Proapoptotic activity of Ukrain is based on Chelidonium majus L. alkaloids and mediated via a mitochondrial death pathway

BACKGROUND: The anticancer drug Ukrain (NSC-631570) which has been specified by the manufacturer as semisynthetic derivative of the Chelidonium majus L. alkaloid chelidonine and the alkylans thiotepa was reported to exert selective cytotoxic effects on human tumour cell lines in vitro. Few clinical trials suggest beneficial effects in the treatment of human cancer. Aim of the present study was to elucidate the importance of apoptosis induction for the antineoplastic activity of Ukrain, to define the molecular mechanism of its cytotoxic effects and to identify its active constituents by mass spectrometry. METHODS: Apoptosis induction was analysed in a Jurkat T-lymphoma cell model by fluorescence microscopy (chromatin condensation and nuclear fragmentation), flow cytometry (cellular shrinkage, depolarisation of the mitochondrial membrane potential, caspase-activation) and Western blot analysis (caspase-activation). Composition of Ukrain was analysed by mass spectrometry and LC-MS coupling. RESULTS: Ukrain turned out to be a potent inducer of apoptosis. Mechanistic analyses revealed that Ukrain induced depolarisation of the mitochondrial membrane potential and activation of caspases. Lack of caspase-8, expression of cFLIP-L and resistance to death receptor ligand-induced apoptosis failed to inhibit Ukrain-induced apoptosis while lack of FADD caused a delay but not abrogation of Ukrain-induced apoptosis pointing to a death receptor independent signalling pathway. In contrast, the broad spectrum caspase-inhibitor zVAD-fmk blocked Ukrain-induced cell death. Moreover, over-expression of Bcl-2 or Bcl-x(L )and expression of dominant negative caspase-9 partially reduced Ukrain-induced apoptosis pointing to Bcl-2 controlled mitochondrial signalling events. However, mass spectrometric analysis of Ukrain failed to detect the suggested trimeric chelidonine thiophosphortriamide or putative dimeric or monomeric chelidonine thiophosphortriamide intermediates from chemical synthesis. Instead, the Chelidonium majus L. alkaloids chelidonine, sanguinarine, chelerythrine, protopine and allocryptopine were identified as major components of Ukrain. Apart from sanguinarine and chelerythrine, chelidonine turned out to be a potent inducer of apoptosis triggering cell death at concentrations of 0.001 mM, while protopine and allocryptopine were less effective. Similar to Ukrain, apoptosis signalling of chelidonine involved Bcl-2 controlled mitochondrial alterations and caspase-activation. CONCLUSION: The potent proapoptotic effects of Ukrain are not due to the suggested "Ukrain-molecule" but to the cytotoxic efficacy of Chelidonium majus L. alkaloids including chelidonine

Nitrogen management guidelines for sugarcane production in Australia: can these be modified for wet tropical conditions using seasonal climate forecasting?

Sugarcane is a highly valuable crop grown in tropical and subtropical climates worldwide primarily for the production of sucrose-based products. The Australian sugarcane industry is located in close proximity to sensitive environments and the apparent declining health of the Great Barrier Reef has been linked to damaging levels of land-based pollutants entering reef waters as a result of sugarcane cultivation undertaken in adjacent catchments. Unprecedented environmental scrutiny of N fertiliser application rates is necessitating improved N fertiliser management strategies in sugarcane.Over time the focus of N fertiliser management has shifted from maximising production to optimising profitability and most recently to improved environmental sustainability. However, current N calculations are limited in their ability to match N fertiliser inputs to forthcoming crop requirements. Seasonal climate forecasts are being used to improve decision-making capabilities across different sectors of the sugarcane value chain. Climate is a key driver of crop growth, N demand and N loss processes, but climate forecasts are not being used to guide N management strategies. Seasonal climate forecasts could be used to develop N management strategies for 'wet' and 'dry' years by guiding application rate, timing and/or frequency of N inputs and the benefit of using alternative forms of N fertiliser. The use of seasonal climate forecasts may allow more environmentally sensitive yet profitable N management strategies to be developed for the Australian sugarcane industry