AQ4N (banoxantrone) is a prodrug that, under hypoxic conditions, is enzymatically converted to a cytotoxic DNA-binding agent, AQ4. Incorporation of AQ4N into conventional chemoradiation protocols therefore targets both oxygenated and hypoxic regions of tumors, and potentially will increase the effectiveness of therapy. This current pharmacodynamic and efficacy study was designed to quantify tumor exposure to AQ4 following treatment with AQ4N, and to relate exposure to outcome of treatment. A single dose of 60 mg/kg AQ4N enhanced the response of RT112 (bladder) and Calu-6 (lung) xenografts to treatment with cisplatin and radiation therapy. AQ4N was also given to separate cohorts of tumor-bearing mice 24 hours before tumor excision for subsequent analysis of metabolite levels. AQ4 was detected by high performance liquid chromatography/mass spectrometry in all treated samples of RT112 and Calu-6 tumors at mean concentrations of 0.23 and 1.07 microg/g, respectively. These concentrations are comparable with those shown to be cytotoxic in vitro. AQ4-related nuclear fluorescence was observed in all treated tumors by confocal microscopy, which correlated with the high performance liquid chromatography/mass spectrometry data. The presence of the hypoxic marker Glut-1 was shown by immunohistochemistry in both Calu-6 tumors and RT112 tumors, and colocalization of AQ4 fluorescence and Glut-1 staining strongly suggested that AQ4N was activated in these putatively hypoxic areas. This is the first demonstration that AQ4N will increase the efficacy of chemoradiotherapy in preclinical models; the intratumoral levels of AQ4 found in this study are comparable with tumor AQ4 levels found in a recent phase I clinical study, which suggests that these levels could be potentially therapeutic.

<p>&Scaron;tavelj (Rumex crispus, Polygonaceae) je vi&scaron;egodi&scaron;nja zeljasta biljka, koja predstavlja bogat izvor fenolnih komponenti. Iako se smatra invazivnim korovom, mlado li&scaron;će &scaron;tavelja je jestivo i često se koristi kao salata. Dalje, upotreba plodova &scaron;tavelja opisana je u srpskoj i turskoj narodnoj medicini u lečenju gastrointestinalnih tegoba. Cilj ovog rada bio je procena in vitro i in vivo antioksidantne/prooksidantne i citotoksične aktivnosti, i određivanje eventualnog in vitro antiinflamatornog efekta vodenog ekstrakta ploda Rumex crispus. Ukupan sadržaj flavonoida određen je spektrofotometrijskom metodom. Kvalifikacija i kvantifikacija flavonoida potvrđena je visokoefikasnom tečnom hromatografijom (HPLC). Antioksidantna aktivnost vodenog ekstrakta ploda &scaron;tavelja procenjena je na osnovu in vitro testova: Ferric-reducing antioxidant power (FRAP), sposobnosti ekstrakta da neutrali&scaron;e slobodne radikale NO&bull;, OH&bull; i DPPH&bull; i uticaja na lipidnu peroksidaciju u lipozomima. Citotoksičnost ispitivanog ekstrakta je određena in vitro na tumorskim ćelijskim linijama: humani karcinom cerviksa (HeLa), adenokarcinom (HT-29) i adenokarcinom dojke (MCF7). Takođe, moguća in vivo hepatoprotektivna i antioksidantna svojstva ekstrakta određena su kod oksidativnog stresa izazvanog CCl4 kod eksperimentalnih životinja. Pored toga, proverena je hipoteza u kojoj testiran ekstrakt pokazuje in vivo antiproliferativnu aktivnost kod Ehrlich-ovih (EAC) i Hepatoma AS30D ćelija, merenjem zapremine ascitesa, procenta vijabilnih ćelija i nivoa nekoliko antioksidantnih enzima. Optimizovan in vitro test za određivanje potencijala inhibicije ciklooksigenaze-1 (COX-1) i 12-lipooksigenaze (12-LOX) preduzet je u svrhu procene antiinflamatornog efekta vodenog ekstrakta ploda R. crispus. HPLC analiza otkrila je da je mikvelianin najdominantniji flavonoidni konstituent ekstrakta. Testirani ekstrakt pokazao je potencijalnu antioksidantnu aktivnost rezultujući velikom moći u neutralizaciji slobodnih radikala, i sposobno&scaron;ću da smanji lipidnu peroksidaciju u lipozomima. Rezultati su ukazali na tkivno-selektivnu citotoksičnost ekstrakta ploda R. crispus in vitro. Najizraženija antitumorska aktivnost primećena je prema HeLa i MCF7 ćelijskim linijama. Podaci sugeri&scaron;u da bi se ispitivani ekstrakt mogao smatrati potencijalnim in vivo hepatoprotektivnim i antioksidantnim agensom, sprečavajući oksidativna o&scaron;tećenja jetre. S druge strane, pomenuti ekstrakt može pokazati in vivo prooksidantna svojstva, uzrokujući oksidativni stres u maligno transformisanim EAC i AS30D ćelijama i smanjujući zapreminu ascitesa i udeo vijabilnih ćelija, u poređenju sa kontrolnom grupom. Promene u aktivnosti antioksidantnih enzima su verovatno posledica indukovanog oksidativnog stresa u EAC i AS30D ćelijama, naročito kod pretretiranih životinja. Vodeni ekstrakt ploda &scaron;tavelja pokazao je COX-1, kao i 12-LOX inhibitornu aktivnost, navodeći da bi ispitivani ekstrakt mogao biti antiinflamatorni agens. Vodeni ekstrakt ploda R. crispus ima potencijalnu antioksidantnu, citotoksičnu i antiinflamatornu aktivnost. Ispoljavanje prooksidantnih svojstava predstavlja mogući mehanizam antiproliferativnog efekta ekstrakta.</p> / <p>Curly dock (Rumex crispus, Polygonaceae) is a wild perennial herbaceous plant, which products are described as a rich source of phenolic compounds. Apart from being considered a seriously invasive weed, young leaves of curly dock are edible and often used as salad. Furthermore, the use of its fruits has been described in Serbian and Turkish traditional medicine against stomach complaints. The objectives of this study were to evaluate in vitro and in vivo antioxidant/prooxidant and cytotoxic activities, and to determine an eventual in vitro anti-inflammatory effect of the aqueous extract of Rumex crispus fruits. Total flavonoid content was determined by spectrophotometric method. Qualification and quantification of flavonoids were confirmed using High performance liquid chromatography (HPLC). The aqueous extract of curly dock fruits was evaluated for its antioxidant activity by in vitro assays for Ferric-reducing antioxidant power (FRAP), NO&bull;, OH&bull; and DPPH&bull;-free radical scavenging activities and the influence on lipid peroxidation in liposomes. The cytotoxicity of tested extract was examined in vitro in human cervix carcinoma (HeLa), colon adenocarcinoma (HT-29) and breast adenocarcinoma (MCF7). Also, the potential in vivo hepatoprotective and antioxidant properties of investigated extract were determined on CCl4-induced oxidative stress in experimental animals. Furthermore, the hypothesis that the examined extract might show in vivo antiproliferative activity in Ehrlich carcinoma (EAC) and Hepatoma AS30D cells was tested by measuring volume of ascites, percentage of viable cells and level of several antioxidant enzymes. The optimized in vitro test for determination of cyclooxygenase-1 (COX-1) and 12-lipoxygenase (12-LOX) inhibition potency was undertaken in order to estimate an anti-inflammatory effect of aqueous extract of R. crispus fruits. HPLC analysis revealed miquelianin as the most abundant flavonoid constituent of the extract. The tested extract might have an antioxidant activity resulting in scavenging of free radicals and ability to decrease lipid peroxidation in liposomes. The results could indicate tissue-selective cytotoxicity of R. crispus fruit extract in vitro. The most prominent antitumor activity was observed towards HeLa and MCF7 cell lines. The data suggested that investigated extract may be considered as potential in vivo hepatoprotective and antioxidant agent due to prevention of the liver injuries induced by oxidative damage. On the other hand, mentioned extract could exhibit in vivo prooxidant property, causing the oxidative stress in malignant transformed EAC and AS30D cells and reducing volume of ascites and percentage of viable cells, in comparison with control group. Changes in activities of antioxidant enzymes might be the results of induced oxidative stress in EAC and AS30D cells, especially in the pretreated animals. The aqueous extract of curly dock fruits showed COX-1, as well as 12-LOX inhibitory activity, suggesting that tested extract might be an anti-inflammatory agent. It could be concluded that aqueous fruit extract of R. crispus might have antioxidant, cytotoxic and anti-inflammatory activities. The prooxidant properties of examined extract could be the mechanism of potential antiproliferative effect of extract.</p>