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Late-phase clinical trials investigating metformin as a cancer therapy are underway. However, there remains controversy as to the mode of action of metformin in tumors at clinical doses. We conducted a clinical study integrating measurement of markers of systemic metabolism, dynamic FDG-PET-CT, transcriptomics, and metabolomics at paired time points to profile the bioactivity of metformin in primary breast cancer. We show metformin reduces the levels of mitochondrial metabolites, activates multiple mitochondrial metabolic pathways, and increases 18-FDG flux in tumors. Two tumor groups are identified with distinct metabolic responses, an OXPHOS transcriptional response (OTR) group for which there is an increase in OXPHOS gene transcription and an FDG response group with increased 18-FDG uptake. Increase in proliferation, as measured by a validated proliferation signature, suggested that patients in the OTR group were resistant to metformin treatment. We conclude that mitochondrial response to metformin in primary breast cancer may define anti-tumor effect.

Multiple myeloma (MM) is a haematological malignancy characterised by the proliferation of clonal plasma cells in the bone marrow, usually leading to the secretion of a monoclonal immunoglobulin termed the M-protein. Multiple myeloma is a complex disease, and despite great advances in both detection and treatment of the disease, there remains a vast amount of interest in improving patient outcomes. In the era of novel agents and clinical trials, patients with MM are living longer and there is a requirement for frequent monitoring to assess treatment response and signs of relapse. Despite these improvements in treatment, even those patients achieving deep responses are still relapsing due to the presence of very low levels of residual disease, termed minimal residual disease (MRD). Increasingly sensitive techniques are needed to detect this MRD and, as MM is a bone marrow based disease, the majority of techniques focus on bone marrow based assays. However, as bone marrow sampling is invasive, painful and unpleasant for patients it is important to investigate whether blood-based assays could be equally or more informative of disease status. This project therefore aimed to obtain evidence as to whether blood-based assays can act as a safe and effective marker for disease in the bone marrow of treated multiple myeloma patients. Firstly, a bone marrow flow cytometry assay was validated to act as the gold standard technique. Next, three blood-based assays were investigated: circulating tumour plasma cell measurement, heavy/light chain measurement and mass spectrometry evaluation. This study shows the significance of blood-based assays at different disease stages in both transplant and nontransplant treated patients. These results lead to the suggestion of a testing algorithm focused on the addition of heavy/light chain measurement which can be pre-emptively used to influence decisions regarding bone marrow analysis and treatment, ultimately improving the patient experience.

Rheumatoid Arthritis (RA) is a chronic, systemic autoimmune disease, primarily affecting the small joints of the hands, wrists and feet. Disease onset is insidious with classical symptoms including pain, morning stiffness, and persistent synovitis; accompanied by fatigue, malaise and weight loss. RA patients also suffer from a number of extra-articular manifestations and additional comorbidities, including rheumatoid nodules and scleritis; as well as an increased risk of cardiovascular disease (CVD), lymphoma and infection. Ultimately, RA is a devastatingly progressive disease, if a diagnosis and an effective therapeutic regimen is not established quickly. Identifying when a therapy is proving ineffective can be difficult, however, and is reliant upon accurate formal joint counts, CRP measurements and patient global assessments. These are potentially insensitive means of assessing remission and may result in delayed changes to therapeutic regimens. The identification of new biomarkers to improve the predictive value of these evaluation models would therefore greatly benefit patient care. Based upon current evidence, it has been hypothesized that RA patients who fail to make a clinical improvement following the initiation of csDMARD and bDMARD therapeutic regimens are identifiable by their profile of the following CD4+ T-cell populations: follicular helper, memory helper, antigen-specific and naturally-occurring regulatory CD4+ T-cells. As such, flow cytometric assays have developed for the identification and enumeration of these respective CD4+ T-cell populations; the relative and absolute counts of which were investigated within a cohort of healthy controls, with normal distributions established. Subsequently, we have identified distinct alterations to the CD4+ T-cell compartments of newly-diagnosed, treatment-naïve and longstanding RA patients, with clear associations with patient clinical outcomes observed. Of notable significance, the relative distribution of CCR4- Th2-like follicular helper CD4+ T-cells within newly-diagnosed, treatment-naïve RA patients at the time of diagnosis, has proven highly predictive of therapeutic efficacy; with changes in the relative distribution of Th17-like follicular helper and Th17 memory helper CD4+ T-cells also associated with changes to disease activity over time. Furthermore, consistent with previous studies, the CMV status of newly-diagnosed, treatment-naïve and longstanding RA patients, is associated with an aggravated clinical course; with changes in CMV-specific CCR4+ Th17 cell responses related to changes in disease activity 12-months after the initiation of a csDMARD therapeutic regimen. No such relationships could, however, be discerned within the naturally-occurring regulatory CD4+ T-cell compartment, limiting the utility of employing naturally-occurring regulatory CD4+ T-cell measurements in therapeutic efficacy monitoring. Nevertheless, these preliminary results indicate that the addition of follicular helper, memory helper and CMV-specific CD4+ T-cell responses to current remission assessment practices, could improve the identification of RA therapy non-responders, enabling the swift adjustment of RA patient therapeutic regimens and, consequently, improving disease management.

Hypoxia is known to induce the release of microparticles in vitro. However, few publications have addressed the role of hypoxia in vivo on circulating levels of microparticles. This randomised, controlled, crossover trial aimed to determine the effect of mild hypoxia on in vivo levels of circulating microparticles in healthy individuals. Blood was obtained from 51 healthy male volunteers (mean age of 26.9 years) at baseline altitude (490 m) and after 24 and 48 h at moderate altitude (2,590 m). The order of altitude exposure was randomised. Flow cytometry was used to assess platelet-poor plasma for levels of circulating microparticles derived from platelets, endothelial cells, leucocytes, granulocytes, monocytes, red blood cells and procoagulant microparticles. Mean (standard deviation) oxygen saturation was significantly lower on the first and second day after arrival at 2,590 m, 91.0 (2.0) and 92.0 (2.0) %, respectively, compared to 490 m, 96 (1.0) %, p

Background : Flow cytometry is the most commonly used technology to measure microvesicles (MVs). Despite reported limitations of this technique, MV levels obtained using conventional flow cytometry have yielded many clinically relevant findings, such as associations with disease severity and ability to predict clinical outcomes. This study aims to determine if MV enumeration by flow cytometry correlates with a measurement of their functional capacity, as this may explain how flow cytometry generates clinically relevant results. Methods : One hundred samples from healthy individuals and patients with obstructive sleep apnoea were analysed by conventional flow cytometry (FACSCalibur) and by three functional MV assays: Zymuphen MP-activity in which data were given as phosphatidylserine equivalent, STA ® Phospholipid Procoag Assay expressed as clotting time and Endogenous Thrombin Potential (ETP) reflecting in vitro thrombin generation. Correlations were determined by Spearman correlation. Results : Absolute counts of lactadherin+ procoagulant MVs generated by flow cytometry weakly correlated with the results obtained from the Zymuphen MP-activity ( r =0.5370, p