Measurement of Vital Signs by Lifelight Software in Comparison to Standard of Care Multisite Development (VISION-MD): Protocol for an Observational Study.

BACKGROUND: Measuring vital signs (VS) is an important aspect of clinical care but is time-consuming and requires multiple pieces of equipment and trained staff. Interest in the contactless measurement of VS has grown since the COVID-19 pandemic, including in nonclinical situations. Lifelight is an app being developed as a medical device for the contactless measurement of VS using remote photoplethysmography (rPPG) via the camera on smart devices. The VISION-D (Measurement of Vital Signs by Lifelight Software in Comparison to the Standard of Care-Development) and VISION-V (Validation) studies demonstrated the accuracy of Lifelight compared with standard-of-care measurement of blood pressure, pulse rate, and respiratory rate, supporting the certification of Lifelight as a class I Conformité Européenne (CE) medical device. OBJECTIVE: To support further development of the Lifelight app, the observational VISION Multisite Development (VISION-MD) study is collecting high-quality data from a broad range of patients, including those with VS measurements outside the normal healthy range and patients who are critically ill. METHODS: The study is recruiting adults (aged ≥16 years) who are inpatients (some critically ill), outpatients, and healthy volunteers, aiming to cover a broad range of normal and clinically relevant VS values; there are no exclusion criteria. High-resolution 60-second videos of the face are recorded by the Lifelight app while simultaneously measuring VS using standard-of-care methods (automated sphygmomanometer for blood pressure; finger clip sensor for pulse rate and oxygen saturation; manual counting of respiratory rate). Feedback from patients and nurses who use Lifelight is collected via a questionnaire. Data to estimate the cost-effectiveness of Lifelight compared with standard-of-care VS measurement are also being collected. A new method for rPPG signal processing is currently being developed, based on the identification of small areas of high-quality signals in each individual. Anticipated recruitment is 1950 participants, with the expectation that data from approximately 1700 will be used for software development. Data from 250 participants will be retained to test the performance of Lifelight against predefined performance targets. RESULTS: Recruitment began in May 2021 but was hindered by the restrictions instigated during the COVID-19 pandemic. The development of data processing methodology is in progress. The data for analysis will become available from September 2022, and the algorithms will be refined continuously to improve clinical accuracy. The performance of Lifelight compared with that of the standard-of-care measurement of VS will then be tested. Recruitment will resume if further data are required. The analyses are expected to be completed in early 2023. CONCLUSIONS: This study will support the refinement of data collection and processing toward the development of a robust app that is suitable for routine clinical use. TRIAL REGISTRATION: ClinicalTrials.gov NCT04763746; https://clinicaltrials.gov/ct2/show/NCT04763746. INTERNATIONAL REGISTERED REPORT IDENTIFIER (IRRID): DERR1-10.2196/41533.

Measurement of Vital Signs Using Lifelight Remote Photoplethysmography: Results of the VISION-D and VISION-V Observational Studies.

BACKGROUND: The detection of early changes in vital signs (VSs) enables timely intervention; however, the measurement of VSs requires hands-on technical expertise and is often time-consuming. The contactless measurement of VSs is beneficial to prevent infection, such as during the COVID-19 pandemic. Lifelight is a novel software being developed to measure VSs by remote photoplethysmography based on video captures of the face via the integral camera on mobile phones and tablets. We report two early studies in the development of Lifelight. OBJECTIVE: The objective of the Vital Sign Comparison Between Lifelight and Standard of Care: Development (VISION-D) study (NCT04763746) was to measure respiratory rate (RR), pulse rate (PR), and blood pressure (BP) simultaneously by using the current standard of care manual methods and the Lifelight software to iteratively refine the software algorithms. The objective of the Vital Sign Comparison Between Lifelight and Standard of Care: Validation (VISION-V) study (NCT03998098) was to validate the use of Lifelight software to accurately measure VSs. METHODS: BP, PR, and RR were measured simultaneously using Lifelight, a sphygmomanometer (BP and PR), and the manual counting of RR. Accuracy performance targets for each VS were defined from a systematic literature review of the performance of state-of-the-art VSs technologies. RESULTS: The VISION-D data set (17,233 measurements from 8585 participants) met the accuracy targets for RR (mean error 0.3, SD 3.6 vs target mean error 2.3, SD 5.0; n=7462), PR (mean error 0.3, SD 4.0 vs mean error 2.2, SD 9.2; n=10,214), and diastolic BP (mean error -0.4, SD 8.5 vs mean error 5.5, SD 8.9; n=8951); for systolic BP, the mean error target was met but not the SD (mean error 3.5, SD 16.8 vs mean error 6.7, SD 15.3; n=9233). Fitzpatrick skin type did not affect accuracy. The VISION-V data set (679 measurements from 127 participants) met all the standards: mean error -0.1, SD 3.4 for RR; mean error 1.4, SD 3.8 for PR; mean error 2.8, SD 14.5 for systolic BP; and mean error -0.3, SD 7.0 for diastolic BP. CONCLUSIONS: At this early stage in development, Lifelight demonstrates sufficient accuracy in the measurement of VSs to support certification for a Level 1 Conformité Européenne mark. As the use of Lifelight does not require specific training or equipment, the software is potentially useful for the contactless measurement of VSs by nonclinical staff in residential and home care settings. Work is continuing to enhance data collection and processing to achieve the robustness and accuracy required for routine clinical use. INTERNATIONAL REGISTERED REPORT IDENTIFIER (IRRID): RR2-10.2196/14326.

Developing the Accuracy of Vital Sign Measurements Using the Lifelight Software Application in Comparison to Standard of Care Methods: Observational Study Protocol.

BACKGROUND: Vital sign measurements are an integral component of clinical care, but current challenges with the accuracy and timeliness of patient observations can impact appropriate clinical decision making. Advanced technologies using techniques such as photoplethysmography have the potential to automate noncontact physiological monitoring and recording, improving the quality and accessibility of this essential clinical information. OBJECTIVE: In this study, we aim to develop the algorithm used in the Lifelight software application and improve the accuracy of its estimated heart rate, respiratory rate, oxygen saturation, and blood pressure measurements. METHODS: This preliminary study will compare measurements predicted by the Lifelight software with standard of care measurements for an estimated population sample of 2000 inpatients, outpatients, and healthy people attending a large acute hospital. Both training datasets and validation datasets will be analyzed to assess the degree of correspondence between the vital sign measurements predicted by the Lifelight software and the direct physiological measurements taken using standard of care methods. Subgroup analyses will explore how the performance of the algorithm varies with particular patient characteristics, including age, sex, health condition, and medication. RESULTS: Recruitment of participants to this study began in July 2018, and data collection will continue for a planned study period of 12 months. CONCLUSIONS: Digital health technology is a rapidly evolving area for health and social care. Following this initial exploratory study to develop and refine the Lifelight software application, subsequent work will evaluate its performance across a range of health characteristics, and extended validation trials will support its pathway to registration as a medical device. Innovations in health technology such as this may provide valuable opportunities for increasing the efficiency and accessibility of vital sign measurements and improve health care services on a large scale across multiple health and care settings. INTERNATIONAL REGISTERED REPORT IDENTIFIER (IRRID): DERR1-10.2196/14326.

Monitoring of chimerism following allogeneic haematopoietic stem cell transplantation (HSCT): technical recommendations for the use of short tandem repeat (STR) based techniques, on behalf of the United Kingdom National External Quality Assessment Service for Leucocyte Immunophenotyping Chimerism Working Group.

Analysis of short tandem repeats (STR) is the predominant method for post-transplant monitoring of donor engraftment. It can enable early detection of disease relapse, level of engraftment and provide useful information on the graft-versus-host disease (GVHD)/graft-versus-tumour (GVT) effect, facilitating therapeutic intervention. Harmonization and standardization of techniques and result interpretation is essential to reduce the impact of laboratory variability on both clinical management and the results of multi-centre clinical trials. However, the United Kingdom National External Quality Assessment Service for Leucocyte Immunophenotyping (UK NEQAS LI) has highlighted significant issues inherent in STR testing that impact upon inter- and intra- laboratory variation. We present here consensus best practice guidelines and recommendations for STR chimerism testing, data interpretation and reporting that have been drawn up and agreed by a consortium of 11 UK and Eire clinical laboratories. This document uses data obtained from the UK NEQAS LI Post-Stem Cell Transplant (SCT) Chimerism Monitoring Programme.

Mimicking the tumour microenvironment: three different co-culture systems induce a similar phenotype but distinct proliferative signals in primary chronic lymphocytic leukaemia cells.

Interactions in the tumour microenvironment can promote chronic lymphocytic leukaemia (CLL) cell survival, proliferation and drug resistance. A detailed comparison of three co-culture systems designed to mimic the CLL lymph node and vascular microenvironments were performed; two were mouse fibroblast cell lines transfected with human CD40LG or CD31 and the third was a human microvascular endothelial cell line, HMEC-1. All three co-culture systems markedly enhanced CLL cell survival and induced a consistent change in CLL cell phenotype, characterized by increased expression of CD38, CD69, CD44 and ITGA4 (CD49d); this phenotype was absent following co-culture on untransfected mouse fibroblasts. In contrast to HMEC-1 cells, the CD40LG and CD31-expressing fibroblasts also induced ZAP70 expression and marked CLL cell proliferation as evidenced by carboxyfluorescein succinimidyl ester labelling and increased Ki-67 expression. Taken together, our data show that co-culture on different stroma induced a remarkably similar activation phenotype in CLL cells but only the CD40LG and CD31-expressing fibroblasts increased ZAP70 expression and CLL cell proliferation, indicating that ZAP70 may play a critical role in this process. This comparative study reveals a number of striking similarities between the co-culture systems tested but also highlights important differences that should be considered when selecting which system to use for in-vitro investigations.

Genetic modification of primary chronic lymphocytic leukemia cells with a lentivirus expressing CD38.

Studies of the role of individual genes in chronic lymphocytic leukemia (CLL) have been hampered by the inability to consistently transfect primary tumor cells. Here, we describe a highly efficient method of genetically modifying primary CLL cells using a VSVG pseudotyped lentiviral vector. We transduced CD38 negative CLL cells with a lentiviral vector encoding CD38 which caused increased surface CD38 expression in all the samples tested (n=17) with no evidence of plasmacytoid differentiation. The mean percentage of positive cells expressing CD38 was 87%+/-8.5% and the mean cell viability 74%+/-17%. This high level of transduction of all the CLL cell samples tested demonstrates the utility of this technique which should prove applicable for the introduction and analysis of other genes in these non-dividing cells.

Rapid recovery of lymphocyte subsets is not associated with protection from relapse of myelodysplastic syndromes and acute myeloid leukaemia after haematopoietic stem cell transplantation using a reduced intensity conditioning regimen and alemtuzumab.

Graft-versus-leukaemia (GvL) and graft-versus-host disease (GvHD) are both caused by alloreactive lymphocytes. We previously reported that GvHD correlated with higher numbers of effector CD4 T cells and Natural Killer cells early after allogeneic transplantation using a regimen comprising fludarabine, busulphan and alemtuzumab. Here, we assessed immune cell subset recovery in these patients in the context of early myeloid malignant disease relapse. Despite the close relationship between the GvL and GvHD immune responses, rapid recovery of lymphocyte subsets was not associated with protection from disease relapse. These results indicated that GvL may be weak in this treatment setting for patients with myelodysplastic syndromes and acute myeloid leukaemia. Consistent with low GvL activity, we previously reported that mixed T cell chimaerism had no detrimental effect on relapse in this treatment setting and instead correlated with better outcome because of reduced GvHD incidence. We now report that patients with significantly higher lymphocyte numbers prior to transplantation subsequently maintained the mixed T cell chimaeric state. This pre-transplant profile, together with absence of the early post-transplant signature indicative of GvHD predisposition, could potentially be used to identify patients suitable for early withdrawal of immunosuppression and prophylactic donor leucocyte infusion to boost GvL activity.

Imbalance of effector and regulatory CD4 T cells is associated with graft-versus-host disease after hematopoietic stem cell transplantation using a reduced intensity conditioning regimen and alemtuzumab.

BACKGROUND: A variety of immune pathways can lead to graft-versus-host disease. A better understanding of the type of immune response causing graft-versus-host disease in defined clinical hematopoietic stem cell transplant settings is required to inform development of methods for monitoring patients and providing them tailored care. DESIGN AND METHODS: Twenty-five patients were recruited presenting with myeloid malignancies and treated with a reduced intensity conditioning transplant regimen with graft-versus-host disease prophylaxis comprising in vivo lymphocyte depletion with alemtuzumab and cyclosporin. A prospective study was performed of lymphocyte subset reconstitution in peripheral blood in relation to the incidence of graft-versus-host disease. RESULTS: Acute graft-versus-host disease was associated with significantly higher numbers of natural killer cells and donor-derived effector CD4 T cells (CD45RO(+) CD27(-)) early (day 30) after transplantation (p=0.04 and p=0.02, respectively). This association was evident before the emergence of clinical pathology in six out of seven patients. Although numbers of regulatory CD4 T cells (CD25(high) Foxp3(+)) were similar at day 30 in all patients, a significant deficit in those who developed acute graft-versus-host disease was apparent relative to effector CD4 T cells (median of 41 effectors per regulatory cell compared to 12 to 1 for patients without graft-versus-host disease) (p=0.03). By day 180, a functional regulatory CD4 T-cell population had expanded significantly in patients who developed chronic graft-versus-host disease, reversing the imbalance (median of 3 effectors per regulatory cell compared to 9.6 to 1 for patients without graft-versus-host disease) (p=0.018) suggesting no overt absence of immune regulation in the late onset form of the disease. CONCLUSIONS: Imbalance of effector and regulatory CD4 T cells is a signature of graft-versus-host disease in this transplantation protocol.

Effects on erythropoiesis of alemtuzumab-containing reduced intensity and standard conditioning regimens.

Haemopoietic cell transplantation (HCT) with reduced-intensity conditioning (RIC) has been associated with delayed disappearance of host anti-A and anti-B isohaemaglutinins and hindrance of donor erythropoiesis in major ABO mismatched transplants. Erythroid recovery, disappearance of recipient type and appearance of donor-type isohaemaglutinins was compared in 84 patients undergoing RIC and 50 patients with standard-conditioning (SCo) HCT. All patients received alemtuzumab as part of their conditioning. The incidence of immune-mediated anaemia and red cell transfusion usage were also compared. Immune factors affecting post-transplant erythroid kinetics showed little variance between different conditioning regimens. Disappearance of recipient isohaemaglutinins and emergence of donor red cells proceeded at similar rates in RIC and SCo transplants; the effects of ABO mismatch were marginal. Pure red cell aplasia, alloimmune haemolysis and autoimmune haemolytic anaemia were not more common in RIC transplants. We believe that alemtuzumab played a critical role in dampening immune reactions of both the host and the donor. Patients in both conditioning groups had similar post-transplant erythroid burst-forming unit (BFU-E) counts; BFU-E chimaerism analysis showed that 90-100% progenitors were of donor origin. However, transfusion requirements were significantly higher in the SCo group, due at least partly to earlier onset of bone marrow hypoplasia.

Delayed attainment of full donor chimaerism following alemtuzumab-based reduced-intensity conditioning haematopoeitic stem cell transplantation for acute myeloid leukaemia and myelodysplastic syndromes is associated with improved outcomes.

This prospective study evaluated the kinetics of lymphoid (CD3) engraftment in 110 patients with acute myeloid leukaemia (AML) and myelodysplastic syndromes (MDS) after allogeneic transplantation and conditioning with fludarabine, busulphan and alemtuzumab, using ciclosporin for post-transplant immunosuppression. Declining donor CD3 chimaerism beyond day+100 was treated with pre-emptive donor lymphocyte infusion (pDLI). The median age of patients was 53.0 years (range: 19-72 years), and the median follow-up was 690 d (range:168-1470 d). Patients achieving full CD3 donor chimaerism (FDC, n = 46) by day+100 had a significantly inferior disease-free survival (DFS) and overall survival (OS) compared to patients with mixed donor chimaerism (MDC, n = 59). Twenty patients had stable MDC and did not require pDLI. Patients attaining early FDC had a higher transplant-related mortality compared to those who maintained stable levels of MDC (P = 0.02), with no difference between the FDC and pDLI groups (P = 0.07). There was no difference in relapse between all three groups (P = 0.21). On multivariate analysis, only CD3 chimaerism status at day+100 and disease status at transplantation had a significant effect on DFS and OS. In patients with AML/MDS undergoing alemetuzumab based-RIC HSCT, prolonged MDC beyond day+100 is associated with an improved OS. Future studies need to be directed towards establishing the underlying factors that dictate T-cell engraftment, expansion and homing post-transplantation.

Outcomes of alemtuzumab-based reduced intensity conditioning stem cell transplantation using unrelated donors for myelodysplastic syndromes.

This prospective study evaluated the outcomes of 75 successive patients receiving a FBC (fludarabine, busulphan, alemtuzumab) reduced-intensity conditioning (RIC) regimen for myelodysplastic syndromes (MDS) using volunteer unrelated donors(VUD). The prognostic significance of a variety of clinical variables including the recently described haematopoietic cell transplantation co-morbidity index (HCT-CI) was assessed. The median age of the cohort was 52.0 years (range: 19-68 years) with a median follow-up of 1038.5 d. Forty-nine patients (65%) had an International Prognostic Scoring System stage of > or = Intermediate-2, 35 (46%) had intermediate or poor risk cytogenetics, and 23 patients(31%) were human leucocyte antigen-mismatched. The actuarial 3-year overall survival (OS) and disease-free survival (DFS) was 43% [95% confidence interval (CI): 37-49] and 41% (95%CI: 35-47) respectively, and the cumulative incidence of extensive chronic graft-versus-host disease was 22%. On multivariate analysis, presence of either one class II mismatch or a two-antigen mismatch adversely influenced transplant-related mortality, DFS and OS. In addition, disease status at transplantation and the haematopoietic cell transplantation-specific comorbidity index were independent variables for overall survival. In contrast, both advanced age and pre-transplant cytogenetic status did not significantly affect overall outcomes. RIC regimens using VUD was associated with durable long-term survival even in older patients with MDS, and the use of a pre-transplant comorbidity index may help to improve patient selection for transplantation.

The AML1-ETO fusion gene promotes extensive self-renewal of human primary erythroid cells.

The t(8;21) translocation, which encodes the AML1-ETO fusion protein (now known as RUNX1-CBF2T1), is one of the most frequent translocations in acute myeloid leukemia, although its role in leukemogenesis is unclear. Here, we report that exogenous expression of AML1-ETO in human CD34(+) cells severely disrupts normal erythropoiesis, resulting in virtual abrogation of erythroid colony formation. In contrast, in bulk liquid culture of purified erythroid cells, we found that while AML1-ETO initially inhibited proliferation during early (erythropoietin [EPO]-independent) erythropoiesis, growth inhibition gave way to a sustained EPO-independent expansion of early erythroid cells that continued for more than 60 days, whereas control cultures became growth arrested after 10 to 13 days (at the EPO-dependent stage of development). Phenotypic analysis showed that although these cells were CD13(-) and CD34(-), unlike control cultures, these cells failed to up-regulate CD36 or to down-regulate CD33, suggesting that expression of AML1-ETO suppressed the differentiation of these cells and allowed extensive self-renewal to occur. In the early stages of this expansion, addition of EPO was able to promote both phenotypic (CD36(+), CD33(-), glycophorin A(+)) and morphologic differentiation of these cells, almost as effectively as in control cultures. However, with extended culture, cells expressing AML1-ETO became refractory to addition of this cytokine, suggesting that a block in differentiation had been established. These data demonstrate the capacity of AML1-ETO to promote the self-renewal of human hematopoietic cells and therefore support a causal role for t(8;21) translocations in leukemogenesis.