title: Elucidating the Modes of Action of the Antitumour New Zealand Marine Natural Product Pterocellin A

authors: Wang, Alice Tansi

subjects: natural product

abstract: Pterocellin A is a novel bioactive alkaloid isolated from the New Zealand marine bryozoan Pterocella vesiculosa. It exhibits potent antitumour activity towards the P388 (murine leukaemia) cell line in vitro with an IC₅₀ value of 477 ng/mL. Further screening at the NCI indicated that it was selectively sensitive towards certain non-small cell lung, melanoma, and breast cancer cell lines, however, the biological mode of action of pterocellin A is unknown.

This project investigated and elucidated the cytotoxicity of pterocellin A using the human cervical cancer cell line HeLa. Cellular toxicity can be manifested as either necrotic or apoptotic cell death. Cancer cells are prone to resisting apoptosis, and one of the key mechanisms recognised in anticancer agents is the induction of apoptosis. Pterocellin A exhibited similar cytotoxicity against the HeLa cell line compared to P388 cells with an IC₅₀ value of 886 ng/mL. In this study, HeLa cells were incubated with various concentrations (up to 2000 ng/mL) of pure pterocellin A, and characteristic markers of apoptotic and necrotic cell death were investigated using several in vitro biochemistry techniques. Assessments were made on the morphological and biochemical changes in HeLa cells after treatment with pterocellin A.

Time-course MTT and LDH assays were carried out and the results showed that only a low level of cytosolic LDH was detected in the supernatant after all the cells had died from pterocellin A treatment at 2000 ng/mL. This indicated the cells maintained membrane integrity upon death which suggested apoptosis. Additionally, morphological changes were observed under the microscope after six hours of treatment. Cell shrinkage and nucleus condensation were observed, as well as apparent membrane blebbing, a key feature of apoptosis. The MTT data were also indicative of mitochondria impairment which could suggest that pterocellin A targets the mitochondria. This idea was supported by the observed changes in the morphology and location of the mitochondria after exposure to pterocellin A. Furthermore, the level of activated caspase-3 in HeLa cells increased after treatment with pterocellin A; activated caspase-3 can only be detected after a series of signalling events following the induction of apoptosis. These data support the notion that pterocellin A is an inducer of apoptosis in HeLa cells. 

This project also utilised the biochemical assays to develop a systematic bioassay screening system at the University of Waikato, and further investigated the bioactive metabolites of P. vesiculosa. The same approach taken to study pure pterocellin A in HeLa cells was successfully applied to the bioassay development. The bioactive fractions separated from the crude extract were analysed by LC-MS and several unknown metabolites were observed, however due to time constraints associated with the project, no further investigation was carried out.

text: Elucidating the Modes of Action of the Antitumour New Zealand Marine Natural Product Pterocellin A Pterocellin A is a novel bioactive alkaloid isolated from the New Zealand marine bryozoan Pterocella vesiculosa. It exhibits potent antitumour activity towards the P388 (murine leukaemia) cell line in vitro with an IC₅₀ value of 477 ng/mL. Further screening at the NCI indicated that it was selectively sensitive towards certain non-small cell lung, melanoma, and breast cancer cell lines, however, the biological mode of action of pterocellin A is unknown.

This project investigated and elucidated the cytotoxicity of pterocellin A using the human cervical cancer cell line HeLa. Cellular toxicity can be manifested as either necrotic or apoptotic cell death. Cancer cells are prone to resisting apoptosis, and one of the key mechanisms recognised in anticancer agents is the induction of apoptosis. Pterocellin A exhibited similar cytotoxicity against the HeLa cell line compared to P388 cells with an IC₅₀ value of 886 ng/mL. In this study, HeLa cells were incubated with various concentrations (up to 2000 ng/mL) of pure pterocellin A, and characteristic markers of apoptotic and necrotic cell death were investigated using several in vitro biochemistry techniques. Assessments were made on the morphological and biochemical changes in HeLa cells after treatment with pterocellin A.

Time-course MTT and LDH assays were carried out and the results showed that only a low level of cytosolic LDH was detected in the supernatant after all the cells had died from pterocellin A treatment at 2000 ng/mL. This indicated the cells maintained membrane integrity upon death which suggested apoptosis. Additionally, morphological changes were observed under the microscope after six hours of treatment. Cell shrinkage and nucleus condensation were observed, as well as apparent membrane blebbing, a key feature of apoptosis. The MTT data were also indicative of mitochondria impairment which could suggest that pterocellin A targets the mitochondria. This idea was supported by the observed changes in the morphology and location of the mitochondria after exposure to pterocellin A. Furthermore, the level of activated caspase-3 in HeLa cells increased after treatment with pterocellin A; activated caspase-3 can only be detected after a series of signalling events following the induction of apoptosis. These data support the notion that pterocellin A is an inducer of apoptosis in HeLa cells. 

This project also utilised the biochemical assays to develop a systematic bioassay screening system at the University of Waikato, and further investigated the bioactive metabolites of P. vesiculosa. The same approach taken to study pure pterocellin A in HeLa cells was successfully applied to the bioassay development. The bioactive fractions separated from the crude extract were analysed by LC-MS and several unknown metabolites were observed, however due to time constraints associated with the project, no further investigation was carried out.

year: 2015

title: (The Futility of) Trying to Predict Carcinogenicity of Chemical Compounds

authors: Pfahringer, Bernhard

subjects: computer science

abstract: This paper describes my submission to one of the sub-problems formulated for the Predictive Toxicology Challenge 2001. The challenge is to predict the carcinogenicity of chemicals based on structural information only. I have only tackled such predictions for bioassays involving male rats. As we currently do not know the true predictions for the test-set, all we can say is that one of the models supplied by us seems to be optimal over some subrange of the ROC spectrum. The successful model uses a voting approach based on most of the sets of structural features made available by various other contestants as well as the organizers in an earlier phase of the Challenge. The WEKA Machine Learning workbench served as the core learning utility. Based on a preliminary examination of our submission we conclude that reliable prediction of carcinogenicity is still a far away goal.

text: (The Futility of) Trying to Predict Carcinogenicity of Chemical Compounds This paper describes my submission to one of the sub-problems formulated for the Predictive Toxicology Challenge 2001. The challenge is to predict the carcinogenicity of chemicals based on structural information only. I have only tackled such predictions for bioassays involving male rats. As we currently do not know the true predictions for the test-set, all we can say is that one of the models supplied by us seems to be optimal over some subrange of the ROC spectrum. The successful model uses a voting approach based on most of the sets of structural features made available by various other contestants as well as the organizers in an earlier phase of the Challenge. The WEKA Machine Learning workbench served as the core learning utility. Based on a preliminary examination of our submission we conclude that reliable prediction of carcinogenicity is still a far away goal.

year: 2001

title: The Price of Mauri: Exploring the validity of Welfare Economics when seeking to measure Mātauranga Māori

authors: Awatere, Shaun

subjects: welfare economics

abstract: Since the 1980s New Zealand has pursued neo-classical or market-based policies with a particular fervour. Market-based options are seen by resource management decision makers as essential frameworks for efficiently allocating resources, an approach that continues to support the view of the inherent dominance of Western knowledge. This is particularly concerning, given that Māori (the indigenous people of New Zealand), have an important role to play in New Zealand resource management and perceive their own knowledge systems have been marginalised. The primary goal of this thesis is to explore the validity of welfare economics when seeking to measure quantitatively Mātauranga Māori or Māori views of the environment through the contingent valuation method. A contingent valuation study is carried out using three separate samples drawn from the general Māori population in Auckland city, a hāpu/sub-tribe indigenous to the Auckland isthmus, and drivers of motor vehicles in Auckland city. Data collection modes include a postal survey and face-to-face interviews. This thesis challenges the validity of political-legal ethnicity constructs to measure Mātauranga Māori. The search for a central tendency will lead to biased, misleading and inaccurate results. The thesis also challenges the validity of contingent valuation to produce true economic measures and to measure and identify Mātauranga Māori. Despite advances in analytical techniques, economic efficiency measures are always deficient, given the difficulty of capturing and anticipating all impacts and valuing them appropriately. Mātauranga Māori is derived from a Māori epistemology and should be considered or analysed with primary reference to this body of knowledge. Economic analysis is only one important cog in the machinery of resource management policy. Given that an economist's contribution to local and regional resource management is most valuable when focusing on the  economic efficiency  of the proposed resource allocation, it is appropriate that other perspectives such as Mātauranga Māori be considered.

text: The Price of Mauri: Exploring the validity of Welfare Economics when seeking to measure Mātauranga Māori Since the 1980s New Zealand has pursued neo-classical or market-based policies with a particular fervour. Market-based options are seen by resource management decision makers as essential frameworks for efficiently allocating resources, an approach that continues to support the view of the inherent dominance of Western knowledge. This is particularly concerning, given that Māori (the indigenous people of New Zealand), have an important role to play in New Zealand resource management and perceive their own knowledge systems have been marginalised. The primary goal of this thesis is to explore the validity of welfare economics when seeking to measure quantitatively Mātauranga Māori or Māori views of the environment through the contingent valuation method. A contingent valuation study is carried out using three separate samples drawn from the general Māori population in Auckland city, a hāpu/sub-tribe indigenous to the Auckland isthmus, and drivers of motor vehicles in Auckland city. Data collection modes include a postal survey and face-to-face interviews. This thesis challenges the validity of political-legal ethnicity constructs to measure Mātauranga Māori. The search for a central tendency will lead to biased, misleading and inaccurate results. The thesis also challenges the validity of contingent valuation to produce true economic measures and to measure and identify Mātauranga Māori. Despite advances in analytical techniques, economic efficiency measures are always deficient, given the difficulty of capturing and anticipating all impacts and valuing them appropriately. Mātauranga Māori is derived from a Māori epistemology and should be considered or analysed with primary reference to this body of knowledge. Economic analysis is only one important cog in the machinery of resource management policy. Given that an economist's contribution to local and regional resource management is most valuable when focusing on the  economic efficiency  of the proposed resource allocation, it is appropriate that other perspectives such as Mātauranga Māori be considered.

year: 2008