Novel titanocene anti-cancer drugs and their effect on apoptosis and the apoptotic pathway in prostate cancer cells

Advanced prostate cancer is not curable by current treatment strategies indicating a significant need for new chemotherapeutic options. Highly substituted ansatitanocene compounds have shown promising cytotoxic activity in a range of cancers. The objectives of this study are to examine the effects of these titanocene compounds on prostate cancer cells. Prostate cell lines were treated with three novel titanocene compounds and compared to titanocene dichloride and cisplatin. Percent apoptosis, viability and cell cycle were assessed using propidium iodide DNA incorporation with flow cytometry. Cytochrome C was assessed by western blotting of mitochondrial and cytoplasmic fractions. Apoptosis Inducing Factor was assessed by confocal microscopy. These novel compounds induced more apoptosis compared to cisplatin in a dose dependent manner. Compound Y had the most significant effect on cell cycle and apoptosis. Despite the release of cytochrome C from the mitochondrial fraction there was no inhibition of apoptosis with the pan caspase inhibitor, ZVAD-FMK. AIF was shown to translocate from the cytosol to the nucleus mediating a caspase independent cell death. Bcl-2 over expressing PC-3 cells, which were resistant to cisplatin induced apoptosis, underwent apoptosis following treatment with all the titanocene compounds. This study demonstrates possible mechanisms by which these novel titanocene compounds can mediate their apoptotic effect in vitro. The fact that they can induce more apoptosis than cisplatin in advanced cancer cell lines would confer an advantage over cisplatin. They represent exciting new agents with future potential for the treatment of advanced prostate cancer.

Patents and Haploid Plants

One of the important themes in any discussion concerning the application of haploids in agricultural biotechnology or elsewhere is the role of Intellectual Property Rights (IPR). This term covers both the content of patents and the confidential expertise, usually related to methodology and referred to as "Trade Secrets". This review will explain the concepts behind patent protection, and will use the international patent databases to analyse the content of these patents and trends over the last 20 years. This analysis from regions including North America, Europe, and Asia reveals a total of more than 30 granted patents and a larger number of applications. The first of these patents dates from 1986, and although the peak of activity was in the late 1990s, there has been continuous interest to the present day. The subject matter of these patents and applications covers methods for anther and pollen culture, ovule culture, the use of specific haploid-inducing genes, the use of haploids as transformation targets, and the exploitation of genes that regulate embryo development. The species mentioned include cereals, vegetables, flowers, spices and trees.

Patents and transgenic plants

One of the recurring themes in any discussion concerning the application of genetic transformation technology is the role of Intellectual Property Rights (IPR). This term covers both the content of patents and the confidential expertise, usually related to methodology and referred to as “Trade Secrets”. This review will explain the concepts behind patent protection, and will discuss the wide-ranging scope of existing patents that cover all aspects of transgenic technology, from selectable markers and novel promoters to methods of gene introduction. Although few of these patents have any significant commercial value, there are a small number of key patents that may restrict the “freedom to operate” of any company seeking to exploit the methods. Over the last twenty years, these restrictions have forced extensive cross-licensing between ag-biotech companies and have been one of the driving forces behind the consolidation of these companies. Although such issues are often considered to be of little interest to the academic scientist working in the public sector, they are of great importance in any debate about the role of “public-good breeding” and of the relationship between the public and private sectors.

Techniques for analysis of proteins by SDS-polyacrylamide gel electrophoresis and Western blotting

The separation of mixtures of proteins by SDS-polyacrylamide gel electrophoresis (SDS-PAGE) is a technique that is widely used—and, indeed, this technique underlies many of the assays and analyses that are described in this book. While SDS-PAGE is routine in many labs, a number of issues require consideration before embarking on it for the first time. We felt, therefore, that in the interest of completeness of this volume, a brief chapter describing the basics of SDS-PAGE would be helpful. Also included in this chapter are protocols for the staining of SDS-PAGE gels to visualize separated proteins, and for the electrotransfer of proteins to a membrane support (Western blotting) to enable immunoblotting, for example. This chapter is intended to complement the chapters in this book that require these techniques to be performed. Therefore, detailed examples of why and when these techniques could be used will not be discussed here.

Effect of earthworm Eisenia fetida and wetland plants on nitrification and denitrification potentials in vertical flow constructed wetland.

The response of nitrification potentials, denitrification potentials, and N removal efficiency to the introduction of earthworms and wetland plants in a vertical flow constructed wetland system was investigated. Addition of earthworms increased nitrification and denitrification potentials of substrate in non-vegetated constructed wetland by 236% and 8%, respectively; it increased nitrification and denitrification potentials in rhizosphere in vegetated constructed wetland (Phragmites austrail, Typha augustifolia and Canna indica), 105% and 5%, 187% and 12%, and 268% and 15% respectively. Denitrification potentials in rhizosphere of three wetland plants were not significantly different, but nitrification potentials in rhizosphere followed the order of C. indica > T. augustifolia > P. australis when addition of earthworms into constructed wetland. Addition of earthworms to the vegetated constructed significantly increased the total number of bacteria and fungi of substrates (P < 0.05). The total number of bacteria was significantly correlated with nitrification potentials (r = 913, P < 0.01) and denitrification potentials (r = 840, P < 0.01), respectively. The N concentration of stems and leaves of C. indica were significantly higher in the constructed wetland with earthworms (P < 0.05). Earthworms had greater impact on nitrification potentials than denitrification potentials. The removal efficiency of N was improved via stimulated nitrification potentials by earthworms and higher N uptake by wetland plants.

Drought and resprouting plants

Many species have the ability to resprout vegetatively after a substantial loss of biomass induced by environmental stress, including drought. Many of the regions characterised by ecosystems where resprouting is common are projected to experience more frequent and intense drought during the 21st Century. However, in assessments of ecosystem response to drought disturbance there has been scant consideration of the resilience and post-drought recovery of resprouting species. Systematic differences in hydraulic and allocation traits suggest that resprouting species are more resilient to drought-stress than nonresprouting species. Evidence suggests that ecosystems dominated by resprouters recover from disturbance more quickly than ecosystems dominated by nonresprouters. The ability of resprouters to avoid mortality and withstand drought, coupled with their ability to recover rapidly, suggests that the impact of increased drought stress in ecosystems dominated by these species may be small. The strategy of resprouting needs to be modelled explicitly to improve estimates of future climate-change impacts on the carbon cycle, but this will require several important knowledge gaps to be filled before resprouting can be properly implemented.

The effect of carrier substrate, dose rate and time of application on biocontrol efficacy of fungal antagonists against Armillaria root rot of strawberry plants.

In a glasshouse experiment using potted strawberry plants (cv. Cambridge Favourite) as hosts, the effect of selected fungal antagonists grown on 25 or 50 g of mushroom compost containing autoclaved mycelia of Agaricus bisporus, or wheat bran was evaluated against Armillaria mellea. Another glasshouse experiment tested the effect of application time of the antagonists in relation to inoculations with the pathogen. A significant interaction was found between the antagonists, substrates and dose rates. All the plants treated with Chaetomium olivaceum isolate Co on 50 g wheat bran survived until the end of the experiment which lasted 482 days, while none of them survived when this antagonist was added to the roots of the plants on 25 g wheat bran or 25 or 50 g mushroom compost. Dactylium dendroides isolate SP had a similar effect, although with a lower host survival rate of 33.3%. Trichoderma hamatum isolate Tham 1 and T. harzianum isolate Th23 protected 33.3% of the plants when added on 50 g and none when added on 25 g of either substrate, while 66.7% of the plants treated with T. harzianum isolate Th2 on 25 g, or T viride isolate TO on 50 g wheat bran, survived. Application of the antagonists on mushroom compost initially resulted in development of more leaves and healthier plants, but this effect was not sustained. Eventually, plants treated with the antagonists on wheat bran had significantly more leaves and higher health scores. The plants treated with isolate Th2 and inoculated with Armillaria at the same time had a survival rate of 66.7% for the duration of the experiment (475 days), while none of them survived that long when the antagonist and pathogen were applied with an interval of 85 days in either sequence. C. olivaceum isolate Co showed a protective effect only, as 66.7% of the plants survived when they were treated with the antagonist 85 days before inoculation with the pathogen, while none of them survived when the antagonist and pathogen were applied together or the infection preceded protection.

Colorectal cancer and the relationship between genes and the environment

Colorectal cancer (CRC) is a significant cause of morbidity and mortality in developed countries, with both genetic and environmental factors contributing to the etiology and progression of the disease. Several risk factors have been identified, including positive family history, red meat intake, smoking, and alcohol intake. Protective factors include vegetables, calcium, hormone replacement therapy, folate, nonsteroidal anti-inflammatory drugs, and physical activity. The interaction between these environmental factors, in particular diet and genes, is an area of growing interest. Currently, oncogenes, tumor suppressor genes, and mismatch repair genes are believed to play an essential role in colorectal carcinogenesis. When considering the genetics of CRC, only 10% of cases are inherited and only 2-6% can be ascribed to the highly penetrant genes, such as APC, hMLH and hMSH2. Lower penetrance genes combined with a Western-style diet contribute to the majority of sporadic CRCs. The purpose of this article is to give a brief overview of the epidemiologic studies that have been conducted and present the major findings. Here, we examine the molecular events in CRC, with particular focus on the interaction between genes and environment, and review the most current research in this area.

Changes in gene expression in Arabidopsis shoots during phosphate starvation and the potential for developing smart plants

Our aim was to generate and prove the concept of "smart" plants to monitor plant phosphorus (P) status in Arabidopsis. Smart plants can be genetically engineered by transformation with a construct containing the promoter of a gene up-regulated specifically by P starvation in an accessible tissue upstream of a marker gene such as beta-glucuronidase (GUS). First, using microarrays, we identified genes whose expression changed more than 2.5-fold in shoots of plants growing hydroponically when P, but not N or K, was withheld from the nutrient solution. The transient changes in gene expression occurring immediately (4 h) after P withdrawal were highly variable, and many nonspecific, shock-induced genes were up-regulated during this period. However, two common putative cis-regulatory elements (a PHO-like element and a TATA box-like element) were present significantly more often in the promoters of genes whose expression increased 4 h after the withdrawal of P compared with their general occurrence in the promoters of all genes represented on the microarray. Surprisingly, the expression of only four genes differed between shoots of P-starved and -replete plants 28 h after P was withdrawn. This lull in differential gene expression preceded the differential expression of a new group of 61 genes 100 h after withdrawing P. A literature survey indicated that the expression of many of these "late" genes responded specifically to P starvation. Shoots had reduced P after 100 h, but growth was unaffected. The expression of SQD1, a gene involved in the synthesis of sulfolipids, responded specifically to P starvation and was increased 100 h after withdrawing P. Leaves of Arabidopsis bearing a SQD1::GUS construct showed increased GUS activity after P withdrawal, which was detectable before P starvation limited growth. Hence, smart plants can monitor plant P status. Transferring this technology to crops would allow precision management of P fertilization, thereby maintaining yields while reducing costs, conserving natural resources, and preventing pollution.