Research on bioactive compounds in Bacillus thuringiensis
Vishna Devi Nadarajah, Shar Mariam Mohammed, Chan Kok Keong, Kanakeswary Krishnan, Soong Zhao Kang, Yow Jee Seng

Historically it is believed that Bacillus thuringiensis (Bt) is a species characterized by the insecticidal activity of its parasporal inclusions. In natural environments, Bt strains producing non-insecticidal parasporal inclusions are more widely distributed than insecticidal ones. However the biological activity of non-insecticidal Bt parasporal inclusions remains relatively unexplored and could potentially be a rich source of therapeutic agents. Indeed, studies have shown that parasporal inclusions from some Bt strains have active and potent antibacterial and anticancer activity. The gene encoding the anti cancer protein (parasporin) has been cloned from Bt isolate designated 84-HS-1-11 and found to encode a polypeptide of 723 amino acid residues with a predicted molecular weight of 81kDa. The sequence of parasporin contains the five conserved blocks commonly found in B. thuringiensis Cry proteins (most Cry proteins are insecticidal, and is expressed as parasporal inclusions in Bt strains); however, only very low homologies (<25%) between parasporin and the existing classes of Cry and Cyt proteins was found. Parasporin exhibited cytocidal activity only when degraded by proteases into smaller molecules of 40 to 60 kDa. Notably, activated parasporin showed strong cytocidal activity towards human leukaemia T cells (MOLT-4) and human uterus cervix cancer cells (HeLa) but were inactive towards normal T cells. Recently, several other Bt isolates (reported by research groups in Japan), were found to produce non-insecticidal and non-haemolytic parasporal inclusions.

Our research group at IMU has identified a Malaysian Bt strain designated Bt 18 to be specifically cytotoxic against CEM-SS cell lines. Bt 18 produces parasporal inclusions, which after trypsin activation is non-haemolytic to human or rat erythrocytes, and is non-cytotoxic to uterus, breast and colorectal cancer cell lines. Importantly, Bt 18 is non toxic to purified T-lymphocytes isolated from healthy individuals, indicating its potential as a therapeutic agent for leukaemia cells. N-terminal sequencing of the trypsin activated Bt 18 inclusion, showed that this protein has a low sequence similarity to a mosquitocidal Bt protein, found in Bt sub species jegathesaniensis, also isolated from the Malaysian soil. Efforts are now underway to identify the gene responsible for the cytotoxic protein expressed in Bt 18.

Studies by our group on the effect of Malaysian mosquitocidal Bt strains on erythrocytes suggests that Bt toxins are lectins which recognize specific plasma membrane glycoconjugate receptors with a terminal residue of either D-mannose (Man), N-acetyl-D-galactosamine (GalNAc), N-acetyl-D-glucosamine (GlcNAc), or even a combination of these monosaccharides. However, as Bt 18 is non-haemolytic against human erythrocytes, the lectin property cannot be determined using the haemolytic inhibition test. Hence another focus area in our group is the use of immunoassays to study the effect of sugars and localisation of Bt 18 on leukaemia cells.

As the long term usage of insecticidal Bt strains can lead to the development of insect resistance, strains that display novel vegetative insecticidal polypeptides will be useful. Unlike the insecticidal Bt d-endotoxins, vegetative insecticidal protein, (Vip), is an 88-kDa protein, which is secreted during the vegetative stage by Bt and it displays high toxicity against a range of insects. Our group, in collaboration with senior researchers from the Entomology Department of IMR and the Medical Faculty of UPM, has recently begun work to isolate bioactive compounds, expressed during the vegetative growth phase of the Malaysian Bt strains, and these compounds will be screened for both mosquitocidal and anti cancer activity.


Development of immunoassays for the detection of Candida antigens in systemic candidiasis
Wong SF, Mak JW, Peter Pook CK, Ng KP, Seow HF, Ngah Zasmy

The clinical spectrum of candidiasis can range from mild superficial infection to life threatening systemic infection with multi-organ involvement. As there are no pathognomonic signs or symptoms of systemic candidiasis, the diagnosis of invasive candidiasis remains a laboratory and clinical challenge. Current serological assays suffer from inadequate sensitivity and specificity or the need for multiple sampling of clinical specimens. Thus, this collaborative study between researchers from IMU, Universiti Putra Malaysia, and University of Malaya, seeks to develop diagnostic assays for systemic candidiasis, identify Candida virulence factors, and study the associated pathological changes using immunohistochemistry.

Monoclonal and polyclonal antibodies were produced against C. albicans and C. parapsilosis. A total number of forty and fifty clones producing monoclonal antibodies against C. albicans and C. parapsilosis respectively were obtained. Two clones from non-heat shocked C. albicans crude extract (IgM) and one clone from heat shocked C. albicans crude extract (IgG3) were propagated in pristane-primed mice to generate ascites fluid. Two clones (IgG1 and IgG2a) from heat shocked C. parapsilosis crude extract were propagated in pristine-primed mice. The monoclonal antibodies were purified from culture medium and ascites fluid, characterized and used in the immunoperoxidase technique. The monoclonal antibodies were able to bind on the surface of the yeast cells and germ tubes.

We induced systemic candidiasis in Balb/c mice through injection of cultured C. albicans harvested from YPD medium supplemented with foetal bovine serum. Histopathological examination showed cystic lesions with large clumps of fungus mainly hyphae and some yeast in the kidneys. Clusters of yeast cells were also detected in lungs, brains and spleen. The relationships between histopathological changes, infective dose, antibody response, and circulating antigens are being investigated.

Two fungal isolates obtained from blood culture of experimentally infected mice were confirmed to be the same species using DNA sequencing and sequence alignment with ATCC and other strains. Western blot results showed that the monoclonal antibodies against non-heat shocked and heat shocked C. albicans were reactive against polypeptides with MW of 20 and 33 kDa respectively. Similar studies with monoclonal antibodies raised against heat shocked C. parapsilosis antigens showed that they were reactive to polypeptides with MW at 30 and 59 kDa.


Mechanism of action of antivirals against nasopharyngeal carcinoma at the molecular level
Chu Wan Loy, Shar Mariam, Mak Joon Wah, Rakesh Naidiu, Pauline Balraj, Patricia Lim Kim Chooi, Phang Siew Moi, Kok Yih Yih, Lai Pey Jiun, Tan Boon Keat

Nasopharyngeal carcinoma (NPC), a type of cancer arising in the epithelial cells of the nasopharynx, has a high incidence in Malaysia, especially amongst Chinese of Southern China origin. It is known that NPC is associated with the Epstein-Barr virus. The major objectives of this project are to screen indigenous microalgae of Malaysia for bioactivity against NPC cell lines and to unravel the mechanisms of action of the bioactive compounds. The project is funded by the Ministry of Science, Technology and Innovation (MOSTI) under the Intensified Research Priority Area (IRPA) programme, and is in collaboration with the University of Malaya and Institute of Medical Research.

The solvent extracts from 19 microalgae have been screened for their cytotoxic effect against NPC cell lines. Results from the screening showed that methanol extracts from two microalgae, Ankistrodesmus convolutus 101 and Synechococcus elongatus UMACC 105 showed antineoplastic activity against NPC cell lines. Fractionation of the crude extracts using chromatographic techniques and further testing with the fractions are being carried out. In addition, methods for the testing of antiviral mechanisms based on the expression of proteins by Epstein-Barr virus have been developed.


Molecular and immunological characterisation of pathogenic and non-pathogenic Acanthamoeba species in Malaysia
Chan Li Li, Mak JW, Init Ithoi

Acanthamoeba species are free-living amoebae that are ubiquitously distributed in nature. Although many Acanthamoeba spp. and strains are not harmful to human, certain isolates, especially the species A. castellanii, A. culbertsoni and A. polyphaga, can cause serious human diseases. These species cause two distinct infections in humans. Acanthamoeba keratitis is a potentially sight-threatening ocular infection that affects both the immunocompetent and immunocompromised individuals, and is often associated with non-hygienic contact lens usage. Granulomatous amoebic encephalitis is a chronic brain disease in immunocompromised patients. As these organisms can also be used as indicators of environmental air or water quality, it is important to differentiate pathogenic from non-pathogenic species and strains. The objectives of this study are to differentiate these organisms based on morphological, immunological, molecular, in vitro culture characteristics, and pathogenicity studies.

Both clinical and environment isolates of Acanthamoeba will be used in the study. The clinical isolates were isolated from patients with keratitis, and were provided by Dr. Kamel from University Kebangsaan Malaysia. The environmental isolates were collected from air condition units of the central cooler using sterile cotton swap. Both types of isolates were cultured on non-nutrient agar seeded with Eschericha coli lawn. Morphological studies were carried out on Trichrome and Iron Haematoxylin stained organisms. Since environmental isolates of Acanthamoeba were in mixed spp., methods for obtaining single cyst and development of single culture line of the protozoan has been established. Currently methods are being developed to axenise the clinical and environmental Acanthamoeba isolates.

Identification of Acanthamoeba isolates at the generic level using published PCR primers has been developed. Direct sequencing on PCR products amplified from one of the single culture line of Acanthamoeba isolate was performed. This was to confirm the reliability of the published PCR primers on detecting Acanthamoeba spp at genus level. Two sets of published PCR primers capable of distinguishing pathogenic and non-pathogenic Acanthamoeba were tested. However, the DNA profile obtained was not similar to those reported.


Reduced tumour growth and metastases following vaccination with tumour-lysate-pulsed dendritic cells (DC)
Ammu Radhakrishnan, Geok Choo Sim, Soon Keng Cheong

The concept that the immune system recognizes and controls cancer has been around for over a century and cancer immunity has continued to be vigorously debated and experimentally tested. The idea that the immune system detects altered or transformed cells, mainly malignant cells underpins much of the current excitement surrounding the development of new immunotherapeutic approaches for the treatment of cancer. Successful tumour growth is a consequence of a balance between the kinetics of host immune response and tumour growth. The main goal of active immunotherapy is to develop an effective anti-tumour immune response in vivo. The initiation of host immune response towards tumour is dependent on the activation of T-lymphocytes.

Dendritic cells (DC) are bone marrow-derived leukocytes that play a key role in activation of T-lymphocytes. The dendritic cells are presumed to die in the lymph nodes after completing their antigen presentation function as they do not appear to exit the lymph node.

The aim of this study was to study the effect on tumour growth and metastasis following vaccinating experimental mice with DC-pulsed with specific tumour lysate from a murine breast cancer cell line and challenged later with the same tumour cells.

We generated DC from cultured murine bone marrow cells. These murine bone marrow-derived DC were pulsed with tumour lysate from 4T1 tumour cells and injected subcutaneously in the left flank of experimental mice for three times prior tumour inoculation. Control mice were not injected with tumour-lysate pulsed. Both control and experimental mice were challenged with 1x104 cells of 4T1 tumour cells by injecting the cells at the mammary pad of six-week old female BALB/c mice. The size of the tumours developed were assessed in a coded, blinded fashion and recorded as tumour volume (mm3) by measuring the largest perpendicular diameters and calculated using established formulae. The data represent the average tumour volume ? SD in each group of mice.

Experimental mice vaccinated with tumour-lysate pulsed DC demonstrated delay in tumour growth as compared to non-treated control mice. In addition, the tumour volume of DC-vaccinated mice was much smaller than the control mice. We also observed that the lung of control mouse with no DC treatment displayed pulmonary metastasis while the lungs taken from DC treated-mouse treated with 4T1 tumour lysate pulsed-DC had a smooth surface and there appeared to be devoid of tumour colonies.

Our study shows that vaccinating mice with DC pulsed with tumour lysate can confer some degree of protection in these mice in terms of tumour onset, tumour size. Although our finding is similar to that reported by researchers studying this effect in other tumour models, there are some advantages and disadvantages on the use of tumour lysate as a source of antigen. The major attraction of this method is that it allows DC to pick up a wide array of tumour peptides, which can result in eliciting immune response simultaneously on many tumour antigens expressed by tumour cells. This approach significantly enhances the anti-tumour response and to some extent prevents tumour escape from host immune system. In addition, the use of whole tumour cells that represent the entire repertoire of tumour Ag could circumvent the need to define specific immunogenic peptides. However, the major drawback of this method is on the concern that some self-peptides might be loaded on MHC molecules at the same time as tumour antigens. This increases the risk of inducing autoimmune or immune response induced self-tolerance to self-antigen and tumour-ag, which subsequently can reduce the efficacy of tumour lysate-pulsed DC in eliciting tumour-specific immune response. Despite these concerns, it is encouraging to note that no adverse effect was observed in our experimental model, similar that reported in some clinical trials using tumour lysate pulsed-DC on cancer patients. These observations suggest that this approach is safe for clinical applications.


Comparing the Allele Frequencies of Single Nucleotide Polymorphisms (SNP) in the Human Interleukin-10 (IL-10) Gene Promoter between Rheumatoid Arthritis (RA) Patients and Normal Subjects in Malaysia
Ammu K. Radhakrishnan, Hee Chee Seng, Gun Suk Chyn, Sushela Devi Somnath, Rakesh Naidu, Esha Gupta

Rheumatoid arthritis (RA) is chronic inflammatory disorder caused by autoimmune responses, where the regulation of their immune responses and recognition has been disrupted. Cytokines have been shown to play an important role in the coordination of the inflammatory processes RA patients. Pro-inflammatory cytokines initiate and maintain inflammation while anti-inflammatory cytokines suppress this process once the stimulus for inflammation is eliminated. It is believed that in RA patients, there is a state of imbalance between these cytokines, leaving inflammation unchecked and this in turn results in destruction of host tissues.

The aim of this study was to identify the frequencies these SNPs in RA patients compared to normal healthy volunteers and later to correlate these findings with IL-10 secretion and also clinical findings.

One-hundred and forty nine patients with physician-diagnosed RA and 149 normal subjects were recruited in this study. Patients with all levels of RA severity were recruited from Seremban Hospital outpatient clinics or general wards and all of them fulfilled the 1987 revised criteria of the American College of Rheumatology (ACR). Normal subjects recruited had no rheumatology or other immunological related diseases and no significant medical illness.

DNA was extracted from peripheral blood of RA patients and normal volunteers using commercially available DNA extraction kit (QIAGEN). The Interleukin-10 (IL-10) romoter region from -1120 to -533 flanking site -824 and -597 was amplified by polymerase chain reaction (PCR) using reported primers. PCR products were checked using 1% agarose gel electrophoresis. SNPs were detected by restriction fragment length polymorphism (RFLP).

Statistical analysis was performed using SPSS and MINITAB statistical packages (SPSS version 10.5, Chicago, IL; MINITAB version 14). Genotype and allele frequencies in the study group were compared by the ?2- test. P<0.05 was taken as the cut-off for statistical significance.

There was no significant deviation of genotype frequencies from the Hardy-Weinberg equilibrium noted in either RA patients or control subjects. SNP at position -824 and -597 of the IL-10 promoter gene were found to be in complete linkage disequilibrium, and therefore the genotype distribution was identical. The genotype and allele distribution at positions -1082, -824 and -597 in RA patients did not differed significantly from that observed for the control group (p>0.05). However, allelic distribution among 3 major ethnic groups in both RA patients and controls differed significantly (p<0.005).

Four haplotypes were identified including a rare haplotype -1082G/-824T/-597A (GTA), which is reportedly only found in the Asian populations. Haplotypes were calculated using EHprogram and our results showed no significant different between controls and patients. The IL-10 SNPs frequencies in our population were identical to that reported in Japanese and Chinese cohorts but contradictory to the reports on Caucasian populations.


Molecular studies on disease biomarkers and pathogenesis of cancer: colorectal carcinoma as a model
Lim Vin Nee, Mak Joon Wah, Rakesh Naidu, Rahman Jamal, Isa Mohd Rose, Kevin Moissinac, Yunus Gul, Patricia Lim Kim Chooi, Greg Tan JS

In Malaysia, colorectal cancers (CRC) have been ranked second in males and sixth in females. Genetic alterations leading to abnormality in protein expression are involved in the tumorigenesis of various cancers. The correlation between expression of VEGF, cyclin D1, cyclin E, CD31, MMP-7 and ß-catenin in the different lesions of CRC has not been completely elucidated. The relationship of these biomarkers with ethnic, gender and age groups in the Malaysian population has not been studied. Their role as prognostic biomarkers needs further investigation. This IRPA funded top-down project seeks to determine their expression in different colorectal neoplastic lesions, and to correlate the protein expression with tumour prognostic factors and progression.

A total of 82 CRC carcinoma specimens have been collected for the study. Most of these specimens were from Chinese, followed by Malay and Indian. The proportion of male and female patients is almost the same, with an M: F ratio of 1.05: 1.0. There were 52 (63.4%) patients who were < 65 years old.

Cyclin D1 and cyclin E immunostaining were predominantly nuclear, whereas VEGF is predominantly cytoplasmic. Some specimens show cyclin E reactivity in both nucleus and cytoplasm; however, only those with nuclear staining was considered as positive. 38 (46%), 51(62%) and 58 (70%) of the CRC carcinoma specimens were positive for cyclin D1, cyclin E and VEGF respectively. There was no significant difference in the percentage of positive expression of cyclin D1, cyclin E and VEGF with the histological grade of tumours (P>0.05). However, cyclin E over expression is associated more frequently with poorly differentiated tumours. The analysis of the co-expression between VEGF, cyclin D1 and cyclin E suggest that their expression were independent of each other and may not be related in the CRC expression (P>0.05).

The further evaluation on the immunostaining for CD31, MMP-7 and ß-catenin expression is in progress. The significance of cytoplasmic expression of cyclin E is being studied.


Immunohistochemical localisation and distribution of placental angiotensin II receptor subtypes in normal and pre-ecclamptic pregnancies
Judson JP, Vishna Devi Nadarajah, Yi Chung Bong, Michael Lau Wei Tiong, Nalliah Sivalingam, J RavindranS

Pre-eclampsia (PE) is an abnormal state of pregnancy characterised by hypertension and fluid retention and albuminuria and is associated with a great degree of morbidity and mortality. Its aetiopathology is poorly understood and delivery of the placenta is the only known cure for this condition at the moment.

Recent studies have proven the existence of a local renin - angiotensin mechanism within the placenta. Immunohistochemical studies have proven the presence of Angiotensin receptors (primarily the receptor subtypes AT1 and AT2) in the placenta. However, not many quantitative and qualitative studies have done to quantify these receptors and analyse their characteristics. The figures of the incidence, morbidity and mortality of pregnancy induced hypertension (PIH) in Malaysia are comparable to the global statistics. Physical characteristics, genetic predisposition, parity, diet and age have all been postulated as factors responsible in some way in the causation of the disease. However such clinical data is sparse and the relationship between biomarkers and these factors has not been studied in the Malaysian population.

A total of 38 samples were collected for this study. The tissues were processed for immunohistochemistry and molecular studies. The results were analysed independently as well as correlated against a previous pilot study conducted in the IMU last year by the same team. It was observed that there was a difference in the level of angiotensin II receptors subtypes' expression in normal and PIH placentas. In the normal placenta, the level of AT1 receptor expression was found to be more or less the same as the level of AT2 receptor expression at the ratio of 1.1:1.0.However, it was observed that there was a definite imbalance in the ratio of AT1:AT2 in PIH/PE placenta caused by an increase in the number of AT1 receptors It was observed that the ratio of AT1:AT2 in PIH placenta changed to 1.8:1.0 by using immunohistochemical staining.

The findings of this study may explain the causes of ischaemic placenta in PIH/PE pregnancies. In normal placenta, there is a balance between AT1 receptors and AT2 receptors which suggests that there is a balance between vasoconstrictory and vasodilatory effect on placental spiral arteries as well. This is to ensure sufficient blood flow to placenta. However, in PIH/PE placenta, AT1 receptors outnumber AT2 receptors. There is less AT2 receptors to counteract the vasoconstriction effect of AT1 receptors in PIH placenta. The increase in AT1 receptors in PIH placenta which causes vasoconstriction compared to AT2 may result in a poorly perfused placenta which may explain the increase in blood pressure observed in PIH/PE patients.

Previous studies by others showed evidence of the AT1 receptor as a 60 kDa molecular weight protein by using Western blot analysis. In addition, Western blot analysis by these workers showed a minor band of 45 kDa. SDS-PAGE analysis of the placenta samples used in our investigation showed a 60 kDa band and a 41 kDa band in both PIH/PE and normal samples. However there was a difference in the protein profile between both placental types, as an additional 50 kDa band was observed to be more prominent in the PIH/PE samples. This band may represent a more potent form of AT1 receptor as it is found to be more dominant in PIH/PE mothers. However, this observation needs to be verified by using a more quantitative method and Western Blot analysis.

The findings in this study warrant further investigations such as fine tuning the Western blot method and analyzing at the molecular level the mRNA expression of AT1 and AT2 receptors in PIH and PE patients. It is hoped that in future, information elucidated from these studies, can aid attempts to develop early diagnostic kits for PIH/PE. Perhaps early detection of PIH in pregnancies can help doctors better manage soon to be mothers and prevent complications.


Respiratory medicine research in IMU
Richard Li-Cher Loh

From 2001, early respiratory research in IMU focused on asthma and most work started as descriptive studies elucidating on management issues such as the assessment of patients with acute exacerbation and the state of asthma control in patients followed up in local hospital and community clinics. These early work provided the impetus into studies of health-related quality of life, influence of ethnicity, perception of breathlessness, and more recently, culminated in the completion of nationwide survey of prescribing patterns practiced by various healthcare professionals, a collaborative work with The Malaysian Thoracic Society, a landmark study providing important benchmark data to study gaps in asthma management from the government and private healthcare perspective.

Parallel to this, was the setting up of sputum induction and processing facility in IMU Clinical School in 2001 to explore this modern technique of investigating airway inflammation in asthma and other respiratory diseases. From this, we published work on the refinement of the induced sputum technique, the profiling of airway inflammation in asthmatics of varying severity, and concluded some studies investigating the effect of inhaled bacterial endotoxin on human airways. The latter two represented the tail-end of a collaborative work with The London Chest Hospital and King's College London, United Kingdom. The sputum research also received national attention when awarded The Ranjit Bhagwan Singh Research Grant in 2001 through the Academy of Sciences Malaysia.

From 2002, respiratory research of IMU had expanded to the area of tuberculosis and non-tuberculosis lung infections, most notably in the area of community-acquired pneumonia in patients requiring hospital admission. In one study, we were the first to prospectively test severity assessment index validated from Western countries in an Asian context and showed their lack of relevance. In another in 2003, we studied the impact of SARS in affecting medical students and healthcare workers in local hospital. Presently, several interesting findings based on two large retrospective studies from our local teaching hospital had been made known in international scientific conferences and one on-going prospective study seeking to better define risk factors for multi-resistant organisms in hospitalized patients with pneumonia. Another recently completed study, emanated from a collaborative effort with The Institute of Respiratory Medicine in Kuala Lumpur, presented preliminary data on the differing microbial aetiologies in patients with and without chronic lung disease, providing a rare Asian perspective of this area.

Although not anticipated to play any huge role, research on lung cancer from the aspect of disease presentation and management issues has also been successfully conducted and published. During this time, in addition to the above, the IMU respiratory research had also completed two large international multi-centre clinical trials on asthma, in which IMU centre achieved the highest number of study recruits among the participating centres in Malaysia, and presently conducting another two international trials, one on asthma and the other on chronic obstructive pulmonary disease.

IMU medical students had contributed significantly to the success of respiratory research in IMU. Notwithstanding the fact that the students have to undertake a research project in their Semester 6 & 7, those who opted for respiratory research enjoyed the experience and had the opportunity to present their work in scientific conferences. Since the inception of respiratory medicine research in IMU in 2001, to date, 14 students had attended and presented in international congresses like The European Respiratory Annual Congresses and World Asthma Meeting, 10 had attended and presented in regional congresses like that of The Asia-Pacific Society of Respirology, and 4 presented in national conferences. Not seen in any other local universities, the IMU student affair policy of providing a sizeable financial subsidy for students presenting in scientific meetings is the single most important factor in making it possible for these students to attend these conferences especially those outside Malaysia.

The success of respiratory medicine research in IMU thus far is attributable to several factors. First is the significant level of internal funding provided by IMU Research and Ethics Committee in providing the much needed seed money to kick start and also to maintain the momentum of the many 'hypothesis generating and testing' projects of ours. Second is the strong support and close partnership with The Ministry of Health Malaysia and senior staff in Seremban Hospital in the conduct of many of these research, as most of these studies were clinical in nature. Third is the employment of IMU part-time research nurse in conducting many of these studies. Throughout these years, respiratory medicine research has had two research nurses, one after another, funded by money obtained from the conduct of pharmaceutical firm sponsored clinical trials. They have proven indispensable to the smooth running and focus of our work. Finally, our own focus and discipline that have helped to overcome obstacles and the charting of any particular research direction.

The future of respiratory medicine research in IMU is likely to continue in present direction of both descriptive and applied experimental nature. To realistically push the frontier of respiratory research, we would need to engage full time researchers, both clinical research fellows and bench-based scientists in pursuits of MD and PhD projects, and other post-doctoral aspirations. We would also need to compete for larger grants like those from government national bodies for this purpose. Presently, a work of ?2-adrenoceptor polymorphisms among asthmatics in relation to ethnicity of Malaysia is concluding and will be the subject of a PhD project for an IMU academic staff. Another work on study into development of model for delivery of pharmacological care in Malaysian context for patients with asthma and COPD has recently begun as a PhD project for another IMU academic staff.