Mapping human haematopoietic stem cells from haemogenic endothelium to birth.

The ontogeny of human haematopoietic stem cells (HSCs) is poorly defined owing to the inability to identify HSCs as they emerge and mature at different haematopoietic sites1. Here we created a single-cell transcriptome map of human haematopoietic tissues from the first trimester to birth and found that the HSC signature RUNX1+HOXA9+MLLT3+MECOM+HLF+SPINK2+ distinguishes HSCs from progenitors throughout gestation. In addition to the aorta-gonad-mesonephros region, nascent HSCs populated the placenta and yolk sac before colonizing the liver at 6 weeks. A comparison of HSCs at different maturation stages revealed the establishment of HSC transcription factor machinery after the emergence of HSCs, whereas their surface phenotype evolved throughout development. The HSC transition to the liver marked a molecular shift evidenced by suppression of surface antigens reflecting nascent HSC identity, and acquisition of the HSC maturity markers CD133 (encoded by PROM1) and HLA-DR. HSC origin was tracked to ALDH1A1+KCNK17+ haemogenic endothelial cells, which arose from an IL33+ALDH1A1+ arterial endothelial subset termed pre-haemogenic endothelial cells. Using spatial transcriptomics and immunofluorescence, we visualized this process in ventrally located intra-aortic haematopoietic clusters. The in vivo map of human HSC ontogeny validated the generation of aorta-gonad-mesonephros-like definitive haematopoietic stem and progenitor cells from human pluripotent stem cells, and serves as a guide to improve their maturation to functional HSCs.

MLLT3 governs human haematopoietic stem-cell self-renewal and engraftment.

Limited knowledge of the mechanisms that govern the self-renewal of human haematopoietic stem cells (HSCs), and why this fails in culture, have impeded the expansion of HSCs for transplantation1. Here we identify MLLT3 (also known as AF9) as a crucial regulator of HSCs that is highly enriched in human fetal, neonatal and adult HSCs, but downregulated in culture. Depletion of MLLT3 prevented the maintenance of transplantable human haematopoietic stem or progenitor cells (HSPCs) in culture, whereas stabilizing MLLT3 expression in culture enabled more than 12-fold expansion of transplantable HSCs that provided balanced multilineage reconstitution in primary and secondary mouse recipients. Similar to endogenous MLLT3, overexpressed MLLT3 localized to active promoters in HSPCs, sustained levels of H3K79me2 and protected the HSC transcriptional program in culture. MLLT3 thus acts as HSC maintenance factor that links histone reader and modifying activities to modulate HSC gene expression, and may provide a promising approach to expand HSCs for transplantation.

Anti-CCR9 chimeric antigen receptor T cells for T-cell acute lymphoblastic leukemia.

T cell acute lymphoblastic leukemia (T-ALL) is an aggressive malignancy of immature T lymphocytes, associated with higher rates of induction failure compared with those in B cell acute lymphoblastic leukemia. The potent immunotherapeutic approaches applied in B cell acute lymphoblastic leukemia, which have revolutionized the treatment paradigm, have proven more challenging in T-ALL, largely due to a lack of target antigens expressed on malignant but not healthy T cells. Unlike B cell depletion, T-cell aplasia is highly toxic. Here, we show that the chemokine receptor CCR9 is expressed in >70% of cases of T-ALL, including >85% of relapsed/refractory disease, and only on a small fraction (<5%) of normal T cells. Using cell line models and patient-derived xenografts, we found that chimeric antigen receptor (CAR) T-cells targeting CCR9 are resistant to fratricide and have potent antileukemic activity both in vitro and in vivo, even at low target antigen density. We propose that anti-CCR9 CAR-T cells could be a highly effective treatment strategy for T-ALL, avoiding T cell aplasia and the need for genome engineering that complicate other approaches.

The impact of continuous lenalidomide maintenance treatment on people living with multiple myeloma-a single-centre, qualitative service evaluation study.

PURPOSE: Continuous lenalidomide maintenance treatment after autologous stem cell transplantation delivers improvement in progression free and overall survival among newly diagnosed multiple myeloma patients and has been the standard of care in the UK since March 2021. However, there is scant information about its impact on patients' day-to-day lives. This service evaluation aimed to qualitatively assess patients receiving lenalidomide treatment at a cancer centre in London, in order that the service might better align with needs and expectations of patients. METHODS: We conducted 20 semi-structured interviews among myeloma patients who were on continuous lenalidomide maintenance treatment at a specialist cancer centre in London. Members of the clinical team identified potentially eligible participants to take part, and convenience sampling was used to select 10 male and 10 female patients, median age of 58 (range, 45-71). The median treatment duration was 11 months (range, 1-60 months). Participants were qualitatively interviewed following the same semi-structured interview guide, which was designed to explore patient experience and insights of lenalidomide. Reflexive thematic analysis was used for data analysis. RESULTS: Four overarching themes were as follows: (i) lenalidomide: understanding its role and rationale; (ii) reframing the loss of a treatment-free period to a return to normal life; (iii) the reality of being on lenalidomide: balancing hopes with hurdles; (iv) gratitude and grievances: exploring mixed perceptions of care and communication. Results will be used to enhance clinical services by tailoring communication to better meet patients' preferences when making treatment decisions. CONCLUSION: This study highlights that most patients feel gratitude for being offered continuous lenalidomide and perceive it as alleviating some fears concerning relapse. It reveals variations in side effects in different age groups; younger patients reported no/negligible side effects, whilst several older patients with comorbidities described significant symptom burden, occasionally leading to treatment discontinuation which caused distress at the perceived loss of prolonged remission. Future research should prioritise understanding the unique needs of younger patients living with multiple myeloma.

The impact of COVID-19 on autologous stem cell transplantation in multiple myeloma: A single-centre, qualitative evaluation study.

Autologous stem cell transplantation (ASCT) is standard of care in biologically fit, newly diagnosed multiple myeloma (MM) patients, offering better therapeutic outcomes and improved quality of life (QoL). However, with the UK's 1st national lockdown on 23/03/2020, several guidelines recommended deferring ASCT due to risks of infection, with resource limitations forcing some units to suspend ASCT entirely. Such changes to patients' treatment plans inevitably altered their lived experience during these uncertain times with expected impact on QoL. We conducted a qualitative study using semi-structured interviews to gain insight into MM patients' understanding of their disease, initial therapy and ASCT, and their response to therapy changes. A clinical snapshot of how COVID-19 affected the MM ASCT service in a single UK institution is also provided, including changes to chemotherapy treatment plans, timing, and prioritisation of ASCT. Framework analysis identified 6 overarching themes: (1) beliefs about ASCT, (2) perceptions of information provided about MM and ASCT, (3) high levels of fear and anxiety due to COVID-19, (4) feelings about ASCT disruption or delay due to COVID-19, (5) perceptions of care, and (6) importance of social support. Example subthemes were beliefs that ASCT would provide a long-remission/best chance of normality including freedom from chemotherapy and associated side-effects, disappointment, and devastation at COVID-related treatment delays (despite high anxiety about infection) and exceptionally high levels of trust in the transplant team. Such insights will help us adjust our service and counselling approaches to be more in tune with patients' priorities and expectations.

Maternal-Neonatal Dyad Outcomes of Maternal COVID-19 Requiring Extracorporeal Membrane Support: A Case Series.

OBJECTIVE: This study aimed to describe two cases of acute respiratory distress syndrome (ARDS) secondary to novel coronavirus disease 2019 (COVID-19) in pregnant women requiring extracorporeal membrane oxygenation (ECMO), and resulting in premature delivery. STUDY DESIGN: The clinical course of two women hospitalized with ARDS due to COVID-19 care in our intensive care (ICU) is summarized; both participants provided consent to be included in this case series. RESULTS: Both women recovered with no clinical sequelae. Neonatal outcomes were within the realm of expected for prematurity with the exception of coagulopathy. There was no vertical transmission to the neonates. CONCLUSION: This case series highlights that ECMO is a feasible treatment in the pregnant woman with severe COVID-19 and that delivery can be performed safely on ECMO with no additional risk to the fetus. While ECMO carries its natural risks, it should be considered a viable option during pregnancy and the postpartum period. KEY POINTS: · COVID-19 may present with a more severe course in pregnancy.. · ECMO may be used in pregnant woman with severe COVID-19.. · Delivery can be performed on ECMO without added fetal risk..

An APRIL-based chimeric antigen receptor for dual targeting of BCMA and TACI in multiple myeloma.

B-cell maturation antigen (BCMA) is a promising therapeutic target for multiple myeloma (MM), but expression is variable, and early reports of BCMA targeting chimeric antigen receptors (CARs) suggest antigen downregulation at relapse. Dual-antigen targeting increases targetable tumor antigens and reduces the risk of antigen-negative disease escape. "A proliferation-inducing ligand" (APRIL) is a natural high-affinity ligand for BCMA and transmembrane activator and calcium-modulator and cyclophilin ligand (TACI). We quantified surface tumor expression of BCMA and TACI on primary MM cells (n = 50). All cases tested expressed BCMA, and 39 (78%) of them also expressed TACI. We engineered a third-generation APRIL-based CAR (ACAR), which killed targets expressing either BCMA or TACI (P < .01 and P < .05, respectively, cf. control, effector-to-target [E:T] ratio 16:1). We confirmed cytolysis at antigen levels similar to those on primary MM, at low E:T ratios (56.2% ± 3.9% killing of MM.1s at 48 h, E:T ratio 1:32; P < .01) and of primary MM cells (72.9% ± 12.2% killing at 3 days, E:T ratio 1:1; P < .05, n = 5). Demonstrating tumor control in the absence of BCMA, we maintained cytolysis of primary tumor expressing both BCMA and TACI in the presence of a BCMA-targeting antibody. Furthermore, using an intramedullary myeloma model, ACAR T cells caused regression of an established tumor within 2 days. Finally, in an in vivo model of tumor escape, there was complete ACAR-mediated tumor clearance of BCMA+TACI- and BCMA-TACI+ cells, and a single-chain variable fragment CAR targeting BCMA alone resulted in outgrowth of a BCMA-negative tumor. These results support the clinical potential of this approach.

Adolescent Idiopathic Scoliosis: Common Questions and Answers.

Adolescent idiopathic scoliosis affects 1% to 3% of U.S. adolescents. It is defined by a lateral curvature of the spine (Cobb angle) of at least 10 degrees in the absence of underlying congenital or neuromuscular abnormalities. Adolescent idiopathic scoliosis may be detected via the forward bend test and should be confirmed with scoliometer measurement. Mild scoliosis is usually asymptomatic; it may contribute to musculoskeletal back pain, but there is no evidence that it causes disability or functional impairment. Patients with severe scoliosis (Cobb angle of 40 degrees or more) may have physical pain, cosmetic deformity, psychosocial distress, or, rarely, pulmonary disorders. Several studies have shown modest benefit from bracing and scoliosis-specific physical therapy to limit progression in mild to moderate scoliosis, but there were no effects on quality of life. Because no high-quality studies have proven that surgery is superior to bracing or observation, it should be reserved for severe cases. There is little evidence that treatments improve patient-oriented outcomes. The U.S. Preventive Services Task Force and the American Academy of Family Physicians found insufficient evidence to assess the balance of benefits and harms of screening for adolescent idiopathic scoliosis in children and adolescents 10 to 18 years of age.

6TiSCH on SCµM: Running a Synchronized Protocol Stack without Crystals.

We report the first time-synchronized protocol stack running on a crystal-free device. We use an early prototype of the Single-Chip micro Mote, SCµM, a single-chip 2×3 mm 2 mote-on-a-chip, which features an ARM Cortex-M0 micro-controller and an IEEE802.15.4 radio. This prototype consists of an FPGA version of the micro-controller, connected to the SCµM chip which implements the radio front-end. We port OpenWSN, a reference implementation of a synchronized protocol stack, onto SCµM. The challenge is that SCµM has only on-chip oscillators, with no absolute time reference such as a crystal. We use two calibration steps - receiving packets via the on-chip optical receiver and RF transceiver - to initially calibrate the oscillators on SCµM so that it can send frames to an off-the-shelf IEEE802.15.4 radio. We then use a digital trimming compensation algorithm based on tick skipping to turn a 567 ppm apparent drift into a 10 ppm drift. This allows us to run a full-featured standards-compliant 6TiSCH network between one SCµM and one OpenMote. This is a step towards realizing the smart dust vision of ultra-small and cheap ubiquitous wireless devices.

Sex hormone drives blood stem cell reproduction.

Stem cells ensure the maintenance of tissue homeostasis throughout life by tightly regulating their self-renewal and differentiation. In a recent study published in Nature, Nakada et al, 2014 unveil an unexpected endocrine mechanism that regulates hematopoietic stem cell (HSC) self-renewal.

Evaluation of B cell maturation antigen as a target for antibody drug conjugate mediated cytotoxicity in multiple myeloma.

B-cell maturation antigen (BCMA, also termed TNFRSF17) is an attractive therapeutic target due to its restricted expression on normal and malignant plasma cells (PC). GSK2857916 (or J6M0-MMAF) is a BCMA-specific antibody conjugated to the microtubule-disrupting agent monomethyl auristatin F (MMAF) via a protease-resistant linker. To evaluate the clinical potential of this agent, tumour cells from seventy multiple myeloma (MM) patients were assessed for BCMA expression by immunohistochemistry and flow cytometry. All patients tested expressed BCMA, at varying levels, and both surface and intracellular expression were observed. BCMA expression is maintained through relapse, extramedullary spread and in residual disease post therapy. BCMA levels may also be prognostically useful as higher levels of BCMA were associated with poorer outcomes, even taking into account genetic risk. We observed rapid internalization of surface BCMA and newly expressed protein by 1 h, suggesting a mechanism for J6M0-MMAF activity even with low surface antigen. J6M0-MMAF mediated cytotoxicity of MM cells varied with dose and antigen levels, with clonogenic progenitors killed at lower doses than mature cells. In comparison, J6M0-MMAF killing of primary CD138(+) myeloma cells occurred with slower kinetics. Our observations support BCMA to be a promising therapeutic target in MM for novel therapies such as J6M0-MMAF.

How healthcare systems evaluate their advance care planning initiatives: Results from a systematic review.

BACKGROUND: Advance care planning initiatives are being implemented across healthcare systems around the world, but how best to evaluate their implementation is unknown. AIM: To identify gaps and/or redundancies in current evaluative strategies to help healthcare systems develop future evaluative frameworks for ACP. DESIGN: Systematic review. METHODS: Peer-reviewed and gray literature searches were conducted till February 2015 to answer: "What methods have healthcare systems used to evaluate implementation of advance care planning initiatives?" A PICOS framework was developed to identify articles describing the implementation and evaluation of a health system-level advance care planning initiative. Outcome measures were mapped onto a conceptual quality indicator framework based on the Institute of Medicine and Donabedian models of healthcare quality. RESULTS: A total of 46 studies met inclusion criteria for analysis. Most articles reported on single parts of a healthcare system (e.g. continuing care). The most common outcome measures pertained to document completion, followed by healthcare resource use. Patient-, family-, or healthcare provider-reported outcomes were less commonly measured. Concordance measures (e.g. dying in place of choice) were reported by only 26% of studies. The conceptual quality indicator framework identified gaps and redundancies in measurement and is presented as a potential foundation from which to develop a comprehensive advance care planning evaluation framework. CONCLUSION: Document completion is frequently used to evaluate advance care planning program implementation; capturing the quality of care appears to be more difficult. This systematic review provides health system administrators with a comprehensive summary of measures used to evaluate advance care planning and may identify gaps in evaluation within their local context.

Characterization of Organic Phosphorus Form and Bioavailability in Lake Sediments using P Nuclear Magnetic Resonance and Enzymatic Hydrolysis.

Lake sediments are known to be a significant source of phosphorus (P) to plankton populations under certain biogeochemical conditions; however, the contribution of sediment organic P (P) to internal P loads remains poorly understood. We investigated P speciation and bioavailability in sediments collected over multiple months from a shallow, eutrophic bay in Lake Champlain (Missisquoi Bay, VT) using solution P nuclear magnetic resonance (NMR) spectroscopy and enzymatic hydrolysis (EH) analysis of sediments collected during years with (2008) and without (2007) algal blooms. Sediments collected during bloom onset (July) and peak bloom (August) months contained the largest proportion of enzyme-labile P, whereas pre- and postbloom sediments were primarily composed of nonlabile P. Monoester P to diester P ratios changed with respect to depth, particularly during bloom periods. Monoester P and DNA accumulation, likely from settling particulate matter, began at the onset of the bloom and continued into October 2008 during the postbloom period. The disappearance of inositol hexakisphosphate stereoisomers and the generation of orthophosphate at lower sediment depths was also evident in August 2008. Principal components analysis of EH and NMR species proportions confirmed differences between sediment cores collected during bloom onset and peak bloom, compared with pre- and postbloom sediments. Large enzyme-labile and P species proportions corresponded to increased sediment P flux and reduced manganese and iron species in porewater. These findings suggest that interseasonal changes in P speciation may influence P mobility in sediments and contribute to important feedback dynamics between biological productivity and sediment water interface geochemistry.