This thesis aimed to evaluate the contribution of several pneumococcal two-component systems to virulence by analysis of null mutants in a murine model of infection. In addition, a putative zinc metalloprotease, ZmpB, located immediately downstream of one of the TCS, was analysed for its role in virulence. Data indicated that one of the systems studied, TCS08, does not contribute significantly to virulence in serotype 2 pneumococcus, but may have a slightly more important role in a serotype 3 background. A second two-component system, TCS09, was found to be essential for virulence in a serotype 2. Despite the completely avirulent phenotype of the mutant, no difference in expression of many of the previously identified pneumococcal virulence-associated genes was detected in the mutant compared to its isogenic parental strain. Microarray analysis indicated that in serotype 2, TCS09 may be involved in nutrient perception in nutrient perception. TCS09 was found to be required for full virulence in a serotype 3 strain. In this strain, mutants appeared to be impaired in their ability to disseminate from the lungs to the blood in a pneumonia model of infection, but were not attenuated in virulence following direct inoculation into the systemic circulation. These data provide evidence that virulence determinants can behave differently based on the genetic background of the parental strain. ZmpB was found to contribute significantly to pneumococcal virulence in a serotype 3 strain. Further analysis of the contribution of this protein to infection found that ZmpB appears to have a role in promoting inflammation. Thus this work has identified ZmpB as being a novel pneumococcal virulence factor. The role of this protein in inflammation is being investigated further. This thesis has thus identified several genes important in the virulence of S. pneumoniae and work is currently ongoing to assess the potential of these genes as future vaccine or drug candidates. Data presented within this work also provides evidence that virulence determinants can behave differently based on the genetic background of the parent bacterial strain. This important observation could have significant implication for the future characterisation of pneumococcal virulence factors and may apply to other bacterial pathogens.

No description available.

This thesis describes three different strategies employed with the aim of optimising targeted antibodies for the treatment of metastatic solid tumours. Whilst the search for improved predictors of response to anti-EGFR antibodies continues, paired primary and metastatic archived tissue from 32 patients with metastatic colorectal cancer was explored for the immunohistochemical expression of EGFR, pEGFR and pMAPK and activating mutations in KRAS, BRAF and PI3KCA. The resulting discordance between expression of pEGFR and pMAPK between primary and metastatic tissue CRC suggests they are unlikely to be useful biomarkers for response unless metastatic tissue is also analysed. Confirmation that mutations in KRAS, BRAF and PI3KCA are concordant in primary and metastatic tissue supports the analysis of archived primary tissue alone for mutation screening. PI3KCA mutations were shown to be present in patients with both wild-type and mutant KRAS, which provides both an additional method for resistance in wild type tumours and a mechanism for high resistance in those with mutant primary tumours, suggesting screening patients for all 3 mutations should be encouraged for future trials of anti-EGFR antibodies. The Phase I biodistribution study of Ley targeting immunoconjugate in advanced epithelial cancers, primarily explored the biodistribution and pharmacokinetics of the immunoconjugate CMD-193 (a humanised anti-Ley antibody conjugated with calicheamicin) in 9 patients with advanced Ley expressing solid tumours. Cycle one was trace labelled with 111In for biodistribution assessment, and subsequent cycles were administered every 3 weeks, to a maximum of 6 cycles, depending on toxicity and response. Tumour targeting was assessed using gamma camera imaging and single photon emission computerised tomography (SPECT), and PK analysis was based on gamma counting of 111In-CMD-193. There were 2 dose cohorts (1.0mg/m2 and 2.6mg/m2), and patients with Ley positive, measurable, advanced and treatment refractory malignancies, were eligible. Nine patients (6 in dose cohort 1, 3 in cohort 2) were enrolled (and received 1-6 cycles of treatment) before the study was terminated. Biodistribution imaging demonstrated initial blood pooling, followed by markedly increased hepatic uptake by day 2 (persisting to day 8), and fast blood clearance. This pattern was seen for all patients and dose levels. There was no significant uptake in tumour visualised in any patient. The overall T 1/2 β of 111In-CMD-193 was 102.88 ± 35.67 hours, with no statistically significant difference between the 2 dose levels. One patient had a partial metabolic response on 18F-FDG PET after 4 cycles, but no radiologic responses were observed. Myelosuppression and effects on liver function were the most significant toxicities, but no severe or unexpected toxicities were observed. The result of this trial highlight the importance of biodistribution and pharmacodynamic assessment in early phase studies of new biologics to assist in clinical development. The Phase I trial of oral capecitabine combined with 131I-huA33 in patients with metastatic colorectal cancer built on the previous development of the humanised antibody huA33 which targets the A33 antigen, known to be expressed in >95% of human colon cancers. This study used radiolabelled huA33 in combination with capecitabine chemotherapy to target chemoradiation to metastatic colorectal cancer, with safety and tolerability being the primary objective. Pharmacokinetics, biodistribution, immunogenicity, and tumour response were also assessed. Eligibility included measurable metastatic colorectal cancer, adequate hematological and biochemical function, and informed consent. An outpatient scout 131I-huA33 dose was followed by a single therapy infusion one week later, when capecitabine was commenced. Dose escalation occurred over 5 dose levels. Patients were evaluated weekly, with tumor response assessment at the end of the 12-week trial. Tumour targeting was assessed using gamma camera and single photon emission computerised tomography (SPECT) imaging. Nineteen patients were enrolled, and although the dose escalation protocol required an amendment following 2 dose-limiting toxicities in the second cohort, subsequent cohorts demonstrated good tolerability. Biodistribution analysis demonstrated excellent tumour targeting of the known tumour sites, expected transient bowel uptake, but no other normal tissue uptake. 131I-huA33 therefore achieves specific targeting of radiotherapy to sites of metastasis and can be safely combined with chemotherapy, providing a promising opportunity to deliver chemoradiation specifically to metastatic disease in colorectal cancer patients.

Background: Malnutrition and the cachexia syndrome are common in people with cancer. A combination of reduced nutritional intake and abnormal metabolism can lead to physical and psychological disturbances which may impair quality of life and reduce survival. Improved patient outcomes are more likely if treatments and nutritional support can be initiated before significant weight loss has occurred. Methods: A three phase, mixed methods study was undertaken. The primary aim was to gain a greater understanding of the complex factors that have an effect on, and can predict, weight loss in people with cancer. Phases I and II involved the psychometric testing of the Cancer Appetite and Symptom Questionnaire (CASQ). The instrument was tested for reliability among patients receiving radiotherapy (n=34). Predictive validity of the CASQ, using ROC curve analysis, was determined in patients with lung or upper GI cancer (n=185). Total CASQ scores (possible range, 0 to 48) were assessed at baseline, together with percentage weight change after 3 months. An exploratory qualitative study, following the principles of grounded theory, was conducted to explore the causes and influencing factors on weight change. Results: When tested for reliability, the intra-class correlation coefficient of the CASQ was 0.80 (95% CI 0.68 to 0.92) and the difference between total CASQ scores at the two time points was -0.20 (95% CI -1.21 to 0.80). The optimum cut point of the total CASQ score to predict >5% weight loss was 31/32 (C statistic = 0.64; sensitivity 65%, specificity 62%, PPV 33%, NPV 86%), and to predict >10% weight loss was 29/30 (C statistic = 0.75; sensitivity 71%, specificity 66%, PPV 19%, NPV 95%). Exploratory modelling using multiple linear regression techniques suggested that BMI, MUST score, age and the CASQ items of enjoyment of food and pain, were most predictive of weight loss. Nine patients with lung or upper GI cancer and three carers participated in semi-structured interviews. Analysis of the data confirmed the vulnerability of this patient group in terms of symptom burden and nutritional risk. From the findings, a conceptual model that explains the influences on weight change in people with cancer was proposed. Conclusions: Patients with lung and upper GI cancer are at high risk of malnutrition. Psychometric testing of the CASQ suggests that the instrument can predict weight loss in this patient group. Due to the low PPV, further refinements are needed before the instrument is able to be used in clinical practice. A conceptual model which explains the complex process of influences on weight change in people with cancer can improve knowledge and understanding, ultimately informing healthcare practice.

*Please note: The abstract, acknowledgments, list of publications, table of contents and abbreviations in this PDF file appear in a different order to that of the print version of this thesis. Cancer patients, treated with systemic adjuvant chemotherapy, have described experiencing persistent deteriorations in cognition. The nature of these effects is unclear, and although a wide range of theories have been advanced, there is currently no treatment. This thesis uses an animal model to investigate the effects of a commonly prescribed chemotherapeutic agent, 5-fluorouracil (5-FU). The cognitive effects of 5-FU were examined using two behavioural tests, the object location recognition test (OLR) and the conditioned emotional response test (CER) both of which require input from the hippocampus, a brain region associated with memory. Memory consolidation by the hippocampus requires the continual production of new neurons (adult neurogenesis) from progenitor cells in the sub granular zone (SGZ) of the dentate gyrus. As an anti mitotic agent, 5-FU could be reducing the cell proliferation required for neurogenesis and this could be a cause of the cognitive deterioration. This hypothesis was tested by quantifying the numbers of proliferating cells (Ki67+) in the SGZ in sections together with the levels of doublecortin (DCX), a neurofilament expressed in developing neurons and brain- derived neurotrophic factor (BDNF), a factor required for new neuron survival and synaptic plasticity, by Western blotting. After developing the methodology (chapter 2); adult male Lister Hooded rats were given five i.v injections of 5-FU (25mg/kg) over a two week period and their behaviour and cellular aspects of the hippocampus compared with saline injected controls (chapter 3). 5-FU treated animals showed significant impairments in their performance of both the OLR and CER behavioural tests. Animals were sacrificed after the behavioural tests were performed and analysis showed they had significantly reduced numbers of dividing cells in the SGZ and non significant reductions in the levels of BDNF and DCX within the hippocampus. These results demonstrate that 5-FU treatment can produce cognitive impairments in this animal model which are similar in nature to those described by patients after chemotherapy. These behavioural changes are correlated with a reduction in the cell proliferation required for hippocampal neurogenesis providing support for the hypothesis that chemotherapy drugs are affecting this aspect of hippocampal function. In order to develop a treatment for the cognitive effects of chemotherapy the antidepressant fluoxetine was co-administered with 5-FU (chapter 4). This approach was based on recent evidence that fluoxetine can increase neurogenesis and protect neurons after damage. As with the experiment described above, performance in the CER test was impaired by five injections of 5-FU (25 mg/kg) as compared with saline treated controls. Similarly, animals treated with six injections of 5-FU (20mg/kg) were unable to discriminate between objects in novel and familiar locations in the OLR task. However co-administration of fluoxetine in drinking water (10mg/kg/day) for three weeks, starting a week before 5-FU treatment, prevented the impaired performance of this task found in the 5-FU only group. 5-FU chemotherapy caused a significant reduction in the number of proliferating cells in the SGZ compared to controls but this reduction was eliminated in the group co administered with fluoxetine. Fluoxetine on its own had no effect on proliferating cell number or behaviour. Moreover hippocampal BDNF or DCX protein levels in the co-treated group (5-FU+fluoxetine) were significantly increased compared to the 5-FU only treated group. These findings suggest that while 5-FU can negatively affect cell proliferation and hippocampal dependent memory, these deficits can be reversed by co- administration of fluoxetine. To understand the long term effects of chemotherapy, the cellular effects of 5-FU treatment were quantified one day, 2 and 6 weeks after the end of two weeks of 5-FU (20mg/kg) treatment (chapter 5). The results showed that 2 weeks of 5-FU treatment did not significantly reduce cell proliferation in the SGZ when quantified one day after the end of treatment. However proliferating cell numbers were significantly reduced compared to controls two and six weeks after the end of treatment. This suggests that 5-FU has a delayed effect on cell proliferation with its maximum effect two weeks after the end of treatment. Cell survival was quantified by BrdU labelling cells immediately prior to 5-FU treatment, and quantifying the numbers of BrdU positive cells at the different time points. BrdU+ cell numbers were significantly reduced at the end of treatment and continued to decline at 2 weeks but stabilised by 6 weeks. These results demonstrate that 5-FU has prolonged effects on neurogenesis after the end of chemotherapy treatment. The effects of 5-FU on cognition and neurogenesis are discussed and correlated with chemotherapy treated patient reports of continued cognitive impairment for months or years after completion of chemotherapy treatment.

Rationale: CMF (cyclophosphamide: CP; methotrexate: MTX; 5-fluorouracil: 5-FU) is a chemotherapy combination associated with the cognitive impairments which many cancer patients experience after treatment. A reduction in hippocampal neurogenesis is a known means by which cytotoxic drugs alter cognition and is the mechanism investigated in the present study. There is currently no way of treating or preventing the cognitive deficits produced by chemotherapy and a simple pharmacological approach to achieving this could potentially have significant benefits for patients. Objectives: The studies in the present thesis use an animal model to investigate the effects of the individual agents in the CMF combination on spatial working memory and the proliferation and survival of neural precursors involved in hippocampal neurogenesis. It was also investigated whether the cognitive impairment produced by chemotherapy could be reversed or prevented by the antidepressant fluoxetine. Methods: In 4 separate experiments, adult male Lister-hooded rats were chronically administered with CP (30mg/kg, 4 or 7 i.v. doses), MTX (75mg/kg, 2 i.v. doses) or 5-FU (25mg/kg, 5 i.p. doses). Some rats were co-administered with fluoxetine (10mg/kg/day, in drinking water) for different time periods. Spatial memory was tested using the novel location recognition (NLR) task and the spontaneous alternation in the T-maze memory tasks. Proliferation and survival of hippocampal cells was quantified using immunohistochemistry and the levels of doublecortin (DCX) and brain-derived neurotrophic factor (BDNF) were quantified in the hippocampus and frontal cortex using Western blotting. Neural stem cells (NSC) were also isolated from the adult mouse hippocampus, to examine the direct effects of 5-FU, fluoxetine and its active metabolite, norfluoxetine (0.01-100µM) in vitro. Results: Rats treated with 5-FU and MTX showed impairment in the NLR task but not the spontaneous alternation in the T-maze task. They also exhibited a reduction in cell proliferation (Ki67-positive cells) and survival (BrdU-positive cells) in the dentate gyrus, compared to saline treated controls, but no difference was seen in the levels of DCX of BDNF. The induced cognitive and cellular impairments were not seen when fluoxetine was co-administered with the chemotherapy. The impairments caused by 5-FU were counteracted when fluoxetine was co-administered before and during 5-FU treatment but not when it was only administered after treatment. CP did not impair performance in the NLR task or hippocampal cell proliferation; however it significantly reduced cell survival. 5-FU, fluoxetine and norfluoxetine all decreased cell viability in vitro. Conclusions: These results demonstrate that MTX and 5-FU have more pronounced effects on spatial memory and hippocampal cell proliferation than CP in the CMF combination. Furthermore these impairments can be reversed by fluoxetine in a mechanism of prevention but not recovery. Although further work is required, it would be beneficial to establish an in vitro model of chemotherapy-induced cognitive impairment to justify this conclusion and to investigate the potential benefits of fluoxetine in cancer patients.

Cachexia is common in patients with incurable cancer, particularly of the lung and upper-gastrointestinal tract, and impacts adversely on treatment options, morbidity, quality of life and survival. Current management of cancer cachexia is inadequate and progress is required. This thesis explores the use of exercise as a proactive supportive therapy with a focus on maintaining physical function. The first piece of work was a systematic review of the use of therapeutic exercise in patients with or cured of cancer. Across 65 exercise studies, the median [IQR] rates of uptake, adherence and completion were 63 [33–80]%, 84 [72–93]% and 87 [80–96]% respectively, with no characteristic influencing the proportion of patients taking up or completing a programme. The main reasons reported for refusal were lack of interest or the impracticality of the programme and for withdrawal were medical complication or deterioration. Overall, only about half of patients offered an exercise programme completed one. This review highlighted a need to modify existing programmes or explore novel alternatives if exercise is to be acceptable to the majority of patients. The second study explored exercise preferences in patients with incurable cancer. A questionnaire was used to determine patients’ perceived capability and preparedness to undertake six different exercise programmes, each illustrated by video clips and accompanying text, and preferences for the delivery of the most preferred programme. All 200 patients considered themselves physically capable of undertaking an exercise programme and two-thirds were prepared to undertake one at that moment in time. The most preferred type of exercise was neuromuscular electrical stimulation (NMES) 36 [35−44]%, followed by walking 22 [16−30]% and resistance training 19 [13−26]% and the majority preferred to undertake exercise at home, alone and unsupervised. This survey suggested that it is realistic to offer therapeutic exercise programmes to patients with incurable cancer and provided rationale to explore NMES in this group. The third study was a randomised controlled pilot study of NMES in patients with non-small cell lung cancer. Sixteen patients were randomised to a control group, which received usual care, or NMES group, which received daily stimulation to the quadriceps for up to 30min (frequency 50Hz, on phase 11−25%) for four weeks. All patients found the NMES device acceptable and median (range) adherence to the recommended programme was 80% (69-100). In the NMES group, quadriceps muscle strength and free-living physical activity improved by a mean of 7.4 Newton metres (22%) and 136 steps (11%) respectively, whilst exercise endurance deteriorated by a mean of 20 metres (4%). This compared favourably with the control group however none of the differences were statistically significant. These findings suggested NMES was an acceptable type of exercise and that further study is warranted in patients with lung cancer. The final piece of work was a feasibility study into the use of a lightweight ActivPAL™ monitor to measure physical activity level. The aims were to determine if this form of assessment is acceptable to patients, the optimal period of monitoring and to explore the added value of the monitor's energy expenditure (EE) estimate over a simple step count. Sixty patients with lung or upper-gastrointestinal cancer wore a monitor for one week. All but one found the monitor acceptable and mean [95% CI] adherence was 98 [94−100] %. Mean daily step count and EE values measured over 2 and 4 days were significantly higher than those from 6 days (p<0.01). Step count was strongly related to stepping EE and non-stepping EE. The ActivPAL™ monitor was shown to be an acceptable method of assessing physical activity level. A mean daily step count obtained over 6 days was recommended for use in future cachexia studies. Collectively, this work supports the use of therapeutic exercise and highlights a particular role for novel approaches, e.g. NMES, which may be more acceptable to patients. Findings can be used to guide future research which ultimately will determine if therapeutic exercise can help patients with cancer to maintain their level of physical activity and independence for as long as possible.

Modulation of DNA base excision repair (BER) has the potential to enhance response to chemotherapy and improve outcomes in tumours such as melanoma, glioma and pancreatic cancer. APE1, a critical protein in BER that processes potentially cytotoxic abasic sites (AP sites), is a promising new target in cancer. In the current study, my aim was to develop small molecule inhibitors of APE1 for cancer therapy. An industry-standard high throughput virtual screening strategy was adopted. The SYBYL8.0 (Tripos, St Louis, MO, USA) molecular modelling software suite was used to build inhibitor templates. Similarity searching strategies were then applied using ROCS 2.3 (Open Eye Scientific, Santa Fe, NM, USA) to extract pharmacophorically related subsets of compounds from a chemically diverse database of 2.6 million compounds. The compounds in these subsets were subjected to docking against the active site of the APE1 model, using the genetic algorithm-based programme GOLD2.7 (CCDC, Cambridge, UK). Predicted ligand poses were ranked on the basis of several scoring functions. The top virtual hits with promising pharmaceutical properties underwent detailed in vitro analyses using fluorescence-based APE1 AP-site cleavage assays and counter screened using endonuclease IV AP-site cleavage assays, fluorescence quenching assays and Whole cell extract AP-site cleavage assays. Biochemical APE1 inhibitors were then subjected to detailed cytotoxicity analyses. Several specific APE1 inhibitors were isolated by this approach. The IC50 for APE1 inhibition ranged between 50 nM and 25 µM. I also demonstrated that APE1 inhibitors lead to accumulation of AP sites in genomic DNA and potentiated the cytotoxicity of alkylating agents in melanoma, glioma and pancreatic cancer cell lines. I have also shown that APE1 inhibitors induce delay in cell cycle progression and caused delay in cancer cell growth. I also demonstrated that APE1 knockdown by shRNA results in decrease cancer cell growth and enhanced cell killing by alkylating agent in Glioma cell line. This PhD project provides evidence that APE1 is an emerging drug target and could have therapeutic application in patients with melanoma, glioma and pancreatic cancer.

Despite considerable research on cancer treatments and preventatives, poor outcomes in cancer patients are common. The vital search for effective cancer drugs often begins in the laboratory, where unfortunately the effects of a drug in humans cannot be perfectly modelled. Epidemiology can play a vital role in determining the real world efficacy of a drug currently used for other purposes before clinical trials begin. This thesis therefore used primarily laboratory evidence to identify potential anti-cancer uses for existing common drugs. The drugs and cancers studied were; tricyclic antidepressants and both incidence and survival in a number of cancer types, particularly glioma; aspirin and colorectal cancer survival; and angiotensin converting enzyme (ACE) inhibitors and hepatocellular carcinoma (HCC) incidence. A series of studies using The General Practice Research Database as a data source assessed any potential associations: A case-control study for tricyclic antidepressant use and cancer incidence; cohort studies to examine mortality in colorectal cancer and glioma in relation to tricyclic use, and for colorectal cancer mortality in aspirin users; and a case-control study in relation to ACE inhibitor use and HCC. A strong, cancer type specific, dose and time dependant protective effect was found for the incidence of glioma and colorectal cancer. This led to a further study examining mortality for these cancer types in tricyclic users. While no significant protective effects in all-cause mortality of tricyclic users were found, a larger study could still find such an effect in glioma. For aspirin and colorectal cancer mortality, a small but significant reduction in mortality was observed, though these effects were not entirely consistent throughout the study. There were no significant associations found between ACE inhibitors and HCC. These findings contribute to the knowledge of the anti-cancer effectiveness of these drugs, and may assist in designing future clinical studies.

Consumption of fruits and vegetables has been shown to reduce the risk of various types of cancer. Macro- and micro-nutrients as well as non-nutritive phytochemicals present in fruits and vegetables have been associated with this effect. This study was conducted to investigate the chemopreventive and chemotherapeutic potential of two types of fruits which are endemic to Borneo Island: Mangifera pajang (bambangan) and Artocarpus odoratissimus (tarap). The first part of the project was to study the antioxidant potential of the crude extracts of the plants in vitro. The fruits were first separated into flesh, kernel and peel for M. pajang and flesh and seed for A. odoratissmus. DPPH (2,2- diphenyl-I-picrylhydrazyl) free radical scavenging and FRAP (ferric reducing / antioxidant power) assay were employed for the antioxidant study. The result showed that the kernel of M. pajang extract displayed strongest antioxidant activity as assessed using both assays, followed by M. pajang peel, A. odoratissmus seed, M. pajang flesh and A. odoratissmus flesh. The presence of selected phytochemicals in the plant extracts was determined in the next chapter. Polyphenols have been identified as major phytochemicals in the plant extracts, and in M. pajang kernel extract represents about 10% of its total weight. Gallic acid, coumaric acid, sinapic acid, caffeic acid, ferulic acid, chlorogenic acid, naringin, hesperidin, rutin, luteolin and diosmin have been identified as the key polyphenol phytochemicals present in the kernel of M. pajang which might be responsible for the superior antioxidant properties as compared to other extracts. Concern that the results for the chemical antioxidant assay do not necessarily reflect cellular activity led to the third part of the project; assessment of the cytoprotective activity of the crude extracts against oxidative stress induced by tert-butyl hydroperoxide (t-BHP). Only M. pajang kernel extract as well as the positive control (quercetin) displayed cytoprotective activity against this toxicant. It seems that non-cell based antioxidant assay does not necessarily reflect the activity in cell-based antioxidant assay. This is shown by lack of cytoprotective activity of both M. pajang peel and A. odoratissimus extracts despite their considerably high antioxidant activity in DPPH free radical scavenging and FRAP assay. In order to study which proteins might be involved in the cytoprotection mechanism, western blotting method was employed to determine the expression of various Cytoprotective proteins [i.e. quinone reductase (NQO I), glutathione peroxidase (OR), methionine sulfoxide reductase A (MSRA), heat shock protein 27 (HSP27) and heat shock protein 70 (HSP 70)]. Different cytoprotective mechanisms were observed by the kernel extracts and quercetin. In the present study, NQOI, OR, MSRA, HSP27 and HSP 70 have been shown to be involved in the cytoprotection activity of quercetin while only OR and MSRA were involved in the cytoprotection activity of M. pajang kernel extracts. Other cytoprotective proteins remain to be studied to fully understand the cytoprotection mechanism of both plant extract and quercetin. Some chemopreventive agents have been shown to suppress cancer proliferation, induce apoptosis in cancer cells as well as inhibit angiogenesis and metastasis in pre-clinical and clinical trials. Thus, the last part of the project was to determine the anti-cancer potential of plant extracts in a variety of cancer cell lines (derived from breast, colon, liver and ovarian carcinoma). The results showed that the kernel extract of M. pajang displayed strong anti proliferative activity in breast cancer cell lines (MCF-7 and MDA-MB-231). The kernel extract induced cell cycle arrest in MCF-7 cells at the sub-G1 (apoptosis) phase of the cell cycle in a time-dependent manner. For MDA-MB-231 cells, the kernel extract induced strong G2-M arrest in cell cycle progression at 24 hours, resulting in substantial sub-G1 (apoptosis) arrest after 48 and 72 hours of incubation. Staining with Annexin V -FITC and propidium iodide revealed that this apoptosis occurred early in both cell types, 36 hours for MCF-7 cells and 24 hours for MDA-MB- 231 cells, with 14.0% and 16.5% of the cells respectively undergoing apoptosis at these times. This apoptosis appeared to be dependent on caspases-2 and -3 in MCF-7 cells and on caspases-2, -3 and -9 in MDA-MB-231 cells. As a conclusion, from the two plants (M. pajang and A. odoratissimus) studied, the extract of M. pajang kernel displayed diverse health benefit properties, antioxidant, chemoprevention and chemotherapeutic potential. M. pajang could be fully utilized for pharmaceutical, nutraceutical as well as food products. Further study (i.e. animal and clinical study, isolation of pure compounds, bioavailability study) are required to determine the efficacy in human population.