cells. Chondrocyte architecture and skeleton length were improved.X-ALD AMN is a late-onset axonopathy of spinal cord caused by ABCD1 mutations. We made an original rAAV9-MAG-humanABCD1-HA (hABCD1) vector and tested its effects in a KO mouse model.hABCD1-HA expression was observed in numerous OL and astrocytes. Neurological deficits were prevented 24 months after injection. C26:0-lysoPC (VLCFA), was lower in spinal cord.In non-human primate, intrathecal injection of the rAAV9-MAG vector induced high hABCD1-HA expression in OL and astrocytes of spinal cord and cerebellum. OL targeting has not been obtained before in primates with other vectors or promoters. This opens the door to the human application of OL targeting in a number of CNS diseases

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Chen, Liu Qi. "Development and Application of AcidoCEST MRI for Evaluating Tumor Acidosis in Pre-Clinical Cancer Models." Diss., The University of Arizona, 2014. http://hdl.handle.net/10150/323450.

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Tumor acidosis is an important biomarker in cancer. We have developed a noninvasive imaging method, termed acidosis Chemical Exchange Saturation Transfer (acidoCEST) MRI to measure extracellular pH (pHe) in the tumor microenvironment. Chapter 1 introduces the importance of measuring tumor acidosis and presents various imaging modalities and their shortcoming to measure pHe. Chapter 2 describes the optimization of acidoCEST MRI for in vivo pHe measurement. The acidoCEST MRI protocol consists of a CEST-FISP acquisition and Lorentzian line shape fittings. We determined the optimal saturation time, saturation power and bandwidth, 5 sec, 2.8 µT and 90 Hz respectively. We also tried various routes of administration to increase contrast agent uptake in the tumor. We decided upon 200 µL bolus followed by 150 µL/hr infusion. The optimized acidoCEST MRI protocol was tested on a mammary carcinoma mouse model of MDA- MB-231. Our method can detect an increase in pHe in the bladder and tumor of the mice treated with bicarbonate. We used this optimized acidoCEST MRI method to measure pHe in lymphoma tumor model of Raji, Ramos and Granta 519 as described in Chapter 3. Pixel-wise pHe maps showed tumor heterogeneity. The pHe of Raji, Ramos and Granta 519 were determined to be mildly acidic with no significant difference. Chapter 4 describes the evolution of pixel-wise analysis in more detail. Besides the pHe map and spatial heterogeneity, we were able to determine the % contrast agent uptake. We monitored these biomarkers in two different mammary carcinoma mouse models, MDA- MB-231 and MCF-7 longitudinally and made comparisons between the different tumor models: MCF-7 were more acidic, more heterogeneous and faster growing than MDA- MB-231.

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Ramirez, Francisco Daniel. "Methodological Rigour in Preclinical Research: Implications for its Scientific Validity and Biomedical Progress." Thesis, Université d'Ottawa / University of Ottawa, 2019. http://hdl.handle.net/10393/39426.

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Preclinical research using animals often precedes and informs clinical trials; however, most attempts to translate findings from “bench-to-bedside” fail. There is growing concern that an important cause of failed translations is that much of preclinical research is not reproducible, with poor experimental methodology believed to be a major contributor. Four studies were conducted: (1) an assessment of reported study designs of preclinical experiments published in leading cardiovascular journals; (2) an examination of sex bias in preclinical cardiovascular research; (3) a comparison of experimental practices between male and female preclinical cardiovascular researchers; and (4) an analysis of the influence of journal initiatives on preclinical research quality. These studies suggest that (1) methodological shortcomings are prevalent and persistent in preclinical cardiovascular research; (2) women’s involvement in preclinical cardiovascular research is positively associated with considering sex as a biological variable; and (3) journals can exert considerable influence on the quality of published data.

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McAllister, Kathryn Anne Lucretia. "Development and validation of touchscreen automated tasks to assess cognition in preclinical models of schizophrenia." Thesis, University of Cambridge, 2012. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.610570.

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Denton, Nicholas Lee Denton. "Modulation of tumor associated macrophages enhances oncolytic herpes virotherapy in preclinical models of Ewing sarcoma." The Ohio State University, 2018. http://rave.ohiolink.edu/etdc/view?acc_num=osu1523892800897524.

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Lahr, Christoph Alexander. "Tissue-engineering humanised bone sarcoma models in rodents-a preclinical study platform for orthopaedic research." Thesis, Queensland University of Technology, 2021. https://eprints.qut.edu.au/207759/1/Christoph%20Alexander_Lahr_Thesis.pdf.

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This thesis is a step forward in preclinical in-vivo disease modelling, designed to find new diagnostic and therapeutic options, to ultimately improve the poor outcome of patients with primary bone cancer. Combining the principles of tissue-engineering, 3D-printing and advanced gene editing techniques the preclinical animal models developed in this thesis have important clinical implications that could shape future innovative treatment plans. Particularly the translation of a humanised osteosarcoma model from a mouse into a newly engineered severely immunocompromised rat will facilitate preclinical primary bone cancer research by opening up new experimental avenues for complex surgical resection and reconstruction models.

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Farr, Christina. "A Caulobacter crescentus microbicide prevents vaginal infection with HIV-1 and HSV-2 in preclinical models." Thesis, University of British Columbia, 2016. http://hdl.handle.net/2429/57896.

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Over 2 million people are infected with HIV each year. The majority of these infections occur in women residing in low-income countries, where their access to and control over preventative measures is often limited. This suggests that female-controlled prevention options for HIV-1 are urgently needed. Microbicides, which can be topically applied to the vaginal tract in advance of sexual activity to protect from HIV-1 infection, represent a promising female-controlled prevention option. We have investigated the development of an HIV-1 specific microbicide using the non-pathogenic, freshwater bacterium Caulobacter crescentus. C. crescentus contains a Surface or S-layer that is easily modified for high-density display of recombinant proteins. We have developed 18 recombinant C. crescentus that display anti-HIV proteins, including decoy receptors and ligands, anti-viral lectins and fusion inhibitors, that can prevent various steps of the HIV-1 attachment and entry process. In vitro testing with these recombinant C. crescentus indicated that 15 were able to provide substantial protection from HIV-1 infection. Studies with immune-competent mice demonstrated that application of C. crescentus to the vaginal tract does not induce the production of inflammatory cytokines or recruitment of immune cells. To test for protection against HIV-1 in vivo we have combined the implantation of human fetal liver and thymus tissue with the intravenous injection of autologous CD34+ cells into NOD-scid IL2Rγnull mice to create humanized Bone Marrow-Liver-Thymus (BLT) mice and we have demonstrated that the peripheral blood of these mice contains human CD45+ cells, including CD4+ and CD8+ T cells, B cells, myeloid cells, and NK cells. Our data indicates that vaginal application of recombinant C. crescentus at the time of HIV-1JR-CSF infection provides protection from HIV-1 infection. Seven of the recombinant C. crescentus were predicted to also prevent infection with herpes simplex virus 2 (HSV-2). HSV-2 is a major co-morbidity for HIV-1 infection, contributing to a 2-4 fold increase in acquisition. Four recombinant C. crescentus provided significant protection from HSV-2 infection in vivo. Taken together this data suggests that a C. crescentus based microbicide could be a safe and effective method to prevent infection with HIV-1 and HSV-2, having considerable impact on public health.
Science, Faculty of
Microbiology and Immunology, Department of
Graduate

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Salvati, Roberto. "Development of Magnetic Resonance Imaging (MRI) methods for in vivo quantification of lipids in preclinical models." Thesis, Rennes 1, 2015. http://www.theses.fr/2015REN1B026/document.

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L'obésité est associée à une augmentation de la morbidité et de la mortalité liée à de nombreuses maladies, y compris le diabète de type 2, l'hypertension et des pathologies hépatiques menant à une surcharge lipidique d’origine non alcoolique. Récemment, l’imagerie par résonance magnétique (IRM) est devenue la méthode de choix pour la quantification non invasive de la graisse. Dans cette thèse, les méthodes d'IRM ont été étudiées sur un scanner préclinique de 4.7T in vitro (fantômes MR) et in vivo (souris). Deux algorithmes de quantifications de la graisse -la méthode de Dixon et l’algorithme IDEAL- ont été considérés. Les performances de l'algorithme IDEAL ont été analysées en fonction de propriétés des tissus (T2*, fraction de graisse et modèle spectral de la graisse), de paramètres d'acquisition IRM (temps d’écho, nombre d'échos) et de paramètres expérimentaux (SNR et carte de champ). Sur les fantômes, l'approche standard single-T2* IDEAL a montré certaines limites qui pourraient être surmontées en optimisant le nombre d'échos. Une nouvelle méthode, pour déterminer les valeurs de vérité terrain pour T2* de l'eau et pour T2* de la graisse, a été proposée. Pour les mesures in vivo, différentes analyses ont été effectuées en utilisant l'algorithme IDEAL sur le foie et les muscles. L'analyse statistique sur les mesures de ROI a montré que le choix optimal du nombre d'échos est égal à trois pour la quantification de la graisse et six ou plus pour la quantification du T2*. Les valeurs de la fraction de graisse, calculées avec l'algorithme IDEAL, étaient statistiquement comparables aux valeurs obtenues avec la méthode de Dixon. Enfin, un procédé pour générer des signaux de référence mimant les systèmes eau-graisse (Fat Virtual Phantom MRI), sans l'aide d'objets physiques, a été proposé. Ces fantômes virtuels, qui présentent des caractéristiques de bruit réalistes, représentent une alternative intéressante aux fantômes physiques pour fournir un signal de référence dans les mesures IRM
Obesity is associated with increased morbidity and mortality linked to many diseases, including type 2 diabetes, hypertension and disease nonalcoholic fatty liver. Recently, 1H magnetic resonance imaging (MRI) has emerged as the method of choice for non-invasive fat quantification. In this thesis, MRI methodologies were investigated for in vitro (MR phantoms) and in vivo (mice) measurements on a 4.7T preclinical scanner. Two algorithms of fat quantifications – the Dixon’s method and IDEAL algorithm – were considered. The performances of the IDEAL algorithm were analyzed as a function of tissue properties (T2*, fat fraction and fat spectral model), MRI acquisition parameters (echo times, number of echoes) and experimental parameters (SNR and field map). In phantoms, the standard approach of single-T2* IDEAL showed some limitations that could be overcome by optimizing the number of echoes. A novel method to determine the ground truth values of T2* of water and T2* of fat was here proposed. For in vivo measurements, different analyses were performed using the IDEAL algorithm in liver and muscle. Statistical analysis on ROI measurements showed that the optimal choice of the number of echoes was equal to three for fat quantification and six or more for T2* quantification. The fat fraction values, calculated with IDEAL algorithm, were statistically similar to the values obtained with Dixon’s method. Finally, a method for generating reference signals mimicking fat-water systems (Fat Virtual Phantom MRI), without using physical objects, was proposed. These virtual phantoms, which display realistic noise characteristics, represent an attractive alternative to physical phantoms for providing a reference signal in MRI measurements

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Badia, Pérez Anna. "Antioxidant nanoparticles for the treatment of Age-related Macular Degeneration: preclinical studies in the DKOrd8 mouse model." Doctoral thesis, Universitat Autònoma de Barcelona, 2019. http://hdl.handle.net/10803/670132.

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La Degeneració Macular Associada a la Edat (DMAE) és una malaltia ocular degenerativa d’aparició tardana que afecta la retina central i causa una pèrdua progressiva de visió. La DMAE és una malaltia multifactorial en la qual l’envelliment és un dels principals factor de risc i on l’estrés oxidatiu, l’autofàgia i el sistema immunitari tenen una important rellevància en la seva patogènesis. L’exposició constant de llum que pateix la màcula exigeix una elevada activitat metabòlica per part dels fotoreceptors i l’EPR, cosa que genera nivells elevats d’estrés oxidatiu amb un increment de les espècies reactives d’oxigen (ROS). Això indueix canvis a la retina, incloent la acumulació de lipofuscina, una desregulació de l’autofàgia, la presència de druses i inflamació crònica; cosa que acaba provocant la mort del EPR i dels fotoreceptors. Malgrat ser la principal causa de ceguera en els països desenvolupats, encara no existeix un tractament efectiu per a la DMAE. La falta de coneixement sobre els mecanismes implicats en l’aparició de la malaltia així com la falta de models animals recapitulant el fenotip n’han estat els principals obstacles. En els últims anys, els compostos antioxidants han adquirit importància donat el rol que l’estrés oxidatiu juga en l’aparició de la malaltia. Sota aquestes premisses, els objectius d’aquesta tesis han estat (i) la generació i caracterització del ratolí Ccl2-/-; Cx3cr1-/-; Crb1rd8\/rd8, un model murí per a la DMAE; i (ii) el desenvolupament d’una teràpia basada en nanopartícules amb gran capacitat antioxidant. En primer lloc, vam generar el model Ccl2-/-; Cx3cr1-/-; Crb1rd8/rd8 o DKOrd8 a partir dels knockout simples. Mitjançant la seva monitorització in vivo i caracterització post-mortem, vam observar com els ratolins DKOrd8 presentaven diverses característiques fenotípiques típiques de la DMAE al mes d’edat. Els ratolins DKOrd8 presentaven alteracions a l’hemisferi inferior del fons d’ull amb disrupció de la capa de fotoreceptors i una disminució progressiva del seu gruix. Mitjançant immunofluorescència vam observar una acumulació de micròglia a l’espai subretinià, una migració cap a les lesions de la retina i un increment de gliosis. A més, els ratolins DKOrd8 exhibien alteracions en el patró d’expressió gènic, amb afectació en gens relacionats amb l’estrés oxidatiu, el sistema immunitari i les funcions neuronals entre d’altres. En relació al desenvolupament de la teràpia antioxidant, vam sintetitzar unes nanopartícules antioxidants (AOxNPs) no tòxiques amb una capacitat reproduïble de disminuir els nivells intracel·lulars de ROS en cultius in vitro. A més, vam establir l’administració tòpica com una ruta eficient per arribar a la retina, comparable a la injecció intravítria. Finalment, el tractament tòpic amb AOxNPs en els ratolins DKOrd8 va demostrar ser efectiva, aconseguint revertir el patró d’expressió alterat, disminuir la infiltració de micròglia i prevenint l’aprimament de la capa nuclear externa. En conjunt, en aquest treball hem aconseguit generar un model fiable que presenta certes característiques típiques de la DMAE i hem demostrat l’efecte beneficiós de l’administració tòpica de les AOxNPs en la millora de les característiques patològiques del model.
La Degeneración Macular Asociada a la Edad (DMAE) es una enfermedad ocular degenerativa de aparición tardía que afecta a la retina central y causa una pérdida progresiva de visión. La DMAE es una enfermedad multifactorial en la que el envejecimiento es uno de los principales factores de riesgo y en la que el estrés oxidativo, la autofagia y el sistema inmunitario tienen una importante relevancia en su patogénesis. La exposición constante a la luz que sufre la mácula exige una elevada actividad metabólica por parte de los fotorreceptores y del EPR, lo que genera niveles elevados de estrés oxidativo con un incremento de las especies reactivas de oxigeno (ROS). Esto induce cambios en la retina, incluyendo la acumulación de lipofuscina, una desregulación de la autofagia, la presencia de drusas e inflamación crónica; cosa que acaba causando la muerte del EPR y de los fotorreceptores. Aunque es la principal causa de ceguera en países desarrollados, aun no existe un tratamiento efectivo para la DMAE. La falta de conocimiento sobre los mecanismos implicados en la aparición de la enfermedad así como la falta de modelos animales recapitulando el fenotipo han sido los principales obstáculos. En los últimos años, los compuestos antioxidantes han adquirido importancia dado el rol que el estrés oxidativo juga en la aparición de la DMAE. Bajo estas premisas, los objetivos de esta tesis han estado (i) la generación y caracterización del ratón Ccl2-/-; Cx3cr1-/-; Crb1rd8/rd8, un modelo murino para DMAE; y (ii) el desarrollo de una terapia basada en nanopartículas con gran capacidad antioxidante. En primer lugar, generamos el modelo Ccl2-/-; Cx3cr1-/-; Crb1rd8\/rd8 o DKOrd8 a partir de los knockout simples. Mediante su monitorización in vivo y caracterización post-mortem, observamos cómo los ratones DKOrd8 presentaban varias características fenotípicas típicas de la DMAE al mes de edad. Los ratones DKOrd8 presentaban alteraciones en el hemisferio inferior del fondo de ojo con disrupción de la capa de fotorreceptores y una disminución progresiva de su grosor. Mediante inmunofluorescencia observamos una acumulación de microglía en el espacio subretiniano y migración hacia las lesiones de la retina y un incremento de gliosis. Además, los ratones DKOrd8 exhibían alteraciones en el patrón de expresión génica, con afectación en genes relacionados con el estrés oxidativo, el sistema inmunitario y las funciones neuronales entre otros. En relación al desarrollo de la terapia antioxidante, sintetizamos unas nanopartículas antioxidantes (AOxNPs) no tóxicas con capacidad reproducible de disminuir los niveles intracelulares de ROS en cultivos in vitro. Además, establecimos la administración tópica como una ruta eficiente para la llegada a la retina, comparable a la inyección intravítrea. Finalmente, el tratamiento tópico con AOxNPs en los ratones DKOrd8 demostró ser efectivo, consiguiendo revertir el patrón de expresión alterado, disminuir la infiltración de microglía y prevenir el adelgazamiento de la capa nuclear externa. En conjunto, en este trabajo hemos conseguido generar un modelo fiable que presenta ciertas características típicas de la DMAE y hemos demostrado el efecto beneficioso de la administración tópica de las AOxNPs en la mejora de las características patológicas del modelo.
Age-related Macular Degeneration (AMD) is a late-onset degenerative eye disease that affects the central retina and causes progressive vision impairment. AMD is a multifactorial disease, with advanced age being the main risk factor and oxidative stress, autophagy and the immune system playing an important role in its pathogenesis. The constant exposure to light in the macula demands a high metabolic rate for photoreceptors and RPE cells, which generate elevated levels of oxidative stress. These increased levels of ROS induce several changes in the retina, including the accumulation of lipofuscin, the deregulation of autophagy, the presence of drusen and chronic inflammatory responses that end up causing RPE cells death and photoreceptor loss. Despite being the major cause of blindness in developed countries, AMD still does not have an effective treatment available. The lack of knowledge about the mechanisms implicated in the onset of the disease and the lack of animal models recapitulating the phenotype of the disease have been the major obstacles. In the last years, antioxidant compounds have gained importance given the role oxidative stress plays on the onset of the disease. Under these premises, the aim of this thesis was set on (i) the generation and characterization of the Ccl2-/-; Cx3cr1-/-; Crb1rd8\/rd8 mouse, a murine model for AMD; and (ii) the development of a therapy based on nanoparticles with high antioxidant capacity. The Ccl2-/-; Cx3cr1-/-; Crb1rd8/rd8 or DKOrd8 model was generated from the single knockout mice. Its posterior in vivo monitoring and post-mortem characterization revealed that DKOrd8 mice mimicked several AMD-like features already at one month of age. DKOrd8 mice presented alterations in the inferior hemisphere of their eye fundus with disruption and age-dependent decrease of the photoreceptor layer; and alterations in their retinal function at an early age that were maintained upon ageing. Immunofluorescence stainings revealed an age-dependent accumulation of microglia in the subretinal space, migration of the microglia towards the inner retinal lesions and increased gliosis. Moreover, DKOrd8 mice exhibit an altered expression pattern with an affectation of genes involved with oxidative stress, immune regulation and neuronal function among others. Regarding the development of the antioxidant therapy, we synthetize non-toxic antioxidant nanoparticles (AOxNPs) with a reproducible capacity of reducing intracellular ROS levels in in vitro cultures and established the topical administration as an efficient delivery route to reach the retina and posterior segment of the eye comparable to the intravitreal injection. Finally, the topical treatment of DKOrd8 mice with AOxNPs was effective in improving the phenotypical characteristics of the mouse model, as it reverted their altered expression profile, reduced microglia infiltration and prevented the thinning of the outer nuclear layer. Altogether, we generated a reliable mouse model that mimicked certain features of AMD and proved the beneficial effect of the topical AOxNPs in improving the pathological characteristics of the model.

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Gaspar, Nathalie. "Preclinical evaluation of different HSP90 inhibitors in adult and pediatric glioblastoma in vitro and in vivo models." Paris 11, 2010. http://www.theses.fr/2010PA11T041.

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Keene, J. M. "Models of dependence and treatment in the social construction and control of addiction." Thesis, Swansea University, 1991. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.637770.

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This study involved the evaluation of the Alcohol and Drug Advice Centre in Swansea, examining not only its general effectiveness but also the theory underlying its practice. The fieldwork took place over a period of two years and six months, 1986 to 1988. The aim was to do a small scale intensive study, which would give insight into the process of treatment, change and maintenance of change, and provide an understanding of individual and interpersonal processes taking place at the Centre. An ethnographic study of forty clients as they passed through the treatment process facilitated the examination of possible variables influencing different outcomes, giving an indication of why particular individuals 'succeeded' in treatment while others 'failed'. The qualitative data collected give an indication of the degree of compatibility between views of clients and the beliefs of staff together with the Alcoholics Anonymous philosophy of the Centre. These data were supplemented by a statistical analysis of client characteristics and attendance. The theory and practice of the Centre is placed in the context of both historical and contemporary theories of addiction. Theoretical controversies and changes in policy and practice are reviewed. The work in this study has demonstrated that there is some correlation between the beliefs and theories of client and therapist on the one hand and successful outcome on the other. The implications of these conclusions for treatment are discussed, focusing on the possibility of pre-treatment 'matching' of clients to particular approaches. It is suggested that different theoretical components that both inform and partly constitute experiences of treatment should be evaluated, not only in terms of internal consistency and external validity but also in terms of usefulness and therapeutic tools or ideologies. The possibility of combining conflicting treatment theories within the provision of an effective service, by a separation of allocation and treatment functions, is considered.

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Zimmerman, Bevin. "Studies of Targeted Therapies Against Human T Lymphotropic Virus Type-1 Adult T-cell Lymphoma in Preclinical Animal Models." The Ohio State University, 2009. http://rave.ohiolink.edu/etdc/view?acc_num=osu1243431025.

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Piggott, Luke. "Investigating the therapeutic potential of cellular FLICE-like inhibitory protein and TRAIL in preclinical models of breast cancer." Thesis, Cardiff University, 2012. http://orca.cf.ac.uk/44561/.

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Apoptosis is an important process in normal mammary gland physiology and evasion of apoptosis has also been identified as a hallmark of cancer. In breast cancer cells apoptotic resistance is an acquired feature that can promote tumour growth and progression. Induction of apoptosis by the extrinsic death ligand TRAIL has been shown to be a promising clinical therapy targeting a number of different cancer cells whilst sparing normal cells. Unfortunately most breast cancers are inherently resistant to TRAIL treatment. Herein it is shown that by reducing the expression of the downstream TRAIL inhibitor c-FLIP, a range of different breast cancer subtypes can be sensitised to TRAIL treatment resulting in significant cancer cell death. Significantly, suppression of c-FLIP in combination with TRAIL (FLIPi/TRAIL) ablated the tumour-initiating breast cancer stem cell (bCSC) subset, as defined by mammosphere formation assay, within cell lines. This selective killing of bCSCs translated to reduced tumour initiation and metastasis in animal transplant models. However, continued culture of FLIPi/TRAIL treated cell lines in adherent conditions resulted in bCSC re-acquisition suggesting a phenotypic plasticity of non-bCSC cells. Re-acquired bCSCs also demonstrated sensitivity to repeated FLIPi/TRAIL treatment and maintaining reduced c-FLIP expression prevented bCSC re-acquisition. These results substantiate the importance of resistance to apoptosis in tumour initiation and metastasis and identify the targeting of c-FLIP proteins as a promising anti-cancer therapeutic approach. Acquired resistance to existing mainstay therapies such as antiestrogens (AEs) (tamoxifen and Faslodex) and aromatase inhibitors (AIs) is an ongoing obstacle in treatment of a large number of breast cancer patients. AE-resistant models of breast cancer and a multiple endocrine-resistant patient sample demonstrated hypersensitivity to TRAIL. This sensitivity was observed in both in vitro and in vivo models of AE resistance and cell death was prevalent in both bulk tumour cells and bCSCs. Sensitisation was not attributed to combination AE/TRAIL treatment suggesting cellular changes during the acquisition of AE resistance are responsible for TRAIL sensitivity in these models. Further investigation suggested that the mechanism of AE-resistant cell sensitivity to TRAIL was not dependant on functional estrogen receptor signalling and is most likely dependant on the AE agent that the cancer cells have acquired resistance to. Interestingly tamoxifen-resistant MCF-7 cells were shown to have reduced c-FLIP protein expression compared to parental cells, further supporting c-FLIP’s potential in cancer therapy. Recent success in the use non-MHC-restricted γδ T cells as a targeted immunotherapy in clinical trials has identified this therapeutic methodology as desirable. Here it is shown that TRAIL is readily expressed by this subset of T cells that also demonstrate cytotoxicity to breast cancer cell lines. Neither the secretion of TRAIL or surface expression of TRAIL appeared to contribute significantly towards γδ T cell cytotoxicity and the majority of breast cancer cell death induced by γδ T cells would seem to be perforin-mediated. The suppression of c-FLIP in target cells increased γδ T cell cytotoxicity but again not via TRAIL. Preliminary results also indicated that the bCSCs of some cell lines were exquisitely sensitive to γδ T cell treatment. In summary these results indicate that targeting c-FLIP and TRAIL can be therapeutically beneficial in a range of different breast cancer subtypes by certain therapeutic strategies.

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Pereyra, Samuel Alejandro. "Pornography Use and Loneliness: Assessing Correlations Using Three Associative Models." BYU ScholarsArchive, 2016. https://scholarsarchive.byu.edu/etd/6417.

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Given the current debate on the discrepancies of pornography research and the inconclusiveness of addiction to pornography, this study examines the associative nature between pornography use and loneliness using a measurement model and two structural equation models where pornography use and loneliness are regressed on each other, respectively. Survey data was collected from a sample of 1,247 participants, who completed an online questionnaire containing questions on pornography use, the University of Los Angeles Loneliness Scale (UCLALS), and other demographic variables. Results from our analyses revealed significant and positive associations between pornography use and loneliness for all three models. Findings provide grounds for future possible models of bidirectionality suggesting a possible addictive cycle between pornography use and loneliness.

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Cannella, Lee Anne. "CHARACTERIZATION OF THE ROLE AND UNDERLYING MECHANISMS OF TRAUMATIC BRAIN INJURY ON REWARD SEEKING BEHAVIOR USING PRECLINICAL ANIMAL MODELS." Diss., Temple University Libraries, 2019. http://cdm16002.contentdm.oclc.org/cdm/ref/collection/p245801coll10/id/568829.

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Biomedical Sciences
Ph.D.
Traumatic brain injury (TBI) is a prominent healthcare concern in the U.S. as millions of TBI-related emergency department visits occur annually. Recent reports estimate more than 5 million Americans currently suffer from life-long disabilities and psychiatric complications associated with TBI. While the risk of TBI has conventionally been considered to be male dominated, analyses of sex-comparable sports indicate that rates of concussions are higher and recovery time is longer following brain injury in females. Following anxiety and depression, substance use disorder (SUD) is the third most common de-novo neuropsychiatric condition diagnosed in both male and female TBI patients. Importantly, during adolescence the primary neuronal networks that regulate reward behaviors and perception of drug-induced euphoria are not fully developed, corroborating epidemiological studies identifying TBI sustained during adolescence as a risk factor for problematic drug use. Yet, to date, little is known about how TBI-induced molecular changes affect brain structures essential for the perception of reward and processing drug-induced euphoria. The following experiments were designed to test the hypothesis that adolescent TBI-induced neuroinflammation in areas such as prefrontal cortex (PFC) and nucleus accumbens (NAc) results in remodeling of neuronal reward networks and affect how the rewarding effects of cocaine shift as a consequence of TBI. Notably, the extent of sex differences in SUD susceptibility in TBI has not be investigated. Therefore, we also investigated whether the immune response stimulated by early-life TBI alters maturation of reward neurocircuits, leading to increased SUD vulnerability in a sex-dependent manner. Following the induction of TBI using the controlled cortical impact (CCI) model of brain injury, we utilized a biased, three-phased cocaine conditioned place preference (CPP) assay to assess the behavioral response to the rewarding effects of cocaine following adolescent injury in male and female C57BL6 mice. Furthermore, we characterized the effect of CCI-TBI on the stimulation of neuroinflammation within the PFC and NAc, comprising the reward pathway. Specifically, our studies revealed a sex-specific increase in 1) sensitivity to the rewarding efficacy of a subthreshold doses of cocaine interpreted from significantly higher cocaine CPP shifts, 2) the activation and phagocytosis of microglia observed by the positive expression of neuronal synaptic proteins in microglia sorted using flow cytometry, 3) increase in permeability of the blood-brain barrier indicated by discontinuous and depleted expression of tight junction proteins that line microvasculature isolated from reward nuclei, 4) decreased neuronal complexity, arborization, and spine density quantified from Golgi-cox stained NAc neurons, 5) changes in expression of genes related to the dopamine system analyzed by qRT-PCR in only male mice injured during adolescence. Additionally, our results imply that high levels of female hormones can promote neuroprotection against increased sensitivity to the rewarding properties of cocaine following injury, associated with decreased neuroinflammatory profiles after TBI in adolescent females. The studies herein aimed to elucidate underlying neuropathological outcomes following TBI in the reward circuitry that could be contributing to increased risk of addiction-like behavior observed clinically. Our findings suggest that TBI during adolescence may enhance the abuse liability of cocaine in adulthood and vulnerability to the rewarding effects of cocaine could be higher as a result of brain injury. Key pathological findings in the NAc such as activated microglial phagocytosis, BBB changes, reduced neuronal complexity, and changes in dopamine gene expression in areas of the reward pathways support the notion that neuroinflammation may contribute to how the rewarding efficacy of cocaine are affected post-TBI during adolescence. The ultimate goal of this research is to 1) advance TBI and SUD literature with the potential to increase awareness and help health care providers inform TBI patients about the increased risk for SUDs, and 2) to translate identified correlated mechanisms into novel targeted therapies that would provide a launching point for the treatment of patients with TBI-related SUD.
Temple University--Theses

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Melemedjian, Ohannes, Marina Asiedu, Dipti Tillu, Raul Sanoja, Jin Yan, Arianna Lark, Arkady Khoutorsky, et al. "Targeting adenosine monophosphate-activated protein kinase (AMPK) in preclinical models reveals a potential mechanism for the treatment of neuropathic pain." BioMed Central, 2011. http://hdl.handle.net/10150/610214.

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Neuropathic pain is a debilitating clinical condition with few efficacious treatments, warranting development of novel therapeutics. We hypothesized that dysregulated translation regulation pathways may underlie neuropathic pain. Peripheral nerve injury induced reorganization of translation machinery in the peripheral nervous system of rats and mice, including enhanced mTOR and ERK activity, increased phosphorylation of mTOR and ERK downstream targets, augmented eIF4F complex formation and enhanced nascent protein synthesis. The AMP activated protein kinase (AMPK) activators, metformin and A769662, inhibited translation regulation signaling pathways, eIF4F complex formation, nascent protein synthesis in injured nerves and sodium channel-dependent excitability of sensory neurons resulting in a resolution of neuropathic allodynia. Therefore, injury-induced dysregulation of translation control underlies pathology leading to neuropathic pain and reveals AMPK as a novel therapeutic target for the potential treatment of neuropathic pain.

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Kreple, Collin John. "Examining the role of ASIC1A in mouse models of addiction and CO2-evoked panic-like behaviors." Diss., University of Iowa, 2015. https://ir.uiowa.edu/etd/1668.

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Acid-sensing ion channel 1A (ASIC1A) is abundant in the nucleus accumbens (NAc), a region known for its role in addiction. Because ASIC1A has been previously suggested to promote associative learning, we hypothesized that disrupting ASIC1A in the NAc would reduce drug-associated learning and memory. However, contrary to this hypothesis, we found that disrupting ASIC1A in the NAc increased cocaine-conditioned place preference, suggesting an unexpected role for ASIC1A in addiction-related behavior. Investigating the underlying mechanisms, we identified a novel postsynaptic current during neurotransmission mediated by ASIC1A and ASIC2 and thus well-positioned to regulate synapse structure and function. Consistent with this possibility, disrupting ASIC1A altered dendritic spine density and glutamate receptor function, and increased cocaine-evoked plasticity in AMPA-to-NMDA ratio, all resembling changes previously associated with cocaine-induced behavior. Together, these data suggest ASIC1A inhibits plasticity underlying addiction-related behavior, and raise the possibility of therapies for drug addiction by targeting ASIC-dependent neurotransmission. The amygdala plays critical roles in the learning and expression of fear-related behavior. Previous studies have implicated the amygdala in CO2-evoked fear-like behavior in mice; however, a more recent study demonstrated that humans lacking the amygdala bilaterally experience fear and panic with CO2-inhalation. Because all subjects lacking the amygdala had panic attacks after inhaling CO2 compared to only 25% of controls, this data suggests the amygdala may play an inhibitory role in CO2-evoked panic. To assess the role of the amygdala in CO2-evoked behaviors in mice, we lesioned the amygdala and optogenetically stimulated different amygdalar nuclei. We found that large unilateral and bilateral amygdala lesions caused the emergence of escape-like jumping behavior in mice exposed to CO2 and a relative deficit in CO2-evoked freezing. This jumping behavior depended on the dorsal periaqueductal gray, a brain area previously associated with panic attacks. Additionally, the putative CO2 chemosensor ASIC1A and ASIC2 are not necessary for CO2-evoked jumping, and may even play an inhibitory role in this behavior. Optogenetic manipulation of the amygdala revealed that stimulation of the basolateral amygdala enhanced jumping behavior and inhibited freezing behavior. This may be due to the basolateral amygdala's ability to inhibit the main output center of the amygdala, the central nucleus. Together, these results suggest that different amygdalar nuclei differentially modulate CO2-evoked behavior by regulating the switch between mobile and immobile defense responses. Additionally, they provide additional evidence that amygdalar dysfunction may contribute to panic disorder.

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Buckel, Lisa [Verfasser], and Aladár [Akademischer Betreuer] Szalay. "Evaluating the combination of oncolytic vaccinia virus and ionizing radiation in therapy of preclinical glioma models / Lisa Buckel. Betreuer: Aladar Szalay." Würzburg : Universität Würzburg, 2012. http://d-nb.info/1102820695/34.

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Martínez, Pérez Carlos. "Evaluation of the antitumour activity of novel flavonoids on pre-clinical models of breast and ovarian cancer." Thesis, University of Edinburgh, 2017. http://hdl.handle.net/1842/25410.

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New drugs are needed for better cancer management. Clinical trials are currently underway to assess the use of flavonoids (natural polyphenols) as anticancer agents. Among them, myricetin has been shown to induce cell cycle arrest and apoptosis in pre-clinical cancer models. We hypothesised that myricetin-derived novel flavonoids designed to enhance this natural potential and improve on the drug-likeness limitations of myricetin might have increased potential for their application in the management of breast and ovarian cancer. The effect of a library of novel flavonoids was screened on 3 panels of breast and ovarian cancer cell lines, representing different molecular subtypes and phenotypes, to assess their potency. The second-generation bi-methoxylated analogue AO-1530-OMe (Oncamex) was identified as the most effective candidate in the library, with sub-micromolar concentrations exerting a strong antiproliferative effect across almost all models studied. Results suggested that changes in the hydroxylation profile, the addition of methoxylations and a decyl alkyl chain were some of the structure-activity relationships contributing to this improved efficacy. Plate assays showed 8 h treatment with Oncamex reduced cell viability and induced cytotoxicity and apoptosis, concomitant with caspase activation and PARP cleavage. Pre-incubation with an antioxidant partially blocked these effects, suggesting the possible involvement of ROS modulation in the mechanism of action of Oncamex. Fluorescence microscopy reported the quick and stable delivery of Oncamex to the mitochondria. Fluorescent probes showed that Oncamex can induce mitochondrial superoxide production at concentrations associated with its antiproliferative effects. Study of the electrochemical properties of Oncamex by cyclic voltammetry supported this. Differential gene expression analysis following a microarray experiment showed Oncamex induces changes in the expression of genes controlling cell cycle and apoptosis. Together with previous results, the findings from this analysis led to the postulation of a model for the mechanism of action of Oncamex: due to its enhanced reactivity and mitochondrial targeting, Oncamex can generate mitochondrial superoxide, leading to mitochondrial dysfunction, membrane permeabilisation and the activation of the JNK pathway and the transcription factor FOXO3, which together contribute to the induction of intrinsic apoptosis and the inhibition of proliferation. Further proliferation assays on cell culture models also reported enhanced effect of Oncamex when administered in combination with paclitaxel and TRAIL. These improved responses were observed in breast and ovarian cancer models, including cells lines characterised by their treatment-resistant phenotype. Cotreatment with Oncamex also improved the effect of tamoxifen on anti-oestrogen resistant LCC9 breast cancer cells. Results from preliminary in vivo studies in mice implanted with the MDA-MB-231 breast cancer xenograft were consistent with an antiproliferative effect of Oncamex (25mg/kg/day) in vivo, as treatment inhibited tumour growth and reduced the expression of the marker of proliferation Ki-67 without signs of systemic toxicity. Tissues from this experiment also allowed for preliminary in vivo validation of the proposed mechanism of action of Oncamex by immunohistochemistry. The in vivo cytostatic effect of Oncamex was confirmed in a second in vivo experiment, which also investigated the effect of Oncamex at higher doses or in combination with paclitaxel. In conclusion, the novel flavonoid Oncamex has shown a promising antiproliferative effect in pre-clinical models of breast and ovarian cancer, including models of treatment-resistant cancers. Preliminary in vivo studies have demonstrated a partial recapitulation of the effect of Oncamex. A mechanistic model has been proposed by which Oncamex induces intrinsic apoptosis through its redox reactivity and mitochondrial targeting. These results support the potential of this prototypic candidate, although possible work in the structure and formulation of this candidate and further study and validation of its mechanism of action is needed for its continued development as an anticancer agent.

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Ruocco, Margherita <1981&gt. "Generation and characterization of mouse models of Small cell lung cancer and Basal cell carcinoma for the preclinical evaluation of new therapies." Doctoral thesis, Alma Mater Studiorum - Università di Bologna, 2013. http://amsdottorato.unibo.it/5623/.

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Background. Human small cell lung cancer (SCLC) accounting for approximately 15-20% of all lung cancers, is an aggressive tumor with high propensity for early regional and distant metastases. Although the initial tumor rate response to chemotherapy is very high, SCLC relapses after approximately 4 months in ED and 12 months in LD. Basal cell carcinoma (BCC) is the most prevalent cancer in the western world, and its incidence is increasing worldwide. This type of cancer rarely metastasizes and the death rate is extraordinary low. Surgery is curative for most of the patients, but for those that develop locally advanced or metastatic BCC there is currently no effective treatment. Both types of cancer have been deeply investigated and genetic alterations, MYCN amplification (MA) among the most interesting, have been found. These could become targets of new pharmacological therapies. Procedures. We created and characterized novel BLI xenograft orthotopic mouse models of SCLC to evaluate the tumor onset and progression and the efficacy of new pharmacological strategies. We compared an in vitro model with a transgenic mouse model of BCC, to investigate and delineate the canonical HH signalling pathway and its connections with other molecular pathways. Results and conclusions. The orthotopic models showed latency and progression patterns similar to human disease. Chemotherapy treatments improved survival rates and validated the in vivo model. The presence of MA and overexpression were confirmed in each model and we tested the efficacy of a new MYCN inhibitor in vitro. Preliminar data of BCC models highlighted Hedgehog pathway role and underlined the importance of both in vitro and in vivo strategies to achieve a better understanding of the pathology and to evaluate the applicability of new therapeutic compounds

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Koschitzki, Kim Christine Cornelia [Verfasser], and Franz [Gutachter] Jakob. "Evaluation of preclinical animal models in bone tissue engineering and their success in clinical translation / Kim Christine Cornelia Koschitzki ; Gutachter: Franz Jakob." Würzburg : Universität Würzburg, 2020. http://d-nb.info/121459400X/34.

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Ordway, Gregory A., Attila Szebeni, Liza J. Hernandez, Jessica D. Crawford, Katalin Szebeni, Michelle J. Chandley, Katherine C. Burgess, Corwin Miller, Erol Bakkalbasi, and Russell W. Brown. "Antidepressant-Like Actions of Inhibitors of Poly(ADP-Ribose) Polymerase in Rodent Models." Digital Commons @ East Tennessee State University, 2017. https://dc.etsu.edu/etsu-works/2768.

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Many patients suffering from depressive disorders are refractory to treatment with currently available antidepressant medications, while many more exhibit only a partial response. These factors drive research to discover new pharmacological approaches to treat depression. Numerous studies demonstrate evidence of inflammation and elevated oxidative stress in major depression. Recently, major depression has been shown to be associated with elevated levels of DNA oxidation in brain cells, accompanied by increased gene expression of the nuclear base excision repair enzyme, poly(ADP-ribose) polymerase-1. Given these findings and evidence that drugs that inhibit poly(ADP-ribose) polymerase-1 activity have antiinflammatory and neuroprotective properties, the present study was undertaken to examine the potential antidepressant properties of poly(ADP-ribose) polymerase inhibitors.

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Martin, Bradley J. "Dopamine and Norepinephrine Transporter Inhibition in Cocaine Addiction: Using Mice Expressing Cocaine-Insensitive Transporters." The Ohio State University, 2011. http://rave.ohiolink.edu/etdc/view?acc_num=osu1312907534.

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Floriano, Maureen Elizabeth. "Models of Addiction and Health Seeking Behaviors: Understanding Participant Utilization of an Overdose Education and Naloxone Distribution Clinic." Case Western Reserve University School of Graduate Studies / OhioLINK, 2021. http://rave.ohiolink.edu/etdc/view?acc_num=case1619772720856071.

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Dissertations / Theses on the topic 'Preclinical models of addiction'

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