Immunohistochemical and Molecular Prognostic/Predictive Markers in Neoplastic Diseases

Traditional morphological examinations are not anymore sufficient for a complete evaluation of tumoral tissue and the use of neoplastic markers is of utmost importance. Neoplastic markers can be classified in: diagnostic, prognostic and predictive markers. Three markers were analyzed. 1) Insulin-like growth factor binding protein 2 (IGFBP2) was immunohistochemically examined in prostatic tissues: 40 radical prostatectomies from hormonally untreated patients with their preoperative biopsies, 10 radical prostatectomies from patients under complete androgen ablation before surgery and 10 simple prostatectomies from patients with bladder outlet obstruction. Results were compared with α-methylacyl-CoA racemase (AMACR). IGFBP2 was expressed in the cytoplasm of untreated adenocarcinomas and, to a lesser extent, in HG-PIN; the expression was markedly lower in patients after complete androgen ablation. AMACR was similarly expressed in both adenocarcinoma and HG-PIN, the level being similar in both lesions; the expression was slightly lower in patients after complete androgen ablation. IGFBP2 may be used a diagnostic marker of prostatic adenocarcinomas. 2) Heparan surface proteoglycan immunohistochemical expression was examined in 150 oral squamous cell carcinomas. Follow up information was available in 93 patients (range: 6-34 months, mean: 19±7). After surgery, chemotherapy was performed in 8 patients and radiotherapy in 61 patients. Multivariate and univariate overall survival analyses showed that high expression of syndecan-1 (SYN-1) was associated with a poor prognosis. In patients treated with radiotherapy, such association was higher. SYN-1 is a prognostic marker in oral squamous cell carcinomas; it may also represent a predictive factor for responsiveness to radiotherapy. 3) EGFR was studied in 33 pulmonary adenocarcinomas with traditional DNA sequencing methods and with two mutation-specific antibodies. Overall, the two antibodies had 61.1% sensitivity and 100% specificity in detecting EGFR mutations. EGFR mutation-specific antibodies may represent a predictive marker to identify patients candidate to tyrosine kinase inhibitors therapy.

Deciphering microRNA code in pain and inflammation: lessons from bladder pain syndrome

MicroRNAs (miRNAs), a novel class of molecules regulating gene expression, have been hailed as modulators of many biological processes and disease states. Recent studies demonstrated an important role of miRNAs in the processes of inflammation and cancer, however, there are little data implicating miRNAs in peripheral pain. Bladder pain syndrome/interstitial cystitis (BPS/IC) is a clinical syndrome of pelvic pain and urinary urgency/frequency in the absence of a specific cause. BPS is a chronic inflammatory condition that might share some of the pathogenetic mechanisms with its common co-morbidities inflammatory bowel disease (IBD), asthma and autoimmune diseases. Using miRNA profiling in BPS and the information about validated miRNA targets, we delineated the signaling pathways activated in this and other inflammatory pain disorders. This review projects the miRNA profiling and functional data originating from the research in bladder cancer and immune-mediated diseases on the BPS-specific miRNAs with the aim to gain new insight into the pathogenesis of this enigmatic disorder, and highlighting the common regulatory mechanisms of pain and inflammation.

Forchheimer's therapeusis of internal diseases.

I. General therapy. Toxicology.--II. Infectious diseases. The intoxications. Constitutional diseases.--III. Digestive system. Respiratory tract. Heart and blood vessels. Blood and ductless glands.--IV. Kidney. Bladder. Male sexual organs. Nervous system. Tropical diseases.--V. Vaccines and serums.

The role of inflammatory mediators in the overactive and bladder pain syndromes

The Overactive Bladder (OAB) and Bladder Pain Syndrome (BPS) are debilitating disorders for which the pathophysiological mechanisms are poorly understood. Injury or dysfunction of the protective urothelial barrier layer, specifically the proteoglycan composition and number, has been proposed as the primary pathological characteristic of BPS. For OAB, the myogenic theory with dysfunction of the muscarinic receptors is the most reiterated hypothesis. For both over activity of the inflammatory response has been posited to play a major role in these diseases. We hypothesise that BPS and OAB are peripheral sensory disorders, with an increase in inflammatory mediators, such as cytokines and chemokines, which are capable of activating, either directly or indirectly, sensory nerve activity causing the disease. The aim of the PhD is to identify potential new therapeutic targets for the treatment of BPS and OAB. We used medium throughput quantitative gene expression analysis of 96 inflammation associated mediators to measure gene expression levels in BPS and OAB bladder biopsies and compared them to control samples. Then we created a novel animal model of disease by specific proteoglycan deglycosylation of the bladder mucosal barrier, using the bacterial enzymes Chondroitinase ABC and Heparanase III. These enzymes specifically remove the glycosaminoglycan side chains from the urothelial proteoglycan molecules. We tested role of the identified mediators in this animal model. In addition, in order to determine on which patients peripheral treatment strategies may work, we assessed the effect of local anaesthetics on patients with bladder pain. Gene expression analysis did not reveal a difference in inflammatory genes in the OAB versus control biopsies. However, several genes were upregulated in BPS versus control samples, from which two genes, FGF7 and CLL21 were correlated with patient clinical phenotypes for ICS/PI symptom and problem indices respectively. In order to determine which patients are likely to respond to treatment, we sought to characterise the bladder pain in BPS patients. Using urodynamics and local anaesthetics, we differentiated patients with peripherally mediated pain and patients with central sensitisation of their pain. Finally to determine the role of these mediators in bladder pain, we created an animal model of disease, which specifically replicates the human pathology: namely disruption in the barrier proteoglycan molecules. CCL21 led to an increase in painrelated behaviour, while FGF7 attenuated this behaviour, as measured by cystometry, spinal c-fos expression and mechanical withdrawal threshold examination. In conclusion, we have identified CCL21 and FGF7 as potential targets for the treatment of BPS. Manipulation of these ligands or their receptors may prove to be valuable previously unexploited targets for the treatment of BPS.

When and how to treat pulmonary non-tuberculous mycobacterial diseases

Nontuberculous mycobacteria are ubiquitous environmental organisms that have been recognised as a cause of pulmonary infection for over 50 years. Traditionally patients have had underlying risk factors for development of disease; however the proportion of apparently immunocompetent patients involved appears to be rising. Not all patients culture-positive for mycobacteria will have progressive disease, making the diagnosis difficult, though criteria to aid in this process are available. The two main forms of disease are cavitary disease (usually involving the upper lobes) and fibronodular bronchiectasis (predominantly middle and lingular lobes). For patients with disease, combination antibiotic therapy for 12-24 months is generally required for successful treatment, and this may be accompanied by drug intolerances and side effects. Published success rates range from 30-82%. As the progression of disease is variable, for some patients, attention to pulmonary hygiene and underlying diseases without immediate antimycobacterial therapy may be more appropriate. Surgery can be a useful adjunct, though is associated with risks. Randomised controlled trials in well described patients would provide stronger evidence-based data to guide therapy of NTM lung diseases, and thus are much needed.

Bayesian methodology for genetics of complex diseases

Genetic research of complex diseases is a challenging, but exciting, area of research. The early development of the research was limited, however, until the completion of the Human Genome and HapMap projects, along with the reduction in the cost of genotyping, which paves the way for understanding the genetic composition of complex diseases. In this thesis, we focus on the statistical methods for two aspects of genetic research: phenotype definition for diseases with complex etiology and methods for identifying potentially associated Single Nucleotide Polymorphisms (SNPs) and SNP-SNP interactions. With regard to phenotype definition for diseases with complex etiology, we firstly investigated the effects of different statistical phenotyping approaches on the subsequent analysis. In light of the findings, and the difficulties in validating the estimated phenotype, we proposed two different methods for reconciling phenotypes of different models using Bayesian model averaging as a coherent mechanism for accounting for model uncertainty. In the second part of the thesis, the focus is turned to the methods for identifying associated SNPs and SNP interactions. We review the use of Bayesian logistic regression with variable selection for SNP identification and extended the model for detecting the interaction effects for population based case-control studies. In this part of study, we also develop a machine learning algorithm to cope with the large scale data analysis, namely modified Logic Regression with Genetic Program (MLR-GEP), which is then compared with the Bayesian model, Random Forests and other variants of logic regression.

Exploring health behaviour determinants of ageing Australians with chronic diseases

Background: Chronic disease presents overwhelming challenges to elderly patients, their families, health care providers and the health care system. The aim of this study was to explore a theoretical model for effective management of chronic diseases, especially type 2 diabetes mellitus and/or cardiovascular disease. The assumed theoretical model considered the connections between physical function, mental health, social support and health behaviours. The study effort was to improve the quality of life for people with chronic diseases, especially type 2 diabetes and/or cardiovascular disease and to reduce health costs. Methods: A cross-sectional post questionnaire survey was conducted in early 2009 from a randomised sample of Australians aged 50 to 80 years. A total of 732 subjects were eligible for analysis. Firstly, factors influencing respondents‘ quality of life were investigated through bivariate and multivariate regression analysis. Secondly, the Theory of Planned Behaviour (TPB) model for regular physical activity, healthy eating and medication adherence behaviours was tested for all relevant respondents using regression analysis. Thirdly, TPB variable differences between respondents who have diabetes and/or cardiovascular disease and those without these diseases were compared. Finally, the TPB model for three behaviours including regular physical activity, healthy eating and medication adherence were tested in respondents with diabetes and/or cardiovascular diseases using Structure Equation Modelling (SEM). Results: This was the first study combining the three behaviours using a TPB model, while testing the influence of extra variables on the TPB model in one study. The results of this study provided evidence that the ageing process was a cumulative effect of biological change, socio-economic environment and lifelong behaviours. Health behaviours, especially physical activity and healthy eating were important modifiable factors influencing respondents‘ quality of life. Since over 80% of the respondents had at least one chronic disease, it was important to consider supporting older people‘s chronic disease self-management skills such as healthy diet, regular physical activity and medication adherence to improve their quality of life. Direct measurement of the TPB model was helpful in understanding respondents‘ intention and behaviour toward physical activity, healthy eating and medication adherence. In respondents with diabetes and/or cardiovascular disease, the TPB model predicted different proportions of intention toward three different health behaviours with 39% intending to engage in physical activity, 49% intending to engage in healthy eating and 47% intending to comply with medication adherence. Perceived behavioural control, which was proven to be the same as self-efficacy in measurement in this study, played an important role in predicting intention towards the three health behaviours. Also social norms played a slightly more important role than attitude for physical activity and medication adherence, while attitude and social norms had similar effects on healthy eating in respondents with diabetes and/or cardiovascular disease. Both perceived behavioural control and intention directly predicted recent actual behaviours. Physical activity was more a volitional control behaviour than healthy eating and medication adherence. Step by step goal setting and motivation was more important for physical activity, while accessibility, resources and other social environmental factors were necessary for improving healthy eating and medication adherence. The extra variables of age, waist circumference, health related quality of life and depression indirectly influenced intention towards the three behaviours mainly mediated through attitude and perceived behavioural control. Depression was a serious health problem that reduced the three health behaviours‘ motivation, mediated through decreased self-efficacy and negative attitude. This research provided evidence that self-efficacy is similar to perceived behavioural control in the TPB model and intention is a proximal goal toward a particular behaviour. Combining four sources of information in the self-efficacy model with the TPB model would improve chronic disease patients‘ self management behaviour and reach an improved long-term treatment outcome. Conclusion: Health intervention programs that target chronic disease management should focus on patients‘ self-efficacy. A holistic approach which is patient-centred and involves a multidisciplinary collaboration strategy would be effective. Supporting the socio-economic environment and the mental/ emotional environment for older people needs to be considered within an integrated health care system.