As one of the natural polyphenols, resveratrol possesses hydroxyl substituted trans-stilbene structure and exerts impact on health by inhibiting multiple human enzymes, such as cyclooxygenase, F1 ATPase, and tyrosinase. Resveratrol has to be bound by human serum albumin (HSA) to keep a high concentration in serum, since its solubility is low in water. To improve water solubility and bioavailability, two resveratrol aliphatic acids and their esters have been designed and synthesized. The solubilities of the resveratrol and its derivatives have been measured using a standard procedure. The two aliphatic acids showed better solubilities in pure water and phosphate buffer (pH 7). The binding affinities of resveratrol derivatives for HSA were also measured, and the drug-protein interaction mechanism was investigated using fluorescence, UV-vis, and NMR spectroscopies. Interestingly, resveratrol hexanoic acid (5) was found to be a much better ligand (Ka = (6.70 ± 0.10) × 106 M-1) for HSA than resveratrol (Ka = (1.64 ± 0.07) × 105 M-1), and there was 41-fold improvement for the binding affinity. It was the first time that the increase of fluorescence of resveratrol moiety was observed during the binding to HSA, suggesting that 5 should be bound tightly by HSA. The UV-vis absorption spectroscopy revealed a maximum absorption shift from 318 to 311 nm with decreasing intensity by 20% upon complexation, suggesting that the π-π conjugation of the stilbene structure was impaired during the binding. Although HSA was reported to have only one binding site for resveratrol, the Job's and molar ratio plots suggested that HSA should bind two molecules of 5. NMR study suggested that phenyl group (B ring) in the center of the molecule of 5 should be involved in the π-π stacking interactions with HSA aromatic amino acid residues. Molecular geometry calculation of 5 with Spartan software showed that the stilbene structure had two conformers, orthogonal and planar ones. The former (E = -1.432 KJ/mol) was more stable than the latter (E = -0.128 KJ/mol), suggesting that the former should be the conformer of 5 in the complexation with HSA.

ZNS-Tumore stellen im Kindesalter die häufigsten soliden sowie die zweithäufigsten Tumorerkrankungen insgesamt nach den Leukämien dar. Sie bedürfen in aller Regel einer multimodalen Therapie, bestehend aus möglichst vollständiger Resektion, Chemo- und Strahlentherapie. Besonders bei Kindern unter drei Jahren kann die kraniale Bestrahlung jedoch zu erheblichen kognitiven Einbußen führen, was unter anderem anhand der Messung des Intelligenzquotienten objektiviert werden kann. Bisher existieren jedoch kaum experimentelle Daten, die belegen, inwieweit Bestrahlung zu einer Reduktion neuraler Vorläuferzellen im Gyrus dentatus führt beziehungsweise die sowohl Kurzzeit- also auch Langzeiteffekte der Bestrahlung dosisabhängig untersuchen. Hinsichtlich der Prävention bestrahlungsinduzierter kognitiver Defizite konnten experimentelle Studien zeigen, dass die Veränderung von Bestrahlungsparametern, der Bestrahlungstechnik, die Gabe neuroprotektiver Substanzen sowie die Wiederherstellung des neuralen Stammzellpools zu einer Verbesserung der kognitiven Eigenschaften führen können. Obwohl Resveratrol als neuroprotektive Substanz bei ischämischen Läsionen des Gehirns sowie einigen neurodegenerativen Erkrankungen bekannt ist, wurde sein neuroprotektives Potential bei radiogen verursachten Läsionen des Gehirns bisher noch nicht näher untersucht. Daher bleibt es offen zu klären, ob die neuralen Vorläuferzellen des Hippokampus, die für komplexe Denkleistungen sowie die Regeneration geschädigter Zellen nötig sind, durch die Gabe dieser Substanz vor dem Zelltod durch Bestrahlung gerettet werden können. In der hier vorliegenden Arbeit war es das Ziel, den Einfluss verschiedener Dosiskonzepte auf das Überleben der neuralen Vorläuferzellen des Gyrus dentatus sowie die Möglichkeit einer Protektion dieser Zellen vor bestrahlungsinduziertem Zelluntergang durch Resveratrol zu untersuchen. Die Experimente wurden mit drei bis sechs Tage alten transgenen Nestin-CFPnuc C57BL/J6 Mäusen durchgeführt, von denen Gewebekulturscheiben, welche die entorhino-hippokampale Formation enthielten, angelegt wurden. Um mit Gewebekulturscheiben arbeiten und reproduktive sowie valide Ergebnisse erzielen zu können, war es von Bedeutung, dass während des gesamten Präparierprozesses die entorhino-hippokampale Formation in den Gehirnen erhalten bleibt. Die Hämytoxylinfärbung zeigte, dass die Zytoarchitektur einer Hirnscheibe in vitro der eines Ganzhirnpräparates entspricht. Insbesondere der Tractus perforans bleibt intakt (Kluge et al., 1998). Die Messung der Zytokine (IL-6, KC, MCP-1) nach Entnahme der Hirne und Präparation ergab 14 Tage nach Gewebepräparation konstante Messwerte, sodass von einem Abschluss der Wundheilung ausgegangen werden konnte. Verhalten Nestin-positiver Zellen in Kultur Der Zeitverlauf Nestin-positiver Zellen in unbestrahlten und unbehandelten Kontrollen über 49 Tage zeigte eine mehrphasische Entwicklung, die in dieser Form bisher noch nie beschrieben worden ist. Bis etwa zur Hälfte der Zeit gab es einen deutlichen Abfall Nestin-positiver Zellen. Dieser schien sich jedoch innerhalb der nächsten zehn Tage zu erholen und ab dem 35. Tag in vitro einen konstanten Verlauf zu zeigen. Diese Beobachtungen beweisen, dass die Gewebekulturscheiben in vitro ähnlich einer Maus in vivo altern (Fukuda et al., 2005). Effekte der Bestrahlung Die Bestrahlung führte über den Zeitraum von sechs Wochen zu einer signifikanten, irreversiblen und dosisabhängigen Reduktion Nestin-positiver Zellen innerhalb aller Bestrahlungsdosen (4,5 Gy, 8 Gy, 12 Gy und 16 Gy). Eine Erklärung für die Abnahme Nestin-positiver Zellen ist der bestrahlungsinduzierte Zelltod, welcher, gemessen an Propidiumiodid-positiven Zellen, einen dosisabhängigen Anstieg zeigte. Aufgrund der geringen absoluten Anzahl Propidiumiodid-positiver Zellen 96 Stunden nach Bestrahlung ist es denkbar, dass der Zelltod womöglich auch zu einem späteren Zeitpunkt einsetzt, wenn sich die durch die Bestrahlung entstandenen Aberrationen und nicht mehr reparablen Schäden über mehrere Zellzyklen manifestiert haben und dann zum Tod der Zelle führen. Entsprechend unseren Ergebnissen führt Bestrahlung ebenso zu einem Einbruch der Neurogenese (NeuN/BrdU) sowie der Proliferation (BrdU/ Ki-67). Diese Beobachtungen machten auch andere Arbeitsgruppen, die ebenfalls negative Langzeiteffekte auf die Neurogenese (Mizumatsu et al., 2003) sowie die Proliferation (Rola et al., 2004) beschrieben. Die Ausdifferenzierung der Stammzellen zu GFAP-positiven Zellen kann durch die Induktion von NF- κB (Ozeki et al., 2012), STAT-3 (Bonni et al., 1997) oder die Aktivierung der Mikroglia im Sinne einer reaktiven Gliosis (Hwang et al., 2006) hervorgerufen werden. Entgegen der Annahme, dass es auch zu einem bestrahlungsinduzierten Abfall ausgereifter Neurone kommen müsste, führte Bestrahlung bis 12 Gy nach 42 Tagen eher zu einer Zunahme. Allerdings ist nicht auszuschließen, dass es unmittelbar nach Bestrahlung zu einem Abfall gekommen ist, der sich aber nach 42 Tagen vollständig erholt hat. Dies könnte auch den Abfall bei einer Dosis von 16 Gy erklären, da in diesem Dosisbereich eben keine Regeneration mehr erwartet werden kann. Effekt von Resveratrol Da es bereits bei einer Einzeldosis von 4,5 Gy zu einem irreversiblen Abfall neuronaler Stammzellen sowie einem Einbruch der Proliferation und Neurogenese kam, sollten neben technischen Maßnahmen zur Neuroprotektion auch andere Möglichkeiten in Betracht gezogen werden. Eine Einzeldosis von 4,5 Gy entspricht in etwa einer Dosis von 10 Gy, die im Rahmen des „hippocampal sparing“ während der fraktionierten Behandlung typischerweise im Bereich des Hippokampus erreicht wird (Gondi et al., 2010b). Die Gabe von Resveratrol führte in unbestrahlten Gewebekulturscheiben zu einer Abnahme Nestin-positiver Zellen im Vergleich zu unbehandelten Kontrollen. In Kombination mit Bestrahlung zeigte sich jedoch, dass die Anzahl Nestin-positiver Zellen durch die Gabe von Resveratrol zum Teil signifikant erhöht werden konnte. Das neuroprotektive Potential von Resveratrol konnte bereits bei 4 Gy bestrahlten hippokampalen Neuronen demonstriert werden (Li et al., 2014). Bei einer Bestrahlungsdosis von 8 Gy, nicht jedoch bei 16 Gy beobachteten wir über den gesamten Zeitraum einen neuroprotektiven Effekt. Es ist anzunehmen, dass die durch 16 Gy hinterlassenen Schäden derart stark ausgeprägt sind, dass Resveratrol, welches nur 48 Stunden verabreicht wurde, nicht in der Lage war, die Stammzellen zu schützen. In der Praxis könnte man Resveratrol über längere Zeit im Rahmen fraktionierter Behandlungen verabreichen, was auch eine Reduktion der Resveratroldosis ermöglichen könnte. In den unbestrahlten Kontrollen führte die Gabe von Resveratrol möglicherweise durch die Aktivierung von Sirt-1 sowie von AMPK zu einer Inhibition der Entwicklung von Stammzellen (Ma et al., 2014; Park et al., 2012). In weiteren Studien sollte geprüft werden, inwieweit Resveratrol auch in vivo Potential besitzt, die neurokognitiven Fähigkeiten durch einen Schutz der Stammzellen vor Bestrahlung zu verbessern.:Inhaltsverzeichnis Abkürzungsverzeichnis II 1. Einführung 1.1 Hintergrund 1.2 Primäre ZNS-Tumore im Kindesalter 1.2.1 Entstehung und Ätiologie 1.2.2 Symptome und Diagnostik 1.2.3 Therapiemodalitäten 1.3 Der Hippokampus 1.3.1 Anatomie der hippokampalen Formation 1.3.2 Funktion des Hippokampus 1.3.3 Neurogenese im Hippokampus 1.4 Beschreibung des Mausmodels sowie der Zellkultur 1.5 Biologische Wirkung ionisierender Strahlung auf Gewebe 1.6 Neuroprotektion vor ionisierender Strahlung 1.7 Zielsetzung der Arbeit 2. Publikation 3. Zusammenfassung 4. Literaturverzeichnis 5. Darstellung des eigenen Beitrags 6. Anhang 36 6.1 Selbstständigkeitserklärung 36 6.2 Lebenslauf 37 6.3 Publikationen 39 6.4 Danksagung 40

Resveratrol (RV) is a naturally occurring phytoalexin of the stilbenoid family produced by some plant species, and present in grape skin, peanuts, and red wine. It has been found to exhibit anti-cancer, anti-inflammatory, anti-viral, anti-aging, cardio protective, and anti-oxidant properties. Bioavailability is a huge setback that limits the potentials of RV. As a result, efforts have been made to design and synthesize RV esters and aliphatic acids in an attempt to increase its bioavailability, solubility in water, and possibly improving its biological activities. Resveratrol esters, 3,5,4'-triacetyloxystilbene (2) and Methyl 1,1',1''- (3,4',5-stilbenyl)-1,6-hexanedioate (3) have been synthesized. Compound 3 is a new compound, synthetic yield is 88%, and purity is above 95% based on NMR integration. Both 2 and 3 are good candidates for biological evaluation. 3 was used as a precursor in the synthesis of resveratrol aliphatic acid, 8-(3',5'-dihydroxylstilbene-4''-oxy)-3,6-dioxocotanoic acid (9). First, 2 was hydrolyzed to resveratrol diester, 3,5-diacetyloxystilbene (4). Mitsunobu reaction of 4 and methyl 8-hydroxy-3,6-dioxooctanoate (7) was then carried out to afford methyl 8-(3',5'-diacetyoxystilben-4''-oxy)-3.6-dioxooctanoate (/5), which was then hydrolyzed to afford 9 in total 43.6 % yield. Structures of all newly synthesized compounds were confirmed by 1H and 13C NMR spectroscopy.

Resveratrol, a naturally derived stilbene, is an interesting compound mostly talked about recently because for its anti-cancer properties. Unfortunately it has some shortcomings due to its low bioavailability and low solubility in water. For this reason, my research is to overcome resveratrol's drawbacks by improving its bioavailability and hydrophilicity. My research is focused on syntheses of novel derivatives of resveratrol such as 3, 5-di-O-isobutyroyl resveratrol and 3, 5-di-O-hexanoyl resveratrol using lipase catalyzed hydrolysis. Therefore, the tri-acylated resveratrols 3, 5, 4'-tri-O-isobutyroyl resveratrol and 3, 5,4'-tri-O-hexanoyl resveratrol were first synthesized. 3,5,4'-tri-O-isobutyroyl resveratrol and 3,5,4'-tri-O-hexanoyl resveratrol were then hydrolyzed using lipase (C. Antarctica) to obtain the products 3,5-di-O-isobutyroyl resveratrol and 3,5-di-O-hexanoyl resveratrol. The four compounds 3,5-di-O-isobutyroyl resveratrol, 3,5-di-O-hexanoyl resveratrol, 3,5,4'-tri-O-hexanoyl resveratrol, and 3,5-di-O-hexanoyl resveratrol were characterized by 1H NMR and 13C NMR.

4’ -ester analogues of resveratrol were synthesized using reaction conditions developed to produce mono-ester products in the presence of two other unprotected phenols. Basic conditions were employed to deprotonate the most acidic 4’ phenol followed by addition of anhydrides or acid chlorides to give the ester product. The reaction favored 4’-ester formation in polar aprotic solvents with DMSO being the optimal solvent. (—)-Englerin A is a guaiane-type sesquiterpene containing two ester side chains. Mimics of (—)-englerin A were proposed that retained the ester side chains while replacing the non-polar core with less complicated structures. These proposed mimic cores would maintain the three-dimensional positioning of the esters which are responsible for the anti-cancer activity of (—)-englerin A. Three mimics were synthesized using the bicyclic terpenes borneol and fenchol. Installation of the second ester on the terpene core was accomplished throught the development and optimization of a unique methylene oxidation using chromium trioxide in glacial acetic acid. These mimics were screened against two kidney cancer cell lines. The compounds were shown to have IC50 (inhibitory concentration for 50 % of cells) values above 30 µM.

The phytoalexin resveratrol and its acetyl analogs have been made using a decarbonylative Heck reaction. The acid chloride derived from 3,5-dihydroxybenzoic acid was coupled with suitable protected 4-hydroxystyrene in the presence of palladium acetate and N,N-bis-(2,6-diisopropylphenyl)-4,5-dihydro imidazolium chloride to give the substituted stilbene in good yield as the key step. Human HL-60 cell assays showed the 4'-acetyl resveratrol variant improved activity (ED50 17 μM) relative to resveratrol (24 μM). Cinchona phase-transfer catalysts (PTC) were developed for glycolate aldol reactions to give differentially protected 1,2-diol products. Silyl enol ether of diphenylmethoxy-2,5-dimethoxyacetophenone reacted to generate benzhydryl-protected products. O-Allyl trifluorobenzyl cinchonium hydrodifluoride (20 mol %) catalyzed the addition of the silyl enol ether to benzaldehyde to give aldol product as a single syn-product in 76% yield and 80% ee. Recrystallization enriched the product to 95% ee, and a Baeyer-Villiger reaction transformed the product into useful ester intermediates. A novel unnatural product, 8,9-Methylamido-Geldanamycin, has been designed and synthesized. Using a convergent route, the total synthesis of the molecule involved only 27 longest linear steps. New synthesis methodologies, including auxiliary controlled asymmetric anti-glycolate aldol, syn-norephedrine aldol, and selective p-quinone formation, were used.

One of the matchless ironies of the human body is its requirement for the highly reactive oxygen molecule, which has been clearly implicated in many diseases and the aging processes. Oxidants produced by metabolic processes damage cells by starting chemical chain reactions including oxidation of DNA and proteins as well as lipid peroxidation. Damage to DNA can cause mutations and lead to cancer if not reversed by DNA repair mechanisms. Damage to proteins causes enzyme inhibition, denaturation and protein degradation. Lipid peroxidation can cause cell lysis as well as creating mutagenic and carcinogenic by-products. The human body contains antioxidants and enzymes that together work to prevent oxidative damage to cellular components. By and large antioxidants either prevent these reactive oxygen species from being formed or remove them before they cause damage. There are many theories currently that tout the superior nature of diverse antioxidant combinations. One such theory is by Dr. Lester Packer of The University of California at Berkley. Dr. Packer puts forth the hypothesis that there is a superlative combination of five antioxidants that have the ability to "recharge" one another both in the blood plasma and intracellularly. This would result in a greater quality of antioxidant protection for an extended time. The current study evaluates Dr. Packer's theory of antioxidant combination from his book The Antioxidant Miracle. The decay rate of the antioxidants vitamin E, vitamin C, lipoic acid, glutathione, and coenzyme Q10 alone and in combination were determined using the ORAC (Oxygen Radical Absorbance Capacity) assay. The majority of the antioxidants retained activity for longer periods of time when tested alone, rather than in combination as Dr. Packer's theory would suggest. The assay was also preformed (using the same antioxidants and combinations) on oxidatively damaged Raji cancer cells. Cell viability and uptake of antioxidants into the cytoplasm were monitored. Finally, a variety of multivitamins were subjected to the ORAC assay and their antioxidant capacity compared to that of the "Packer Combination". The results suggest that multivitamins are superior antioxidants than the Packer ratio listed in The Antioxidant Miracle.

No description available.

No description available.

Sirtuin-1 (SirT1) is a nicotinamide adenine dinucleotide (NAD+)-dependent lysine deacetylase expressed ubiquitously in the body. In the vasculature, SirT1 is present in endothelial and vascular smooth muscle cells (VSMCs), where it has been shown to promote anti-inflammatory and anti-oxidant effects. As a result, SirT1 is known to play a protective role in the vasculature wall from pathologies such as atherosclerosis, arterial stiffness, and aortic aneurysm. Hence, SirT1 is considered an attractive therapeutic target for vascular diseases and potentially, aging-related and metabolic diseases. However, currently available SirT1 activators have failed to reach the clinic. Thus, novel approaches to activate SirT1 are needed. In this study, we first sought to optimize a novel fluorescence-based SirT1 activity assay, with which to reliably assess intracellular SirT1 activity and the efficacy of SirT1 activators and inhibitors. We next sought to use the SirT1 activity assay to screen novel compounds identified by an in silico docking analysis and hypothesized to activate SirT1. Lastly, we generated adeno-associated viruses (AAV) overexpressing wildtype (WT) or a redox-resistant (3M) SirT1 to analyze the effects of overexpressing SirT1 in VSMCs, in normal and oxidative stress conditions. For the activity assay, our results showed that an optimal standard curve range was between 0 ng and 12 ng of substrate (acetylated-p53 peptide). After testing different commercially available human recombinant SirT1s, the Anaspec SirT1 of the highest concentration showed a decrease in measured fluorescence for acetylated-p53 peptide with higher SirT1 (ng), indicating the enzyme and the assay were functional. However, when novel small molecules (A4, B4, and G3) hypothesized to activate SirT1 were added to reactions, the total p53 peptide fluorescence values increased compared to the control, suggesting some interference of the molecules with the assay detection. After AAV infection in VSMCs, SirT1 expression, measured by HA-tag, increased for AAV WT (n=3, p=0.04) and similarly for AAV 3M SirT1, indicating that the AAVs efficiently infect VSMCs. SirT1 activity, measured by Western Blot as decreased acetylated-histone (H3), also appeared to increase for both AAV WT and AAV 3M. A similar trend was shown for VSMCs under oxidant stress conditions (n=2). In conclusion, we successfully established a standard curve range for a novel SirT1 activity assay. Further trials are needed to ensure activity assay reproducibility before testing the efficacy of SirT1 activators and inhibitors. Infection of AAV WT and 3M SirT1 led to an increase in the expression and activity of SirT1 in VSMCs. The expression of SirT1 by AAV may be a promising therapeutic option for in vivo prevention and treatment of vascular diseases. / 2026-03-01T00:00:00Z