Exploratory Analysis of TP53\textit{TP53} Mutations in Circulating Tumour DNA as Biomarkers of Treatment Response for Patients with Relapsed High-Grade Serous Ovarian Carcinoma: A Retrospective Study

$\textbf{Background:}$ Circulating tumour DNA (ctDNA) carrying tumour-specific sequence alterations may provide a minimally invasive means to dynamically assess tumour burden and response to treatment in cancer patients. Somatic $\textit{TP53}$ mutations are a defining feature of high-grade serous ovarian carcinoma (HGSOC). We tested whether these mutations could be used as personalised markers to monitor tumour burden and early changes as a predictor of response and time to progression (TTP). $\textbf{Methods and Findings:}$ We performed a retrospective analysis of serial plasma samples collected during routine clinical visits from 40 patients with HGSOC undergoing heterogeneous standard of care treatment. Patient-specific $\textit{TP53}$ assays were developed for 31 unique mutations identified in formalin-fixed paraffin-embedded tumour DNA from these patients. These assays were used to quantify ctDNA in 318 plasma samples using microfluidic digital PCR. The $\textit{TP53}$ mutant allele fraction (TP53MAF) was compared to serum CA-125, the current gold-standard response marker for HGSOC in blood, as well as to disease volume on computed tomography scans by volumetric analysis. Changes after one cycle of treatment were compared with TTP. The median TP53MAF prior to treatment in 51 relapsed treatment courses was 8% (interquartile range [IQR] 1.2%-22%) compared to 0.7% (IQR 0.3%-2.0%) for seven untreated newly diagnosed stage IIIC/IV patients. TP53MAF correlated with volumetric measurements (Pearson $r$ = 0.59, $p$ 32 cm$^3$, ctDNA was detected at ≥20 amplifiable copies per millilitre of plasma. In 49 treatment courses for relapsed disease, pre-treatment TP53MAF concentration, but not CA-125, was associated with TTP. Response to chemotherapy was seen earlier with ctDNA, with a median time to nadir of 37 d (IQR 28-54) compared with a median time to nadir of 84 d (IQR 42-116) for CA-125. In 32 relapsed treatment courses evaluable for response after one cycle of chemotherapy, a decrease in TP53MAF of >60% was an independent predictor of TTP in multivariable analysis (hazard ratio 0.22, 95% CI 0.07-0.67, $p$ = 0.008). Conversely, a decrease in TP53MAF of ≤60% was associated with poor response and identified cases with TTP < 6 mo with 71% sensitivity (95% CI 42%-92%) and 88% specificity (95% CI 64%-99%). Specificity was improved when patients with recent drainage of ascites were excluded. Ascites drainage led to a reduction of TP53MAF concentration. The limitations of this study include retrospective design, small sample size, and heterogeneity of treatment within the cohort. $\textbf{Conclusions:}$ In this retrospective study, we demonstrated that ctDNA is correlated with volume of disease at the start of treatment in women with HGSOC and that a decrease of ≤60% in TP53MAF after one cycle of chemotherapy was associated with shorter TTP. These results provide evidence that ctDNA has the potential to be a highly specific early molecular response marker in HGSOC and warrants further investigation in larger cohorts receiving uniform treatment.This work was supported by Cancer Research UK Grant numbers: A15601 (JDB), A11906 (NR), A20240 (NR), A18072 (JDB). JDB was supported by the National Institute for Health Research Cambridge Biomedical Research Centre. CAP was supported in part by the Academy of Medical Sciences, the Wellcome Trust, British Heart Foundation and Arthritis Research UK

Detection of heat shock protein 70 in choroidal neovascular membranes secondary to age related macular degeneration

<p>Abstract</p> <p>Background</p> <p>Heat shock proteins are acute phase proteins that are upregulated in inflammation or following thermal stress. We analyzed the presence of the heat shock protein 70 (Hsp 70) in choroidal neovascular (CNV) membranes secondary to AMD after treatment with verteporphin photodynamic therapy (PDT) or transpupillary thermo therapy (TTT) to determine whether treatment correlated with the presence of Hsp70.</p> <p>Results</p> <p>CNV membranes were removed by pars plana vitrectomy (ppV) and subretinal extraction. The membranes were analysed by light microscopy and the presence of Hsp 70 was examined using histochemistry. HeLa Cells served as controls.</p> <p>Of the 14 membranes analysed 11 were Hsp70 positive and 3 negative. In the no pre-treatment group of 8 membranes 6 were Hsp70 positive and 2 negative; in the PTD group all 4 membranes were positive and in the TTT group 1 membrane was positive and 1 membrane was negative for Hsp70.</p> <p>Conclusion</p> <p>Hsp70 is present in the most CNV membranes secondary to AMD. Pre-treatment of the membrane with PTD or TTT does not appear to influence the expression of Hsp70.</p

What Lies behind the Wish to Hasten Death? A Systematic Review and Meta-Ethnography from the Perspective of Patients

BACKGROUND: There is a need for an in-depth approach to the meaning of the wish to hasten death (WTHD). This study aims to understand the experience of patients with serious or incurable illness who express such a wish. METHODS AND FINDINGS: Systematic review and meta-ethnography of qualitative studies from the patient's perspective. Studies were identified through six databases (ISI, PubMed, PsycINFO, CINAHL, CUIDEN and the Cochrane Register of Controlled Trials), together with citation searches and consultation with experts. Finally, seven studies reporting the experiences of 155 patients were included. The seven-stage Noblit and Hare approach was applied, using reciprocal translation and line-of-argument synthesis. Six main themes emerged giving meaning to the WTHD: WTHD in response to physical/psychological/spiritual suffering, loss of self, fear of dying, the desire to live but not in this way, WTHD as a way of ending suffering, and WTHD as a kind of control over one's life ('having an ace up one's sleeve just in case'). An explanatory model was developed which showed the WTHD to be a reactive phenomenon: a response to multidimensional suffering, rather than only one aspect of the despair that may accompany this suffering. According to this model the factors that lead to the emergence of WTHD are total suffering, loss of self and fear, which together produce an overwhelming emotional distress that generates the WTHD as a way out, i.e. to cease living in this way and to put an end to suffering while maintaining some control over the situation. CONCLUSIONS: The expression of the WTHD in these patients is a response to overwhelming emotional distress and has different meanings, which do not necessarily imply a genuine wish to hasten one's death. These meanings, which have a causal relationship to the phenomenon, should be taken into account when drawing up care plans

A Genome-Wide Analysis of Promoter-Mediated Phenotypic Noise in Escherichia coli

Gene expression is subject to random perturbations that lead to fluctuations in the rate of protein production. As a consequence, for any given protein, genetically identical organisms living in a constant environment will contain different amounts of that particular protein, resulting in different phenotypes. This phenomenon is known as “phenotypic noise.” In bacterial systems, previous studies have shown that, for specific genes, both transcriptional and translational processes affect phenotypic noise. Here, we focus on how the promoter regions of genes affect noise and ask whether levels of promoter-mediated noise are correlated with genes' functional attributes, using data for over 60% of all promoters in Escherichia coli. We find that essential genes and genes with a high degree of evolutionary conservation have promoters that confer low levels of noise. We also find that the level of noise cannot be attributed to the evolutionary time that different genes have spent in the genome of E. coli. In contrast to previous results in eukaryotes, we find no association between promoter-mediated noise and gene expression plasticity. These results are consistent with the hypothesis that, in bacteria, natural selection can act to reduce gene expression noise and that some of this noise is controlled through the sequence of the promoter region alon

Diagnostic challenges of early Lyme disease: Lessons from a community case series

<p>Abstract</p> <p>Background</p> <p>Lyme disease, the most common vector-borne infection in North America, is increasingly reported. When the characteristic rash, erythema migrans, is not recognized and treated, delayed manifestations of disseminated infection may occur. The accuracy of diagnosis and treatment of early Lyme disease in the community is unknown.</p> <p>Methods</p> <p>A retrospective, consecutive case series of 165 patients presenting for possible early Lyme disease between August 1, 2002 and August 1, 2007 to a community-based Lyme referral practice in Maryland. All patients had acute symptoms of less than or equal to 12 weeks duration. Patients were categorized according to the Centers for Disease Control and Prevention criteria and data were collected on presenting history, physical findings, laboratory serology, prior diagnoses and prior treatments.</p> <p>Results</p> <p>The majority (61%) of patients in this case series were diagnosed with early Lyme disease. Of those diagnosed with early Lyme disease, 13% did not present with erythema migrans; of those not presenting with a rash, 54% had been previously misdiagnosed. Among those with a rash, the diagnosis of erythema migrans was initially missed in 23% of patients whose rash was subsequently confirmed. Of all patients previously misdiagnosed, 41% had received initial antibiotics likely to be ineffective against Lyme disease.</p> <p>Conclusion</p> <p>For community physicians practicing in high-risk geographic areas, the diagnosis of Lyme disease remains a challenge. Failure to recognize erythema migrans or alternatively, viral-like presentations without a rash, can lead to missed or delayed diagnosis of Lyme disease, ineffective antibiotic treatment, and the potential for late manifestations.</p

Health care costs, utilization and patterns of care following Lyme disease

BACKGROUND:Lyme disease is the most frequently reported vector borne infection in the United States. The Centers for Disease Control have estimated that approximately 10% to 20% of individuals may experience Post-Treatment Lyme Disease Syndrome - a set of symptoms including fatigue, musculoskeletal pain, and neurocognitive complaints that persist after initial antibiotic treatment of Lyme disease. Little is known about the impact of Lyme disease or post-treatment Lyme disease symptoms (PTLDS) on health care costs and utilization in the United States. OBJECTIVES:1) to examine the impact of Lyme disease on health care costs and utilization, 2) to understand the relationship between Lyme disease and the probability of developing PTLDS, 3) to understand how PTLDS may impact health care costs and utilization. METHODS:This study utilizes retrospective data on medical claims and member enrollment for persons aged 0-64 years who were enrolled in commercial health insurance plans in the United States between 2006-2010. 52,795 individuals treated for Lyme disease were compared to 263,975 matched controls with no evidence of Lyme disease exposure. RESULTS:Lyme disease is associated with $2,968 higher total health care costs (95$3,798 higher total health care costs (95% CI: 3,542-4,055, p<.001) and 66% more outpatient visits (95% CI: 64%-69%, p<.001) over a 12-month period, relative to those with no PTLDS-related diagnoses. CONCLUSIONS:Lyme disease is associated with increased costs above what would be expected for an easy to treat infection. The presence of PTLDS-related diagnoses after treatment is associated with significant health care costs and utilization

Phenotypic Variation and Bistable Switching in Bacteria

Microbial research generally focuses on clonal populations. However, bacterial cells with identical genotypes frequently display different phenotypes under identical conditions. This microbial cell individuality is receiving increasing attention in the literature because of its impact on cellular differentiation, survival under selective conditions, and the interaction of pathogens with their hosts. It is becoming clear that stochasticity in gene expression in conjunction with the architecture of the gene network that underlies the cellular processes can generate phenotypic variation. An important regulatory mechanism is the so-called positive feedback, in which a system reinforces its own response, for instance by stimulating the production of an activator. Bistability is an interesting and relevant phenomenon, in which two distinct subpopulations of cells showing discrete levels of gene expression coexist in a single culture. In this chapter, we address techniques and approaches used to establish phenotypic variation, and relate three well-characterized examples of bistability to the molecular mechanisms that govern these processes, with a focus on positive feedback.