Purpose: Combretastatin A4 phosphate (CA4P) and its structural analog, combretastatin A1 phosphate (CA1P), are soluble prodrugs capable of interacting with tubulin and causing rapid vascular shutdown within tumors. CA4P has completed Phase I clinical trials, but recent preclinical studies have shown that CA1P displays a greater antitumor effect than the combretastatin A4 (CA4) analog at equal doses. The aim of this study, therefore, is to compare pharmacokinetics and metabolism of the two compounds to determine whether pharmacokinetics plays a role in their differential activity. Experimental Design: NMRI mice bearing MAC29 tumors received injection with either CA4P or CA1P at a therapeutic dose of 150 mg·kg-1, and profiles of both compounds and their metabolites analyzed by a sensitive and specific liquid chromatography/mass spectroscopy method. Results: The metabolic profile of both compounds is complex, with up to 14 metabolites being detected for combretastatin A1 (CA1) in the plasma. Many of these metabolites have been identified by liquid chromatography/mass spectroscopy. Initial studies, however, focused on the active components CA4 and CA1, where plasma and tumor areas under the curve were 18.4 and 60.1 µg·h·ml-1 for CA4, and 10.4 and 13.1 µg·h·ml-1 for CA1, respectively. In vitro metabolic comparisons of the two compounds strongly suggest that CA1 is metabolized to a more reactive species than the CA4. Conclusions: Although in vitro studies suggest that variable rates of tumor-specific prodrug dephosphorylation may explain these differences in pharmacokinetics profiles, the improved antitumor activity and altered pharmacokinetic profile of CA1 may be due to the formation of a more reactive metabolite.

no / Purpose: Combretastatin A4 phosphate (CA4P) and its structural analog, combretastatin A1 phosphate (CA1P), are soluble prodrugs capable of interacting with tubulin and causing rapid vascular shutdown within tumors. CA4P has completed Phase I clinical trials, but recent preclinical studies have shown that CA1P displays a greater antitumor effect than the combretastatin A4 (CA4) analog at equal doses. The aim of this study, therefore, is to compare pharmacokinetics and metabolism of the two compounds to determine whether pharmacokinetics plays a role in their differential activity. Experimental Design: NMRI mice bearing MAC29 tumors received injection with either CA4P or CA1P at a therapeutic dose of 150 mg/kg-1 , and profiles of both compounds and their metabolites analyzed by a sensitive and specific liquid chromatography/mass spectroscopy method. Results: The metabolic profile of both compounds is complex, with up to 14 metabolites being detected for combretastatin A1 (CA1) in the plasma. Many of these metabolites have been identified by liquid chromatography/mass spectroscopy. Initial studies, however, focused on the active components CA4 and CA1, where plasma and tumor areas under the curve were 18.4 and 60.1 microgram/h/ml-1 for CA4, and 10.4 and 13.1 microgram/h/ml-1 for CA1, respectively. In vitro metabolic comparisons of the two compounds strongly suggest that CA1 is metabolized to a more reactive species than the CA4. Conclusions: Although in vitro studies suggest that variable rates of tumor-specific prodrug dephosphorylation may explain these differences in pharmacokinetics profiles, the improved antitumor activity and altered pharmacokinetic profile of CA1 may be due to the formation of a more reactive metabolite.

Ficus thonningii is a nutraceutical that is extensively used in ethnomedicine. Nursing mothers use F. thonningii leaves as nutritional and medicinal supplements and are at risk of exposing their infants to its constituent phytochemicals. The exposure of the sensitive neonatal gastrointestinal tract (GIT) to these phytochemicals can result in irreversible changes in growth and development. The objectives of this study were to determine the effects of crude F. thonningii extracts on; growth, morphology and morphometry of the abdominal viscera and clinical biochemistry of neonatal rats. Forty, suckling Sprague Dawley rats of either sex were randomly divided into 5 groups. Each group was orally gavaged once daily with either low (50 mg/kg b.w) or high (500 mg/kg b.w) doses of aqueous or methanolic extracts of F. thonningii, for 7 days. The control rats received distilled water. The pups were euthanased and tissues were collected and weighed. Samples of the liver, caecum and proximal small intestine were preserved and processed for histology. Plasma biochemical parameters were analysed colorimetrically. Data was presented as means + SD. F. thonningii extracts exhibited trophic effects on the stomach and ceacal mucosa of rats but had no significant growth-promoting effects on the small intestine and visceral organs. Histological analysis of the intestine, liver and caeca revealed no mucosal damage. Clinical chemistry parameters were not abnormally altered. There was a significant decrease (p<0.05, ANOVA) in the plasma concentrations of basal (non-fasting) glucose in the pups on the high methanolic extracts. However, the triglyceride and cholesterol levels were unaltered by the treatments. The findings suggest that F. thonningii extracts exhibit trophic effects on the mucosal layers of the stomach and caecum. F. thonningii extracts also possess glucose-lowering activity. At low doses, F. thonningii extracts can be safely used without the risk of any disruption in the structural integrity of the neonatal rat GIT and function of the liver and kidneys.

No description available.

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.

La transplantation d’îlots permet la restauration d’une insulino-sécrétion endogène chez les patients diabétiques de type 1 par greffe allogénique et limite les conséquences métaboliques d’une pancréatectomie en cas d’autogreffe. Le site de référence intrahépatique présente néanmoins de nombreuses limites. Dans le cadre d’autogreffe, le risque hémorragique accru chez les patients récemment opérés liée à l’injection nécessairement conjointe des îlots et d’héparine en intraportal doit faire privilégier une technique de greffe mini invasive. De nombreux sites d’implantation ont été décrits. L’hypothèse de ce travail de thèse était que la voie intramusculaire offrirait par rapport à la voie portale, l’avantage de simplifier l’acte de transplantation, de réduire le traumatisme pour le patient et d’améliorer la viabilité des îlots en limitant les processus inflammatoires immédiats et en optimisant les processus de néo-vascularisation.Dans une première partie, nous avons pu démontrer, dans un model préclinique, que le site intramusculaire permet la survie, la revascularisation et la sécrétion des îlots autogreffés. Nous avons décrit une technique de greffe permettant d’ameliorer le contrôle glycémique d’animaux autogreffés après pancréatectomie totale. Bien qu’inférieure à la voie intraportale, les tests fonctionnels nous ont permis de valider le site intramusculaire pour la greffe d’îlots autologues. Dans une deuxième partie, nous décrivons un cas clinique original confirmant la possible transposition en clinique de l’autogreffe d’ilots en intramusculaire après pancréatectomie partielle. Ce cas clinique, confirme la faisabilité et suggère son innocuité. Il était cependant difficile dans ce contexte de pancréatectomie partielle d’établir un rapport entre l’absence de développement de diabète et la greffe. Pour ce faire, nous décrivons dans une troisième partie, une étude pilote sur l’évaluation de la fonction des îlots autogreffés dans le muscle chez 8 patients ayant subi une pancréatectomie partielle. Dans ce but nous avons comparé la sécrétion d’insuline après stimulation par l’arginine mesurée simultanément dans le bras greffé et le bras non greffé après l’autogreffe par des tests de stimulation à l’arginine. Malgré une faible quantité d’ilots greffés, nous avons documenté une fonction primaire du greffon chez plus de la moitié des patients, ainsi que sa persistance à plus d’un an. Enfin, nous avons également montré que le gradient d’insulinémie entre le bras greffé et le bras systémique était corrélé avec la masse d’ilots greffés.Le muscle est donc un site phare pour le développement d’un site alternatif lors de greffe d’ilots intramusculaire. Le site intramusculaire permet un formidable site d’évaluation des îlots. Cette procédure, résolument mini-invasive, est particulièrement attractive par son extrême accessibilité aux biopsies, à l’imagerie et aux explantations. Cette accessibilité permet d’élargir les indications de greffe telles que l’autogreffe d’îlots provenant de pancréas tumoraux. / The liver may not be an optimal site for islet transplantation due to obstacles by an instant blood-mediated inflammatory response (IBMIR), and low revascularization of transplanted islets. Therefore, intramuscular islet transplantation (IMIT) offers an attractive alternative, based on its simplicity, enabling easier access for noninvasive graft imaging and cell explantation. In this study, we explored the outcome of autologous IMIT in the minipig (n = 30). Using the intramuscular injection technique, we demonstrated by direct histological evidence the rapid revascularization of islets autotransplanted into the gracilius muscle. Islet survival assessment was performed using immunohistochemistry staining for insulin and glucagon up to a period of 6 months. Furthermore, we showed the crucial role of minimizing mechanical trauma to the myofibers and limiting exocrine contamination. Intramuscular islet graft function after transplantation was confirmed by documenting the acute insulin response to intravenous glucose in 5/11 pancreatectomized animals. Graft function after IMIT remained however significantly lower than the function measured in 12 out of 18 minipigs who received a similar islet volume in the liver through intraportal infusion. Collectively, these results demonstrated in a clinically relevant preclinical model, suggest IMIT as a promising alternative to intraportal infusion for the transplantation of &#946; cells in certain medical situations.

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.

No description available.

No description available.

Sepsis is a life-threatening condition characterized by organ dysfunction due to an uncontrolled response to infection. Despite decades of research, the mortality rate remains high, emphasizing the need for an improved understanding of sepsis pathophysiology and improvements in preclinical animal research. Recently, extracellular histones, major mediators of organ dysfunction and death, have emerged as a potential therapeutic target for sepsis. In this thesis, we reported that the ability of heparin to neutralize the cytotoxic and procoagulant effects of histones is size-dependent but independent of the antithrombin- binding pentasaccharide. In contrast, the ability of heparin to neutralize histone-mediated impairment of activated protein C generation is independent of size and anticoagulant activity. These findings suggest that heparin variants may have differential therapeutic potential in vascular disease states that are associated with elevated levels of histones. Before testing the therapeutic efficacy of the heparin variants in vivo, we aimed to develop and standardize a murine model of sepsis that can be utilized in a multi-center platform. As one of the lead sites for the National Preclinical Sepsis Platform (NPSP), we optimized a 72-hour model of abdominal sepsis using supportive treatments. As sepsis predominately impacts the elderly, we also explored the impact of aging on the host response to sepsis using our fecal induced peritonitis (FIP) model. Aged FIP mice exhibited a higher mortality rate compared to young FIP mice. The worsened organ injury and poor survival in aged mice may be attributed to heightened inflammation in aged mice. We also observed trends in increased bacterial loads, increased coagulation, elevated cell free DNA, and decreased ADAMTS13 activity in aged septic mice. These findings help to improve our understanding of how aging impacts the host response to sepsis, which may be translated into therapeutic strategies that considers advanced age as a risk factor for sepsis. / Thesis / Candidate in Philosophy