RESUMO
Prion disease is caused by misfolding of the prion protein (PrP) into pathogenic self-propagating conformations, leading to rapid-onset dementia and death. However, elimination of endogenous PrP halts prion disease progression. In this study, we describe Coupled Histone tail for Autoinhibition Release of Methyltransferase (CHARM), a compact, enzyme-free epigenetic editor capable of silencing transcription through programmable DNA methylation. Using a histone H3 tail-Dnmt3l fusion, CHARM recruits and activates endogenous DNA methyltransferases, thereby reducing transgene size and cytotoxicity. When delivered to the mouse brain by systemic injection of adeno-associated virus (AAV), Prnp-targeted CHARM ablates PrP expression across the brain. Furthermore, we have temporally limited editor expression by implementing a kinetically tuned self-silencing approach. CHARM potentially represents a broadly applicable strategy to suppress pathogenic proteins, including those implicated in other neurodegenerative diseases.
Assuntos
Encéfalo , Metilação de DNA , Dependovirus , Inativação Gênica , Histonas , Proteínas Priônicas , Animais , Humanos , Camundongos , Encéfalo/metabolismo , Dependovirus/genética , DNA (Citosina-5-)-Metiltransferases/metabolismo , DNA (Citosina-5-)-Metiltransferases/genética , Histonas/metabolismo , Doenças Priônicas/genética , Doenças Priônicas/metabolismo , Proteínas Priônicas/genética , Proteínas Priônicas/metabolismo , TransgenesRESUMO
A 13-year-old male domestic short-hair cat presented for evaluation of labored breathing, hyporexia, and lethargy. Pertinent initial diagnostics yielded leukocytosis, characterized by neutrophilia and monocytosis. Numerous small, round, magenta granules were observed within all neutrophils in Wright-Giemsa-stained blood films on the day of presentation and the day thereafter. No other neutrophil morphologic abnormalities were present, making cytoplasmic toxicity highly unlikely. Hyperadrenocorticism was diagnosed based on the lack of suppression in a low-dose dexamethasone suppression test, and without other diagnostics, the cat was discharged on trilostane therapy. Neutrophil granules did not stain with Alcian blue pH 1.0, periodic acid-Schiff (PAS), PAS and Alcian blue pH 2.5, and toluidine blue. Electron microscopy identified no differences in the morphology of the secretory granules or other neutrophil features. Metabolic screening tests of the cat's urine did not identify a genetic metabolic disorder. However, serum α- and ß -hexosaminidase (HexA and HexB) activities were 4.3% and 0% of normal controls, respectively, which is supportive of GM2-gangliosidosis, that is, Sandhoff disorder. However, the historical, clinical, and electron microscopy findings did not provide evidence to confirm this genetic defect. To the author's knowledge, this is the first case of magenta-staining granules within neutrophils in a breed other than a Birman, Siamese, or Himalayan.
Assuntos
Doenças do Gato , Neutrófilos , Animais , Gatos , Masculino , Neutrófilos/patologia , Doenças do Gato/patologia , Doenças do Gato/diagnóstico , Doenças do Gato/sangue , Grânulos Citoplasmáticos/ultraestrutura , Grânulos Citoplasmáticos/patologiaRESUMO
Developing vehicles that efficiently deliver genes throughout the human central nervous system (CNS) will broaden the range of treatable genetic diseases. We engineered an adeno-associated virus (AAV) capsid, BI-hTFR1, that binds human transferrin receptor (TfR1), a protein expressed on the blood-brain barrier. BI-hTFR1 was actively transported across human brain endothelial cells and, relative to AAV9, provided 40 to 50 times greater reporter expression in the CNS of human TFRC knockin mice. The enhanced tropism was CNS-specific and absent in wild-type mice. When used to deliver GBA1, mutations of which cause Gaucher disease and are linked to Parkinson's disease, BI-hTFR1 substantially increased brain and cerebrospinal fluid glucocerebrosidase activity compared with AAV9. These findings establish BI-hTFR1 as a potential vector for human CNS gene therapy.
Assuntos
Antígenos CD , Encéfalo , Capsídeo , Técnicas de Transferência de Genes , Vetores Genéticos , Glucosilceramidase , Receptores da Transferrina , Animais , Humanos , Camundongos , Antígenos CD/metabolismo , Antígenos CD/genética , Barreira Hematoencefálica/metabolismo , Encéfalo/metabolismo , Capsídeo/metabolismo , Proteínas do Capsídeo/metabolismo , Proteínas do Capsídeo/genética , Dependovirus , Células Endoteliais/metabolismo , Técnicas de Introdução de Genes , Terapia Genética , Receptores da Transferrina/metabolismo , Receptores da Transferrina/genética , Glucosilceramidase/genética , Doença de Gaucher/genética , Doença de Gaucher/terapia , Doença de Parkinson/genética , Doença de Parkinson/terapiaRESUMO
A circular economy based on symbiotic relationships among sectors, where the waste from one is resource to another, holds promise for cost-effective and sustainable production. This research explores such a model for the agriculture, energy, and construction sectors in California. Here, we develop new an understanding for the synergistic utilization mechanisms for rice hull, a byproduct from rice production, as a feedstock for electricity generation and rice hull ash (RHA) used as a supplementary cementitious material in concrete. A suite of methods including experimental analysis, techno-economic analysis (TEA), and life-cycle assessment (LCA) were applied to estimate the cost and environmental performance of the system. TEA results showed that the electricity price required for break even on expenses without selling RHA is $0.07/kWh, lower than the market price. As such, RHA may be available at little to no cost to concrete producers. Our experimental results showed the viability of RHA to be used as a supplementary cementitious material, meaning it can replace a portion of the cement used in concrete. LCA results showed that replacing 15% of cement with RHA in concrete can reduce carbon dioxide equivalent (CO2e) emissions by 15% while still meeting material performance targets. While the substitution rate of RHA for cement may be modest, RHA generated from California alone could mitigate 0.2% of total CO2e from the entire cement production sector in the United States and 1% in California.
RESUMO
BACKGROUND: High/intermediate-risk pulmonary embolism (PE) confers increased risk of cardiovascular morbidity and mortality. International guidelines recommend the formation of a PE response team (PERT) for PE management because of the complexity of risk stratification and emerging treatment options. However, there are currently no available Australian data regarding outcomes of PE managed through a PERT. AIMS: To analyse the clinical and outcome data of patients from an Australian centre with high/intermediate-risk PE requiring PERT-guided management. METHODS: We performed a retrospective observational study of 75 consecutive patients with high/intermediate-risk PE who had PERT involvement, between August 2018 and July 2021. We recorded clinical and interventional data at the time of PERT and assessed patient outcomes up to 30 days from PERT initiation. We used unpaired t tests to compare right to left ventricular (RV/LV) ratios by computed tomography criteria or transthoracic echocardiogram (TTE) at baseline and after interventions. RESULTS: Data were available for 74 patients. Initial computed tomography pulmonary angiography RV/LV ratio was increased at 1.65 ± 0.5 and decreased to 1.30 ± 0.29 following PERT-guided interventions (P < 0.001). TTE RV/LV ratio also decreased following PERT-guided management (1.09 ± 0.19 vs 0.93 ± 0.17; P < 0.001). 20% of patients had any bleeding complication, but two-thirds were mild, not requiring intervention. All-cause mortality was 6.8%, and all occurred within the first 7 days of admission. CONCLUSION: The PERT model is feasible in a large Australian centre in managing complex and time-critical PE. Our data demonstrate outcomes comparable with existing published international PERT data. However, successful implementation at other Australian institutions may require adequate centre-specific resource availability and the presence of multispeciality input.
Assuntos
Embolia Pulmonar , Humanos , Embolia Pulmonar/terapia , Embolia Pulmonar/mortalidade , Masculino , Feminino , Estudos Retrospectivos , Pessoa de Meia-Idade , Idoso , Austrália/epidemiologia , Ecocardiografia , Equipe de Assistência ao Paciente , Idoso de 80 Anos ou mais , Adulto , Angiografia por Tomografia Computadorizada , Medição de RiscoRESUMO
Plant-based product replacements are gaining popularity. However, the long-term health implications remain poorly understood, and available methods, though accurate, are expensive and burdensome, impeding the study of sufficiently large cohorts. To identify dietary transitions over time, we examine anonymised loyalty-card shopping records from Co-op Food, UK. We focus on 10,626 frequent customers who directly replaced milk with alternative milk. We then use product nutritional information to estimate weekly nutrient intake before and after the transition. 83% who converted to alternative milk saw a fall in iodine (44%), calcium (30%) and vitamin B12 (39%) consumption, with 57% reducing iodine purchase by more than 50%. The decline is even higher for those switching dairy and meat products. Our findings suggest that dietary transitions - such as replacing milk with alternative milk - could lead to nutritional deficiencies, notably iodine, which, if not addressed, may represent a significant public health concern, particularly in countries which do not mandate salt iodisation.
Assuntos
Iodo , Magnoliopsida , Desnutrição , Produtos da Carne , Dieta/efeitos adversos , Ingestão de AlimentosRESUMO
BACKGROUND: Pleural biopsy is the gold standard for diagnosis of pleural malignancy but a significant proportion will have an inconclusive biopsy despite ongoing clinical suspicion of malignancy. We investigated whether positron emission tomography-computed tomography (PET-CT) targeted pleural biopsy is superior to standard CT-guided pleural biopsy following an initial non-diagnostic biopsy. METHODS: The TARGET trial was a multicentre, parallel group randomised trial. Patients with a previous inconclusive pleural biopsy but an ongoing suspicion of pleural malignancy were randomised (1:1) to receive either CT-guided biopsy (standard care) or PET-CT followed by a targeted CT biopsy (intervention). The primary outcome was pleural malignancy correctly identified from the trial biopsy. RESULTS: Between September 2015 and September 2018, 59 participants were randomised from eight UK hospital sites: 29 to CT-only followed by targeted biopsy and 30 to PET-CT followed by targeted biopsy. The proportion of pleural malignancy correctly identified was similar between the groups (risk ratio 1.03 (95% CI 0.83-1.29); p=0.77). The sensitivity of the trial biopsy to identify pleural malignancy was 79% (95% CI 54-94%) in the CT-only group versus 81% (95% CI 54-96%) in the PET-CT group. CONCLUSIONS: The results do not support the practice of PET-CT to guide pleural biopsies in patients with a previous non-diagnostic biopsy. The diagnostic sensitivity in the CT-only group was higher than anticipated and supports the practice of repeating a CT-guided biopsy following an inconclusive result if clinical suspicion of malignancy persists.
Assuntos
Doenças Pleurais , Neoplasias Pleurais , Humanos , Tomografia por Emissão de Pósitrons combinada à Tomografia Computadorizada , Tomografia Computadorizada por Raios X , Biópsia Guiada por Imagem/métodos , Biópsia , Neoplasias Pleurais/diagnóstico por imagem , Neoplasias Pleurais/patologiaRESUMO
Rationale: Assessing the early use of video-assisted thoracoscopic surgery (VATS) or intrapleural enzyme therapy (IET) in pleural infection requires a phase III randomized controlled trial (RCT). Objectives: To establish the feasibility of randomization in a surgery-versus-nonsurgery trial as well as the key outcome measures that are important to identify relevant patient-centered outcomes in a subsequent RCT. Methods: The MIST-3 (third Multicenter Intrapleural Sepsis Trial) was a prospective multicenter RCT involving eight U.K. centers combining on-site and off-site surgical services. The study enrolled all patients with a confirmed diagnosis of pleural infection and randomized those with ongoing pleural sepsis after an initial period (as long as 24 h) of standard care to one of three treatment arms: continued standard care, early IET, or a surgical opinion with regard to early VATS. The primary outcome was feasibility based on >50% of eligible patients being successfully randomized, >95% of randomized participants retained to discharge, and >80% of randomized participants retained to 2 weeks of follow-up. The analysis was performed per intention to treat. Measurements and Main Results: Of 97 eligible patients, 60 (62%) were randomized, with 100% retained to discharge and 84% retained to 2 weeks. Baseline demographic, clinical, and microbiological characteristics of the patients were similar across groups. Median times to intervention were 1.0 and 3.5 days in the IET and surgery groups, respectively (P = 0.02). Despite the difference in time to intervention, length of stay (from randomization to discharge) was similar in both intervention arms (7 d) compared with standard care (10 d) (P = 0.70). There were no significant intergroup differences in 2-month readmission and further intervention, although the study was not adequately powered for this outcome. Compared with VATS, IET demonstrated a larger improvement in mean EuroQol five-dimension health utility index (five-level edition) from baseline (0.35) to 2 months (0.83) (P = 0.023). One serious adverse event was reported in the VATS arm. Conclusions: This is the first multicenter RCT of early IET versus early surgery in pleural infection. Despite the logistical challenges posed by the coronavirus disease (COVID-19) pandemic, the study met its predefined feasibility criteria, demonstrated potential shortening of length of stay with early surgery, and signals toward earlier resolution of pain and a shortened recovery with IET. The study findings suggest that a definitive phase III study is feasible but highlights important considerations and significant modifications to the design that would be required to adequately assess optimal initial management in pleural infection.The trial was registered on ISRCTN (number 18,192,121).
Assuntos
Doenças Transmissíveis , Doenças Pleurais , Sepse , Humanos , Cirurgia Torácica Vídeoassistida/efeitos adversos , Estudos de Viabilidade , Doenças Transmissíveis/etiologia , Sepse/tratamento farmacológico , Sepse/cirurgia , Sepse/etiologia , Terapia EnzimáticaRESUMO
Manatees (Antillean-, Amazonian, and African-) and dugongs belong to the Order Sirenia, and when combined with elephants and rock hyraxes, form the Paenungulata. A bilobed mononuclear cell has previously been identified in elephants and rock hyraxes, but not in manatees and dugongs, with cytochemical staining identifying these cells as bilobed monocytes in elephants. The objective of this study was to characterize leukocytes (white blood cells, WBC) and platelets in blood films of Florida manatees (Trichechus manatus latirostris; n = 8) using one routine hematological (Wright-Giemsa) and eight cytochemical stains: alkaline phosphatase (ALP), α-naphthyl butyrate esterase (ANBE), chloroacetate esterase (CAE), Luna, myeloperoxidase (MPx), periodic acid-Schiff (PAS), Sudan black B (SBB), and toluidine blue (TB). Heterophils and lymphocytes comprised most of the WBC, with low numbers of eosinophils, basophils, and monocytes. Additionally, 1-3% of the WBC were bilobed mononuclear cells. Bilobed mononuclear cell proportions were similar to rock hyraxes, but lower than elephants (approximate range 20-60%). Heterophils and eosinophils were positive for MPx, ALP, SBB, and PAS, with heterophils also being positive for CAE. Most of the lymphocytes were positive for ANBE and they were variably positive for CAE. Monocytes and bilobed mononuclear cells had similar cytochemical staining reactions (variably positive for all stains, except Luna and TB), supporting a monocytic origin, like elephants. Platelets were ANBE- and PAS-positive. Luna stain was useful for identifying eosinophils and TB was uninformative. This study provides new information on the morphological features and cytochemical staining characteristics of WBC and platelets and will aid in obtaining accurate hematological data of Florida manatees.
RESUMO
INTRODUCTION: Malignant pleural effusion (MPE) is common, with 50 000 new cases per year in the UK. MPE causes disabling breathlessness and indicates advanced disease with a poor prognosis. Treatment approaches focus on symptom relief and optimising quality of life (QoL). Patients who newly present with MPE commonly require procedural intervention for both diagnosis and therapeutic benefit.Thoracoscopic pleural biopsies are highly sensitive in diagnosing pleural malignancy. Talc poudrage may be delivered at thoracoscopy (TTP) to prevent effusion recurrence by effecting pleurodesis. Indwelling pleural catheters (IPCs) offer an alternative strategy for fluid control, enabling outpatient management and are often used as 'rescue' therapy following pleurodesis failure or in cases of 'trapped lung'. It is unknown whether combining a TTP with IPC insertion will improve patient symptoms or reduce time spent in the hospital.The randomised thoracoscopic talc poudrage + indwelling pleural catheters versus thoracoscopic talc poudrage only in malignant pleural effusion trial (TACTIC) is the first randomised controlled trial (RCT) to examine the benefit of a combined TTP and IPC procedure, evaluating cost-effectiveness and patient-centred outcomes such as symptoms and QoL. The study remains in active recruitment and has the potential to radically transform the pathway for all patients presenting with MPE. METHODS AND ANALYSIS: TACTIC is an unblinded, multicentre, RCT comparing the combination of TTP with an IPC to TTP alone. Co-primary outcomes are time spent in the hospital and mean breathlessness score over 4 weeks postprocedure. The study will recruit 124 patients and aims to define the optimal pathway for patients presenting with symptomatic MPE. ETHICS AND DISSEMINATION: TACTIC is sponsored by North Bristol NHS Trust and has been granted ethical approval by the London-Brent Research Ethics Committee (REC ref: 21/LO/0495). Publication of results in a peer-reviewed journal and conference presentations are anticipated. TRIAL REGISTRATION: ISRCTN 11058680.
Assuntos
Derrame Pleural Maligno , Humanos , Cateteres de Demora , Dispneia/etiologia , Pleura , Derrame Pleural Maligno/etiologia , Ensaios Clínicos Controlados Aleatórios como Assunto , Talco/uso terapêuticoRESUMO
Introduction: A discussion of 'waves' of the COVID-19 epidemic in different countries is a part of the national conversation for many, but there is no hard and fast means of delineating these waves in the available data and their connection to waves in the sense of mathematical epidemiology is only tenuous. Methods: We present an algorithm which processes a general time series to identify substantial, significant and sustained periods of increase in the value of the time series, which could reasonably be described as 'observed waves'. This provides an objective means of describing observed waves in time series. We use this method to synthesize evidence across different countries to study types, drivers and modulators of waves. Results: The output of the algorithm as applied to epidemiological time series related to COVID-19 corresponds to visual intuition and expert opinion. Inspecting the results of individual countries shows how consecutive observed waves can differ greatly with respect to the case fatality ratio. Furthermore, in large countries, a more detailed analysis shows that consecutive observed waves have different geographical ranges. We also show how waves can be modulated by government interventions and find that early implementation of NPIs correlates with a reduced number of observed waves and reduced mortality burden in those waves. Conclusion: It is possible to identify observed waves of disease by algorithmic methods and the results can be fruitfully used to analyse the progression of the epidemic.
RESUMO
Acute ischaemia of the glans penis is a rare and serious complication following circumcision. We report the case of a teenage boy with glanular ischaemia shortly after circumcision with dorsal penile nerve block. This was successfully treated with total 11 days of topical 2% nitroglycerin ointment, 14 days of oral pentoxifylline 400 mg three times a day and 3 days of epidural (0.2% ropivocaine). There was marked clinical improvement at 4 days with a few patches of cyanosis remaining. Surgical intervention was not required, and the patient was discharged with follow-up review. At 12 days, there was complete resolution of ischaemia and the glans penis appeared normal. We suggest that oral, topical and epidural regimen of vasodilators and anti-sympathomimetic agents can be used in combination for acute ischaemia of the glans penis.
Assuntos
Circuncisão Masculina , Pentoxifilina , Masculino , Humanos , Adolescente , Pentoxifilina/uso terapêutico , Nitroglicerina/uso terapêutico , Pênis , Circuncisão Masculina/efeitos adversos , Isquemia/tratamento farmacológico , Isquemia/etiologiaAssuntos
Ascite , Paracentese , Humanos , Ascite/etiologia , Ascite/terapia , Ultrassonografia , Cirrose HepáticaRESUMO
Developing vehicles that efficiently deliver genes throughout the human central nervous system (CNS) will broaden the range of treatable genetic diseases. We engineered an AAV capsid, BI-hTFR1, that binds human Transferrin Receptor (TfR1), a protein expressed on the blood-brain barrier (BBB). BI-hTFR1 was actively transported across a human brain endothelial cell layer and, relative to AAV9, provided 40-50 times greater reporter expression in the CNS of human TFRC knock-in mice. The enhanced tropism was CNS-specific and absent in wild type mice. When used to deliver GBA1, mutations of which cause Gaucher disease and are linked to Parkinson's disease, BI-hTFR1 substantially increased brain and cerebrospinal fluid glucocerebrosidase activity compared to AAV9. These findings establish BI-hTFR1 as a promising vector for human CNS gene therapy.
RESUMO
INTRODUCTION: Mesothelioma is a heterogeneous disease that can be challenging to monitor and prognosticate. ASSESS-meso is a multicentre, prospective, longitudinal observational cohort study of patients with mesothelioma. The primary aim is to describe different clinical phenotypes and investigate predictive and prognostic factors, including biomarkers from blood and pleural fluid. The secondary aim is to provide a resource for future trials and substudies. METHODS AND ANALYSIS: We aim to recruit 700 patients with a histological, cytological or clinicopathological diagnosis of mesothelioma, at any anatomical site (pleural, peritoneal, pericardial, etc). Longitudinal data will be collected, including clinical information, radiological investigations, blood tests and patient-reported outcome measures for breathlessness, chest pain and sweats. Preplanned analyses will use Cox proportional hazards method to evaluate factors associated with survival, linear and logistic regression models to investigate associations with symptoms, and analysis of variance modelling to explore changes in symptoms over time. ETHICS AND DISSEMINATION: Ethical approval has been granted by the Research Ethics Committee South West-Central Bristol (17-SW-0019) and Health Research Authority (IRAS ID 220360). A study steering committee has been established and results will be published OpenAccess in peer-reviewed journals. TRIAL REGISTRATION NUMBER: ISRCTN: 61861764.
Assuntos
Neoplasias Pulmonares , Mesotelioma Maligno , Mesotelioma , Humanos , Estudos Prospectivos , Mesotelioma/diagnóstico , Biomarcadores , Demografia , Estudos Observacionais como AssuntoRESUMO
An Environmental Justice (EJ) analysis was carried out using full Chemical Transport Models (CTMs) over Los Angeles, California, to determine how the combination of domain size and spatial resolution affects predicted air pollution disparities in present day and future simulations when data support from measurements is not available. One set of simulations used the Weather Research and Forecasting (WRF) model coupled with Chemistry (WRF/Chem) with spatial resolution ranging from 250 m to 36 km, comparable to census tract sizes, over domains ranging in size from 320 km2 to 10,000 km2. A second set of simulations used the UCD/CIT CTM with spatial resolution ranging from 4 km to 24 km over domains ranging in size from 98,000 km2 to 1,000,000 km2. Overall WRF/Chem model accuracy improved approximately 9% as spatial resolution increased from 4 km to 250 m in present-day simulations, with similar results expected for future simulations. Exposure disparity results are consistent with previous findings: the average Non-Hispanic White person in the study domain experiences PM2.5 mass concentrations 6-14% lower than the average resident, while the average Black and African American person experiences PM2.5 mass concentrations that are 3-22% higher than the average resident. Predicted exposure disparities were a function of the model configuration. Increasing the spatial resolution finer than approximately 1 km produced diminishing returns because the increased spatial resolution came at the expense of reduced domain size in order to maintain reasonable computational burden. Increasing domain size to capture regional trends, such as wealthier populations living in coastal areas, identified larger exposure disparities but the benefits were limited. CTM configurations that use spatial resolution/domain size of 1 km/103 km2 and 4 km/104 km2 over Los Angeles can detect a 0.5 µg m-3 exposure difference with statistical power greater than 90%. These configurations represent a balanced approach between statistical power, sensitivity across socio-economic groups, and computational burden when predicting current and future air pollution exposure disparities in Los Angeles.
RESUMO
BACKGROUND: Malignant pleural mesothelioma (MPM) is an aggressive thoracic malignancy with a poor prognosis. Systemic immunotherapy is an effective frontline treatment for MPM, and there is a scientific rationale supporting the possible efficacy of local, i.e. intra-pleural immune modulators. Trial of intra-pleural bacterial immunotherapy (TILT) investigated the feasibility of performing a randomised trial of intra-pleural bacterial immunotherapy in people with MPM, using the trials within cohorts (TwiC) methodology. METHODS: TILT was a multicentre, three-armed, randomised, feasibility TwiC of intra-pleural OK432, BCG, or usual care in people with MPM. Eligible participants were identified from within the ASSESS-meso study, a prospective, longitudinal, observational cohort study, and were randomly selected to be offered a single dose of OK432 or BCG, via an indwelling pleural catheter. The primary outcome was feasibility, evaluated against prespecified recruitment, attrition and data completeness targets. The acceptability of trial processes and interventions was assessed during qualitative interviews with participants and family members at the end of the trial. TILT was registered prospectively on the European Clinical Trials Registry (EudraCT number 2016-004,727-23) and the ISRCTN Register on 04 December 2017. RESULTS: Seven participants were randomised from a planned sample size of 12; thus, the 66% recruitment rate target was not met. Two participants withdrew after randomisation, breaching the pre-stated attrition threshold of 10%. It was not possible to maintain blinding of control participants, which negated a fundamental tenet of the TwiC design. The trial processes and methodology were generally acceptable to participants and relatives, despite several recipients of intra-pleural bacterial agents experiencing significant local and systemic inflammatory responses. CONCLUSION: It was possible to design a clinical trial of an investigational medicinal product based on the TwiC design and to obtain the necessary regulatory approvals. However, whilst acceptable to participants and relatives, the TwiC design was not a feasible method of investigating intra-pleural bacterial immunotherapy in people with MPM. Future trials investigating this topic should consider the eligibility constraints and recruitment difficulties encountered. TRIAL REGISTRATION: TILT was registered prospectively on the European Clinical Trials Registry (EudraCT number 2016-004727-23 ) and the ISRCTN Register ( 10432197 ) on 04 December 2017.