Gas phase biomarkers for disease diagnostics by ion-mobility

The detection of Volatile Organic Compounds (VOCs) has shown great potential as a noninvasive and rapid diagnostic tool for early cancer and disease detection. The use of VOCs for early disease detection has several advantages and can provide valuable insights into disease pathophysiology. This PhD research aimed to investigate the presence of VOCs in urine samples and the development of chemical fingerprints for various cancers, namely, bladder cancer, colorectal cancer, prostate cancer and hepatocellular cancer, and diseases such as fibrosis and urinary tract infection. This study investigated the possibility of using analytical techniques such as Gas Chromatography-Ion Mobility Spectrometry (GC-IMS), Gas Chromatography-Time of Flight-Mass Spectrometry (GC-TOF-MS), and Electronic Noses (eNoses) for the identification of these biomarkers. The study findings demonstrate the presence of particular VOCs in urine samples. We found that GC-IMS and GC-TOF-MS was able to distinguish between some of the cancers with 100% sensitivity and 100% specificity. During this research, a total of 46 VOCs were identified as relevant for the identification of these cancer groups, with some VOCs being specific to each type of cancer. 13 VOCs with the highest concentration in urine samples of bladder cancer patients were identified in the course of this study. The electronic noses utilized in this research were the AlphaMOS FOX 4000 and PEN3 eNose. The AlphaMOS FOX 4000 was able to distinguish between some of the cancer groups with 100% sensitivity and 100% specificity. Although, PEN3 eNose did not exhibit 100% sensitivity in distinguishing between the different cancer groups, it still demonstrated high levels of sensitivity and specificity. Overall, the study contributes to the advancement of research on the detection of VOCs and provides insights into the potential for using analytical techniques for the detection of VOCs in urine samples. However, there are some limitations to these technologies such as some of them require specialized training and expertise to operate and interpret results accurately and the cost of some of these technologies can be high, making them less accessible. Another limitation is that the detection of VOCs can be affected by factors such as sample collection, handling, and storage, which can impact on the accuracy of results. The second aim of this research presents the development of an in-house device based on Photo Ionisation Detector (PID) for the detection of VOCs. The thesis focuses on the xxii evaluation of the performance of the developed PID+ based device for detecting ISB. Throughout this thesis, we have examined the electronics of PID+. Experiments were conducted to assess the performance of the PID+ at various flow rates, ISB (Isobutylene) gas concentrations, bias voltages of detector plates, and amplifier circuit gains. The results of the study demonstrate the feasibility and efficacy of the developed device in detecting ISB with high response. It was observed that increasing the flow rate of the sample gas has the potential to improve the response time of the detector. Additionally, it was observed that the concentration of the target VOCs, in this case ISB gas, was a crucial factor that affected the response of the detector, with higher concentrations resulting in higher sensitivities. However, it was noted that elevating either the flow rate or the concentration of the target VOCs beyond a certain threshold could lead to the saturation of the output. During the testing of the two versions of PID+, the performance of the PID+ was significantly improved. Nevertheless, more extensive research is required to verify the validity of the results of this study with respect to various VOCs

Mechanistic Modeling of Mycobacterium tuberculosis Infection in Murine Models for Drug and Vaccine Efficacy Studies.

Tuberculosis (TB) drug, regimen, and vaccine development rely heavily on preclinical animal experiments, and quantification of bacterial and immune response dynamics is essential for understanding drug and vaccine efficacy. A mechanism-based model was built to describe Mycobacterium tuberculosis H37Rv infection over time in BALB/c and athymic nude mice, which consisted of bacterial replication, bacterial death, and adaptive immune effects. The adaptive immune effect was best described by a sigmoidal function on both bacterial load and incubation time. Applications to demonstrate the utility of this baseline model showed (i) the important influence of the adaptive immune response on pyrazinamide (PZA) drug efficacy, (ii) a persistent adaptive immune effect in mice relapsing after chemotherapy cessation, and (iii) the protective effect of vaccines after M. tuberculosis challenge. These findings demonstrate the utility of our model for describing M. tuberculosis infection and corresponding adaptive immune dynamics for evaluating the efficacy of TB drugs, regimens, and vaccines

Non-invasive detection and staging of colorectal cancer using a portable electronic nose

Electronic noses (e-nose) offer potential for the detection of cancer in its early stages. The ability to analyse samples in real time, at a low cost, applying easy–to-use and portable equipment, gives e-noses advantages over other technologies, such as Gas Chromatography-Mass Spectrometry (GC-MS). For diseases such as cancer with a high mortality, a technology that can provide fast results for use in routine clinical applications is important. Colorectal cancer (CRC) is among the highest occurring cancers and has high mortality rates, if diagnosed late. In our study, we investigated the use of portable electronic nose (PEN3), with further analysis using GC-TOF-MS, for the analysis of gases and volatile organic compounds (VOCs) to profile the urinary metabolome of colorectal cancer. We also compared the different cancer stages with non-cancers using the PEN3 and GC-TOF-MS. Results obtained from PEN3, and GC-TOF-MS demonstrated high accuracy for the separation of CRC and non-cancer. PEN3 separated CRC from non-cancerous group with 0.81 AUC (Area Under the Curve). We used data from GC-TOF-MS to obtain a VOC profile for CRC, which identified 23 potential biomarker VOCs for CRC. Thus, the PEN3 and GC-TOF-MS were found to successfully separate the cancer group from the non-cancer group

Exploratory study using urinary volatile organic compounds for the detection of hepatocellular carcinoma

Hepatocellular carcinoma (HCC) biomarkers are lacking in clinical practice. We therefore explored the pattern and composition of urinary volatile organic compounds (VOCs) in HCC patients. This was done in order to assess the feasibility of a potential non-invasive test for HCC, and to enhance our understanding of the disease. This pilot study recruited 58 participants, of whom 20 were HCC cases and 38 were non-HCC cases. The non-HCC cases included healthy individuals and patients with various stages of non-alcoholic fatty liver disease (NAFLD), including those with and without fibrosis. Urine was analysed using gas chromatography–ion mobility spectrometry (GC–IMS) and gas chromatography–time-of-flight mass spectrometry (GC–TOF-MS). GC–IMS was able to separate HCC from fibrotic cases with an area under the curve (AUC) of 0.97 (0.91–1.00), and from non-fibrotic cases with an AUC of 0.62 (0.48–0.76). For GC-TOF-MS, a subset of samples was analysed in which seven chemicals were identified and tentatively linked with HCC. These include 4-methyl-2,4-bis(p-hydroxyphenyl)pent-1-ene (2TMS derivative), 2-butanone, 2-hexanone, benzene, 1-ethyl-2-methyl-, 3-butene-1,2-diol, 1-(2-furanyl)-, bicyclo(4.1.0)heptane, 3,7,7-trimethyl-, [1S-(1a,3β,6a)]-, and sulpiride. Urinary VOC analysis using both GC–IMS and GC-TOF-MS proved to be a feasible method of identifying HCC cases, and was also able to enhance our understanding of HCC pathogenesis

Minimal Gluten Exposure Alters Urinary Volatile Organic Compounds in Stable Coeliac Disease

Coeliac disease (CD) patients are distinguishable from healthy individuals via urinary volatile organic compounds (VOCs) analysis. We exposed 20 stable CD patients on gluten-free diet (GFDs) to a 14-day, 3 g/day gluten challenge (GCh), and assessed urinary VOC changes. A control cohort of 20 patients continued on GFD. Urine samples from Days 0, 7, 14, 28 and 56 were analysed using Lonestar FAIMS and Markes Gas Chromatography−Time of Flight−Mass Spectrometer (GC-TOF-MS). VOC signatures on D (day) 7−56 were compared with D0. Statistical analysis was performed using R. In GCh patients, FAIMS revealed significant VOC differences for all time points compared to D0. GC-TOF-MS revealed significant changes at D7 and D14 only. In control samples, FAIMS revealed significant differences at D7 only. GC-TOF-MS detected no significant differences. Chemical analysis via GC-MS-TOF revealed 12 chemicals with significantly altered intensities at D7 vs. D0 for GCh patients. The alterations persisted for six chemicals at D14 and one (N-methyltaurine) remained altered after D14. This low-dose, short-duration challenge was well tolerated. FAIMS and GC-TOF-MS detected VOC signature changes in CD patients when undergoing a minimal GCh. These findings suggest urinary VOCs could have a role in monitoring dietary compliance in CD patients

Urinarv volatiles and chemical characterisation for the non-invasive detection of prostate and bladder cancers

Bladder cancer (BCa) and prostate cancer (PCa) are some of the most common cancers in the world. In both BCa and PCa, the diagnosis is often confirmed with an invasive technique that carries a risk to the patient. Consequently, a non-invasive diagnostic approach would be medically desirable and beneficial to the patient. The use of volatile organic compounds (VOCs) for disease diagnosis, including cancer, is a promising research area that could support the diagnosis process. In this study, we investigated the urinary VOC profiles in BCa, PCa patients and non-cancerous controls by using gas chromatography-ion mobility spectrometry (GC-IMS) and gas chromatography time-of-flight mass spectrometry (GC-TOF-MS) to analyse patient samples. GC-IMS separated BCa from PCa (area under the curve: AUC: 0.97 (0.93–1.00)), BCa vs. non-cancerous (AUC: 0.95 (0.90–0.99)) and PCa vs. non-cancerous (AUC: 0.89 (0.83–0.94)) whereas GC-TOF-MS differentiated BCa from PCa (AUC: 0.84 (0.73–0.93)), BCa vs. non-cancerous (AUC: 0.81 (0.70–0.90)) and PCa vs. non-cancerous (AUC: 0.94 (0.90–0.97)). According to our study, a total of 34 biomarkers were found using GC-TOF-MS data, of which 13 VOCs were associated with BCa, seven were associated with PCa, and 14 VOCs were found in the comparison of BCa and PCa

Non-Invasive Detection and Staging of Colorectal Cancer Using a Portable Electronic Nose

Electronic noses (e-nose) offer potential for the detection of cancer in its early stages. The ability to analyse samples in real time, at a low cost, applying easy–to-use and portable equipment, gives e-noses advantages over other technologies, such as Gas Chromatography-Mass Spectrometry (GC-MS). For diseases such as cancer with a high mortality, a technology that can provide fast results for use in routine clinical applications is important. Colorectal cancer (CRC) is among the highest occurring cancers and has high mortality rates, if diagnosed late. In our study, we investigated the use of portable electronic nose (PEN3), with further analysis using GC-TOF-MS, for the analysis of gases and volatile organic compounds (VOCs) to profile the urinary metabolome of colorectal cancer. We also compared the different cancer stages with non-cancers using the PEN3 and GC-TOF-MS. Results obtained from PEN3, and GC-TOF-MS demonstrated high accuracy for the separation of CRC and non-cancer. PEN3 separated CRC from non-cancerous group with 0.81 AUC (Area Under the Curve). We used data from GC-TOF-MS to obtain a VOC profile for CRC, which identified 23 potential biomarker VOCs for CRC. Thus, the PEN3 and GC-TOF-MS were found to successfully separate the cancer group from the non-cancer group

Electronic Nose for Bladder Cancer Detection

This study outlines the use of an electronic nose as a method for the detection of VOCs as biomarkers of bladder cancer. Here, an AlphaMOS FOX 4000 electronic nose was used for the analysis of urine samples from 15 bladder cancer and 41 non-cancerous patients. The FOX 4000 consists of 18 MOS sensors that were used to differentiate the two groups. The results obtained were analysed using s MultiSens Analyzer and RStudio. The results showed a high separation with sensitivity and specificity of 0.93 and 0.88, respectively, using a Sparse Logistic Regression and 0.93 and 0.76 using a Random Forest classifier. We conclude that the electronic nose shows potential for discriminating bladder cancer from non-cancer subjects using urine samples

Mechanistic Modeling of Mycobacterium tuberculosis Infection in Murine Models for Drug and Vaccine Efficacy Studies.

Tuberculosis (TB) drug, regimen, and vaccine development rely heavily on preclinical animal experiments, and quantification of bacterial and immune response dynamics is essential for understanding drug and vaccine efficacy. A mechanism-based model was built to describe Mycobacterium tuberculosis H37Rv infection over time in BALB/c and athymic nude mice, which consisted of bacterial replication, bacterial death, and adaptive immune effects. The adaptive immune effect was best described by a sigmoidal function on both bacterial load and incubation time. Applications to demonstrate the utility of this baseline model showed (i) the important influence of the adaptive immune response on pyrazinamide (PZA) drug efficacy, (ii) a persistent adaptive immune effect in mice relapsing after chemotherapy cessation, and (iii) the protective effect of vaccines after M. tuberculosis challenge. These findings demonstrate the utility of our model for describing M. tuberculosis infection and corresponding adaptive immune dynamics for evaluating the efficacy of TB drugs, regimens, and vaccines

Pharmacodynamic Correlates of Linezolid Activity and Toxicity in Murine Models of Tuberculosis.

BackgroundLinezolid (LZD) is bactericidal against Mycobacterium tuberculosis, but it has treatment-limiting toxicities. A better understanding of exposure-response relationships governing LZD efficacy and toxicity will inform dosing strategies. Because in vitro monotherapy studies yielded conflicting results, we explored LZD pharmacokinetic/pharmacodynamic (PK/PD) relationships in vivo against actively and nonactively multiplying bacteria, including in combination with pretomanid.MethodsLinezolid multidose pharmacokinetics were modeled in mice. Dose-fractionation studies were performed in acute (net bacterial growth) and chronic (no net growth) infection models. In acute models, LZD was administered alone or with bacteriostatic or bactericidal pretomanid doses. Correlations between PK/PD parameters and lung colony-forming units (CFUs) and complete blood counts were assessed.ResultsOverall, time above minimum inhibitory concentration (T>MIC) correlated best with CFU decline. However, in growth-constrained models (ie, chronic infection, coadministration with pretomanid 50 mg/kg per day), area under the concentration-time curve over MIC (AUC/MIC) had similar explanatory power. Red blood cell counts correlated strongly with LZD minimum concentration (Cmin).ConclusionsAlthough T>MIC was the most consistent correlate of efficacy, AUC/MIC was equally predictive when bacterial multiplication was constrained by host immunity or pretomanid. In effective combination regimens, administering the same total LZD dose less frequently may be equally effective and cause less Cmin-dependent toxicity