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Tissue engineering holds great promise for regenerative medicine, drug discovery and as an alternative to animal models. However, as soon as the dimensions of engineered tissue exceed the diffusion limit of oxygen and nutriments, a necrotic core forms leading to irreversible damage. To overcome this constraint, the establishment of a functional perfusion network is essential and is a major challenge to be met. In this work, we explore a promising Digital Light Processing (DLP) bioprinting approach to encapsulate endothelial progenitor cells (EPCs) in 3D photopolymerized hydrogel scaffolds to guide them towards vascular network formation. We observed that EPCs encapsulated in the appropriate photopolymerized hydrogel can proliferate and self-organize within a few days into branched tubular structures with predefined geometry, forming capillary-like vascular tubes or trees of various diameters (in the range of 10 to 100 μm). Presenting a monolayer wall of endothelial cells strongly connected by tight junctions around a central lumen, these structures can be microinjected with fluorescent dye and are stable for several weeksin vitro. Interestingly, our technology has proven to be versatile in promoting the formation of vascular structures using a variety of vascular cell lines, including EPCs, human vascular endothelial cells (HUVECs) and human dermal lymphatic endothelial cells (HDLECs). We have also demonstrated that these vascular structures can be recovered and manipulated in an alginate patch without altering their shape or viability. This opens new opportunities for future applications, such as stacking these endothelial vascular structures with other cell sheets or multicellular constructs to yield bioengineered tissue with higher complexity and functionality.

Advanced therapy medicinal products (ATMPs) represent a new class of biological medicines. The European regulation has classified ATMPs into three categories: gene therapy medicinal products, somatic cell therapy medicinal products, and Tissue-Engineered products. If one of these categories incorporates a medical device, the medicine is defined as a Combined ATMP. The specificity and complexity of these innovative drugs have required a complete reorganization of hospital and pharmaceutical circuits, from patient eligibility to drug administration. Indeed, increased interaction and collaboration between different healthcare professionals are essential in order to guarantee quality and safety of these innovative medicines.

The public French Cord Blood Banks Network was established in 1999 with the objective of standardizing the practices governing umbilical cord blood (UCB) banking in France. The Network adopted a strategy to optimize its inventory and improve the quality of its banked units based on a quality improvement process using outcome data regularly provided by Eurocord. This study aimed to describe the results, over 10 years, of UCBT facilitated by a national network that used the same criteria of UCB collection and banking and to assess how modifications of banking criteria and unit selection might influence transplant outcomes. Nine hundred and ninety-nine units (593 single-unit and 203 double-unit grafts) were released by the Network to transplant 796 patients with malignant (83%) and non-malignant (17%) diseases. Median cell dose exceeded 3.5 × 107 TNC/kg in 86%. There was a trend to select units more recently collected and with higher cell dose. Neutrophil engraftment was 88.2% (85.7–90.7) and 79.3% (72.6–86.5) respectively for malignant and non-malignant diseases with a trend to faster recovery with higher cell doses. The respective 3-year transplant-related mortality were 31.1% (27.5–35.1) and 34.3% (27.0–43.5). OS was 49% ± 4 in malignant and 62% ± 4 in non-malignant disorders. In multivariate analysis, cell dose was the only unit-related factor associated with outcomes. Our results reflect the benefit on clinical outcomes of the strategy adopted by the Network to bank units with higher cell counts.

Umbilical cord-derived mesenchymal stem/stromal cells (UC-MSCs) emerge as a perspective for therapeutic use in immune and inflammatory diseases. Indeed, immunomodulatory and anti-inflammatory properties, associated to fewer ethical, availability, and safety issues, position UC-MSCs as a promising active substance to develop medicinal products. Since 2007, UC-MSC-based products are classified as advanced therapy medicinal products (ATMP) according to the European Regulation 1394/2007/EC. This new regulatory status required a total adaptation of stakeholders wishing to develop UC-MSC-based ATMPs. Cell production in tissue and cell banks has been replaced by the manufacturing of a medicine, in authorized establishments, according to the good manufacturing practices (GMP) specific to ATMPs. After a brief description of UC-MSCs, we described in this review their recent use in a large panel of immune and inflammatory pathologies, including early and late phase clinical trials. Despite the use of the same product, we noticed an important heterogeneity in terms of indication, posology and study design. Then, we discussed regulatory and manufacturing challenges for stakeholders, especially in terms of process harmonization and cells characterization. Our aim was to point that despite MSCs use for several decades, the development of an UC-MSC-based ATMP remains at this day a real challenge for both academic institutions and pharmaceutical companies.

Summary Background Genome-edited donor-derived allogeneic anti-CD19 chimeric antigen receptor (CAR) T cells offer a novel form of CAR-T-cell product that is available for immediate clinical use, thereby broadening access and applicability. UCART19 is one such product investigated in children and adults with relapsed or refractory B-cell acute lymphoblastic leukaemia. Two multicentre phase 1 studies aimed to investigate the feasibility, safety, and antileukaemic activity of UCART19 in children and adults with relapsed or refractory B-cell acute lymphoblastic leukaemia. Methods We enrolled paediatric or adult patients in two ongoing, multicentre, phase 1 clinical trials to evaluate the safety and antileukaemic activity of UCART19. All patients underwent lymphodepletion with fludarabine and cyclophosphamide with or without alemtuzumab, then children received UCART19 at 1·1–2·3 × 106 cells per kg and adults received UCART19 doses of 6 × 106 cells, 6–8 × 107 cells, or 1·8–2·4 × 108 cells in a dose-escalation study. The primary outcome measure was adverse events in the period between first infusion and data cutoff. These studies were registered at ClinicalTrials.gov, NCT02808442 and NCT02746952. Findings Between June 3, 2016, and Oct 23, 2018, seven children and 14 adults were enrolled in the two studies and received UCART19. Cytokine release syndrome was the most common adverse event and was observed in 19 patients (91%); three (14%) had grade 3–4 cytokine release syndrome. Other adverse events were grade 1 or 2 neurotoxicity in eight patients (38%), grade 1 acute skin graft-versus-host disease in two patients (10%), and grade 4 prolonged cytopenia in six patients (32%). Two treatment-related deaths occurred; one caused by neutropenic sepsis in a patient with concurrent cytokine release syndrome and one from pulmonary haemorrhage in a patient with persistent cytopenia. 14 (67%) of 21 patients had a complete response or complete response with incomplete haematological recovery 28 days after infusion. Patients not receiving alemtuzumab (n=4) showed no UCART19 expansion or antileukaemic activity. The median duration of response was 4·1 months with ten (71%) of 14 responders proceeding to a subsequent allogeneic stem-cell transplant. Progression-free survival at 6 months was 27%, and overall survival was 55%. Interpretation These two studies show, for the first time, the feasibility of using allogeneic, genome-edited CAR T cells to treat patients with aggressive leukaemia. UCART19 exhibited in-vivo expansion and antileukaemic activity with a manageable safety profile in heavily pretreated paediatric and adult patients with relapsed or refractory B-cell acute lymphoblastic leukaemia. The results this study are an encouraging step forward for the field of allogeneic CAR T cells, and UCART19 offers the opportunity to treat patients with rapidly progressive disease and where autologous CAR-T-cell therapy is unavailable. Funding Servier.

The prognosis for adults with relapsed or refractory B-cell acute lymphoblastic leukaemia remains poor. UCART19, an allogeneic genome-edited anti-CD19 chimeric antigen receptor (CAR) T-cell product derived from healthy donors and available for immediate clinical use, offers a potential therapeutic option for such patients. The CALM trial is a first-in-human study evaluating the safety and antileukaemic activity of UCART19 in adult patients with relapsed or refractory B-cell acute lymphoblastic leukaemia.This phase 1, open-label study was conducted at eight centres across France, the UK, the USA, and Japan. Adult patients aged 16-70 years with CD19-positive relapsed or refractory B-cell acute lymphoblastic leukaemia who had morphological relapse or a minimal residual disease level of at least 1 × 10Between Aug 1, 2016, and June 30, 2020, 25 patients were enrolled in the study and treated with UCART19. Median duration of follow-up was 12·8 months (IQR 2·8-24·8). Median age was 37 years (IQR 28-45). 14 (56%) patients were male and 11 (44%) female. 17 (68%) patients were White, two (8%) Black, two (8%) Asian, and four (16%) from other racial or ethnic groups. Three patients developed dose-limiting toxicities (one at each dose level); one had grade 4 cytokine release syndrome and two had grade 4 prolonged cytopenias. Grade 3 or higher cytokine release syndrome was reported in six (24%) patients and grade 3 or higher neurological toxicity in one (4%) patient. Grade 3 or higher infections occurred in seven (28%) patients, and grade 4 prolonged cytopenia in four (16%) patients. Two (8%) patients developed grade 1 acute cutaneous graft-versus-host disease. 14 patients died, nine from progressive disease and five from infections or other complications, of which four were considered to be related to UCART19 or lymphodepletion, or both. After a median of follow-up of 12·8 months (IQR 2·8-24·8), overall response rate was 48% (95% CI 28-69; 12 of 25 patients), duration of response and median relapse-free survival were 7·4 months (95% CI 1·8 to not calculable), progression-free survival was 2·1 months (95% CI 1·2-2·8), and overall survival was 13·4 months (95% CI 4·8-23·0).UCART19 had a manageable safety profile, and showed evidence of antileukaemic activity in heavily pretreated adult patients with relapsed or refractory B-cell acute lymphoblastic leukaemia. This study shows that allogeneic off-the-shelf CAR T cells can be used safely to treat patients with relapsed B-cell acute lymphoblastic leukaemia.Servier.

Abstract: Tumor recurrence is a colossal challenge in clinical oncology. This multifactorial problem is attributed to the emergence of additional genetic mutations and the presence of dormant cancer cells. However, the plasticity of non-stem cancer cells and the acquisition of cancer stem cell (CSC) functionality is another contributing factor to tumor recurrence. Herein, I focus attention on the mechanisms that fuel cancer cell de-differentiation and the interplay between intra-cellular regulators and tumor microenvironment (TME) landscape that promotes cancer cell stemness. Our understanding of the mechanisms underlying tumor cell de-differentiation is crucial for developing innovative therapeutic strategies that prevent cancer from ever recurring.

International audience; Extracellular vesicles (EV) are emergent therapeutic effectors that have reached clinical trial investigation. To translate EV-based therapeutic to clinic, the challenge is to demonstrate quality, safety, and efficacy, as required for any medicinal product. EV research translation into medicinal products is an exciting and challenging perspective. Recent papers, provide important guidance on regulatory aspects of pharmaceutical development, defining EVs for therapeutic applications and critical considerations for the development of potency tests. In addition, the ISEV Task Force on Regulatory Affairs and Clinical Use of EV-based Therapeutics as well as the Exosomes Committee from the ISCT are expected to contribute in an active way to the development of EV-based medicinal products by providing update on the scientific progress in EVs field, information to patients and expert resource network for regulatory bodies. The contribution of our work group "Extracellular Vesicle translatiOn to clinicaL perspectiVEs - EVOLVE France", created in 2020, can be positioned in complement to all these important initiatives. Based on complementary scientific, technical, and medical expertise, we provide EV-specific recommendations for manufacturing, quality control, analytics, non-clinical development, and clinical trials, according to current European legislation. We especially focus on early phase clinical trials concerning immediate needs in the field. The main contents of the investigational medicinal product dossier, marketing authorization applications, and critical guideline information are outlined for the transition from research to clinical development and ultimate market authorization.

Background Severe acute respiratory syndrome coronavirus-2 (SARS–CoV-2)-induced acute respiratory distress syndrome (ARDS) causes high mortality. Umbilical cord-derived mesenchymal stromal cells (UC-MSCs) have potentially relevant immune-modulatory properties, whose place in ARDS treatment is not established. This phase 2b trial was undertaken to assess the efficacy of UC-MSCs in patients with SARS–CoV-2-induced ARDS. Methods This multicentre, double-blind, randomized, placebo-controlled trial (STROMA–CoV-2) recruited adults (≥ 18 years) with SARS–CoV-2-induced early (6 UC-MSCs/kg or placebo (0.9% NaCl) over 5 days after recruitment. For the modified intention-to-treat population, the primary endpoint was the partial pressure of oxygen to fractional inspired oxygen (PaO2/FiO2)-ratio change between baseline (day (D) 0) and D7. Results Among the 107 patients screened for eligibility from April 6, 2020, to October 29, 2020, 45 were enrolled, randomized and analyzed. PaO2/FiO2 changes between D0 and D7 did not differ significantly between the UC-MSCs and placebo groups (medians [IQR] 54.3 [− 15.5 to 93.3] vs 25.3 [− 33.3 to 104.6], respectively; ANCOVA estimated treatment effect 7.4, 95% CI − 44.7 to 59.7; P = 0.77). Six (28.6%) of the 21 UC-MSCs recipients and six of 24 (25%) placebo-group patients experienced serious adverse events, none of which were related to UC-MSCs treatment. Conclusions D0-to-D7 PaO2/FiO2 changes for intravenous UC-MSCs-versus placebo-treated adults with SARS–CoV-2-induced ARDS did not differ significantly. Repeated UC-MSCs infusions were not associated with any serious adverse events during treatment or thereafter (until D28). Larger trials enrolling patients earlier during the course of their ARDS are needed to further assess UC-MSCs efficacy in this context. Trial registration: NCT04333368. Registered 01 April 2020, https://clinicaltrials.gov/ct2/history/NCT04333368.

Throughout mammal erythroid differentiation, erythroblasts undergo enucleation and organelle clearance becoming mature red blood cell. Organelles are cleared by autophagic pathways non-specifically targeting organelles and cytosolic content or by specific mitophagy targeting mitochondria. Mitochondrial functions are essential to coordinate metabolism reprogramming, cell death, and differentiation balance, and also synthesis of heme, the prosthetic group needed in hemoglobin assembly. In mammals, mitochondria subcellular localization and mitochondria interaction with other structures as endoplasmic reticulum and nucleus might be of importance for the removal of the nucleus, that is, the enucleation. Here, we aim to characterize by electron microscopy the changes in ultrastructure of cells over successive stages of human erythroblast differentiation. We focus on mitochondria to gain insights into intracellular localization, ultrastructure, and contact with other organelles. We found that mitochondria are progressively cleared with a significant switch between PolyE and OrthoE stages, acquiring a rounded shape and losing contact sites with both ER (MAM) and nucleus (NAM). We studied intracellular vesicle trafficking and found that endosomes and MVBs, known to be involved in iron traffic and heme synthesis, are increased during BasoE to PolyE transition; autophagic structures such as autophagosomes increase from ProE to OrthoE stages. Finally, consistent with metabolic switch, glycogen accumulation was observed in OrthoE stage.

Cholangiocytes, biliary epithelial cells, are known to spontaneously self-organize into spherical cysts with a central lumen. In this work, we explore a promising biocompatible stereolithographic approach to encapsulate cholangiocytes into geometrically-controlled 3D hydrogel structures to guide them towards the formation of branched tubular networks. We demonstrate that within the appropriate mix of hydrogels, normal rat cholangiocytes can proliferate, migrate and organize into branched tubular structures, form walls consisting of a cell monolayer, transport fluorescent dyes into the luminal space and show markers of epithelial maturation such as primary cilia. The resulting structures have dimensions typically found in the intralobular and intrahepatic bile ducts and are stable for weeks, without any requirement of bulk supporting material, thereby offering total access to the basal side of these biliary epithelial constructs.

Objectives Germ cell tumor (GCT) patients with brain metastases (BM) have a poor prognosis and high risk of treatment failure. Optimal therapies for these patients remain controversial. The aim of this study was to report the outcomes of all GCT patients with BM treated with high-dose chemotherapy (HDCT) in our French expert center for GCT. Methods We carried out a retrospective study of 35 GCT patients with BM who were treated from 2003 to 2019 with HDCT, followed by infusions of autologous peripheral blood hematopoietic stem cells. Results The overall survival at 2 years was 36.9% (95% confidence interval, 19.7-54). The median overall survival was 12 months and the median progression-free survival was 8 months. No variables were associated with better survival in the univariable analysis. Among the 35 patients included in our study, 31 completed HDCT and 4 stopped treatments after mobilization. Eleven patients (11) showed favorable responses (complete, partial, or stable disease) to HDCT and 20 patients died of disease progression (17) or toxicities (3). Among the 11 patients with favorable responses to HDCT, 8 (72.7%) had metachronous BM, mostly isolated. The majority of these patients did not receive local treatment at diagnosis or at relapse. Conclusions Together, our study reveals that GCT patients can experience long-term survival even in the presence of BM. Metachronous BM can also be cured with HDCT even in the absence of local treatment. Biological and radiologic responses to mobilization could be a predictor of favorable responses to HDCT.

In recent years, 3D cell culture models such as spheroid or organoid technologies have known important developments. Many studies have shown that 3D cultures exhibit better biomimetic properties compared to 2D cultures. These properties are important for in-vitro modeling systems, as well as for in-vivo cell therapies and tissue engineering approaches. A reliable use of 3D cellular models still requires standardized protocols with well-controlled and reproducible parameters. To address this challenge, a robust and scaffold-free approach is proposed, which relies on multi-trap acoustic levitation. This technology is successfully applied to Mesenchymal Stem Cells (MSCs) maintained in acoustic levitation over a 24-h period. During the culture, MSCs spontaneously self-organized from cell sheets to cell spheroids with a characteristic time of about 10 h. Each acoustofluidic chip could contain up to 30 spheroids in acoustic levitation and four chips could be ran in parallel, leading to the production of 120 spheroids per experiment. Various biological characterizations showed that the cells inside the spheroids were viable, maintained the expression of their cell surface markers and had a higher differentiation capacity compared to standard 2D culture conditions. These results open the path to long-time cell culture in acoustic levitation of cell sheets or spheroids for any type of cells.

Objective To assess the superiority of adipose tissue-derived stromal vascular fraction (AD-SVF) injection into the fingers vs placebo in reducing hand disability in systemic sclerosis (SSc) patients. Methods We performed a double-blind, multicentre, phase II trial from October 2015 to January 2018 in France. SSc patients with a Cochin Hand Function Scale (CHFS) ≥20/90 were randomized 1:1 to receive injection of AD-SVF or placebo. AD-SVF was obtained using the automated processing Celution 800/CRS system. The placebo was lactated Ringer’s solution. The primary efficacy end point was the change of the CHFS score from baseline to 3 months. Secondary efficacy endpoints included the CHFS score at 6 months, hand function, vasculopathy, hand pain, skin fibrosis, sensitivity of the finger pulps, Scleroderma Health Assessment Questionnaire, patients and physician satisfaction, and safety. Results Forty patients were randomized. The AD-SVF and placebo groups were comparable for age, sex ratio, disease duration, skin fibrosis of the hands and main cause of hand disability. After 3 months’ follow-up, hand function significantly improved in both groups with no between-group difference of CHFS (mean change of −9.2 [12.2] in the AD-SVF group vs −7.6 [13.2] in the placebo group). At 6 months, hand function improved in both groups. Conclusion This study showed an improvement of hand function in both groups over time, with no superiority of the AD-SVF. Considering the limits of this trial, studies on a larger population of patients with homogeneous phenotype and hand handicap should be encouraged to accurately assess the benefit of AD-SVF therapy. Trial registration ClinicalTrials.gov, https://clinicaltrials.gov, NCT02558543. Registered on September 24, 2015.

International audience

International audience; In recent years, 3D cell culture models such as spheroid or organoid technologies have known important developments. Many studies have shown that 3D cultures exhibit better biomimetic properties compared to 2D cultures. These properties are important for in-vitro modeling systems, as well as for in-vivo cell therapies and tissue engineering approaches. A reliable use of 3D cellular models still requires standardized protocols with well-controlled and reproducible parameters. To address this challenge, a robust and scaffold-free approach is proposed, which relies on multi-trap acoustic levitation. This technology is successfully applied to Mesenchymal Stromal Cells (MSCs) maintained in acoustic levitation over a 24-hour period. During the culture, MSCs spontaneously self-organized from cell sheets to cell spheroids with a characteristic time of about ten hours. Each acoustofluidic chip could contain up to 30 spheroids in acoustic levitation and four chips could be ran in parallel, leading to the production of 120 spheroids per experiment. Various biological characterizations showed that the cells inside the spheroids were viable, maintained the expression of their cell surface markers and had a higher differentiation capacity compared to standard 2D culture conditions. These results open the path to long-time cell culture in acoustic levitation of cell sheets or spheroids for any type of cells.; In recent years, 3D cell culture models such as spheroid or organoid technologies have known important developments. Many studies have shown that 3D cultures exhibit better biomimetic properties compared to 2D cultures. These properties are important for in-vitro modeling systems, as well as for in-vivo cell therapies and tissue engineering approaches. A reliable use of 3D cellular models still requires standardized protocols with well-controlled and reproducible parameters. To address this challenge, a robust and scaffold-free approach is proposed, which relies on multi-trap acoustic levitation. This technology is successfully applied to Mesenchymal Stromal Cells (MSCs) maintained in acoustic levitation over a 24-hour period. During the culture, MSCs spontaneously self-organized from cell sheets to cell spheroids with a characteristic time of about ten hours. Each acoustofluidic chip could contain up to 30 spheroids in acoustic levitation and four chips could be ran in parallel, leading to the production of 120 spheroids per experiment. Various biological characterizations showed that the cells inside the spheroids were viable, maintained the expression of their cell surface markers and had a higher differentiation capacity compared to standard 2D culture conditions. These results open the path to long-time cell culture in acoustic levitation of cell sheets or spheroids for any type of cells.

Author(s): Delagreverie, Heloise M; Gerard, Laurence; Chaillon, Antoine; Roelens, Marie; Djerroudi, Lounes; Salmona, Maud; Larghero, Jerome; Galicier, Lionel; Simon, Francois; Oksenhendler, Eric; Moins-Teisserenc, Helene; Delaugerre, Constance | Abstract: ObjectivesAutologous stem cell transplantation following high-dose chemotherapy (HDC/ASCT) is the prime model to study the impact of HDC in HIV-1-infected participants. We analyzed the impact of HDC/ASCT on the resurgent reservoir composition and origin.DesignWe included retrospectively a homogenous group of HIV-1-infected patients treated for high-risk lymphoma in a reference center with similar chemotherapy regimens.MethodsThirteen participants treated with HDC/ASCT from 2012 to 2015 were included. A median seven longitudinal blood samples per participant were available. Total HIV-1 DNA levels in peripheral blood mononuclear cells (PBMCs) were quantified by quantitative PCR. In six participants with sustained viral suppression, the highly variable C2V3 viral region was subjected to next-generation sequencing. Maximum-likelihood phylogeny trees were generated from the reconstructed viral haplotypes. Lymphocyte subsets were studied by flow cytometry.ResultsPBMC-associated HIV-1 DNA levels were stable over time. Viral diversity decreased along lymphoma treatment, but increased promptly back to prechemotherapy numbers after HDC/ASCT. Blood viral populations from all time-points were intermingled in phylogeny trees: the resurgent reservoir was similar to pre-HDC circulating proviruses. Memory subsets were the main contributor to the early restoration of the CD4+ T-cell pool, with a delayed increase in naive cell counts.ConclusionsThe characterization of HIV-1 reservoir in blood revealed a fast and consistent replenishment from memory CD4+ T cells after HDC/ASCT. As HDC/ASCT is increasingly involved in HIV cure trials with gene-modified hematopoietic stem cells, the management of infected T cells in HIV-positive autologous transplants will be critical.

Resume Plus de cinq ans apres les premieres publications Americaines rapportant un taux important de reponses cliniques dans des hemopathies lymphoides B de haut grade ou a un stade avance, les equipes Francaises peinent toujours a donner acces a ces traitements. L’un des obstacles – et pas le moindre – reste la mise en place d’une organisation respectant la reglementation Europeenne concernant les medicaments de therapies innovantes, ainsi que les exigences nationales. Nous revoyons ici ces aspects pour les deux scenarios possibles : les CAR-T cells prepares industriellement, et les CAR-T Cells produits comme des medicaments experimentaux.

Tisagenlecleucel therapy has shown promising efficacy for relapsed/refractory (R/R) B-cell precursor acute lymphoblastic leukemia (BCP-ALL). However, relapses occur in 30-50% of patients. Determinants for CD19pos versus CD19neg relapses are poorly characterized. We report on 51 patients with R/R BCP-ALL (median age 17 years) infused with tisagenlecleucel after lymphodepletion. Complete remission rate at D28 was 96%. Prior blinatumomab increased the risk of early failure at D28. The 18-month cumulative incidence of relapse (CIR), event-free survival (EFS), and overall survival (OS) were 51%, 44%, and 74%, respectively, at a median follow-up of 15.5 months. Factors associated with a high tumor burden (occurrence of cytokine release syndrome) and prior blinatumomab were associated with an increased CIR, and a shorter EFS and OS. Pre-lymphodepletion high disease burden (MRD ≥ 10-2, SHR 10.4, p = 0.03) and detectable MRD at D28 (SHR 7.2, p = 0.006) correlated with an increased risk of CD19neg relapse. Low disease burden (SHR 5.3, p = 0.03) and loss of B-cell aplasia (BCA) (SHR 21.7, p = 0.004) predicted an increased risk of CD19pos relapses. These data highlight the impact of prior therapy on patient outcome. Finally, detectable MRD at D28 and loss of BCA both define patients at high risk of relapse for whom additional interventions are needed.

CAR-T cells belong to a new class of biological medicines, referred to as Advanced Therapy Medicinal Products (ATMPs). Despite the cellular component, according to the regulatory definition, CAR-T cells are gene therapy medicines, a sub-category of ATMPs, since their therapeutic effect is the result of their genetic modification. The specificity and the complexity of these innovative drugs have required a complete reorganization of the hospital and pharmaceutical circuits, from the cell collection to the drug administration to the patient. Indeed, increased interaction and collaboration between different healthcare professionals is essential in order to guarantee the quality and safety of these innovative medicines.

This article is distributed under the terms of an Attribution–Noncommercial–Share Alike–No Mirror Sites license for the first six months after the publication date (see http://www.rupress.org/terms/). After six months it is available under a Creative Commons License (Attribution–Noncommercial–Share Alike 4.0 International license, as described at https://creativecommons.org/licenses/by-nc-sa/4.0/).; International audience; The fate of hematopoietic stem and progenitor cells (HSPCs) is regulated by their interaction with stromal cells in the bone marrow. However, the cellular mechanisms regulating HSPC interaction with these cells and their potential impact on HSPC polarity are still poorly understood. Here we evaluated the impact of cell–cell contacts with osteoblasts or endothelial cells on the polarity of HSPC. We found that an HSPC can form a discrete contact site that leads to the extensive polarization of its cytoskeleton architecture. Notably, the centrosome was located in proximity to the contact site. The capacity of HSPCs to polarize in contact with stromal cells of the bone marrow appeared to be specific, as it was not observed in primary lymphoid or myeloid cells or in HSPCs in contact with skin fibroblasts. The receptors ICAM, VCAM, and SDF1 were identified in the polarizing contact. Only SDF1 was independently capable of inducing the polarization of the centrosome–microtubule network.

Changes in gene expression due to cell heterogeneity and response to stressors in specific microenvironments remains poorly understood. Current methodologies that seek to interrogate gene expression at a molecular level require sampling of cellular transcriptome and therefore lysis of the cell preventing serial analysis of cellular transcriptome. To address this area of unmet need, we have recently developed a new technology allowing transcriptomic analysis over time without cellular destruction. Our novel method, TRACE-seq (TRanscriptomic Analysis Captured in Extracellular vesicles using sequencing), is characterized by a cell-type specific transgene expression. It provides data on a representative part of the cell transcriptome inside extracellular vesicles. Thus, the transcriptome of cells expressing TRACE can be followed over time in a non-destructive manner, which is a powerful tool for many fields of fundamental and translational biology research.

Background Tissue engineering is an attractive alternative to conventional esophageal replacement techniques using intra-abdominal organs which are associated with a substantial morbidity. The objective was to evaluate the feasibility of esophageal replacement by an allogenic decellularized esophagus in a porcine model. Secondary objectives were to evaluate the benefit of decellularized esophagus recellularization with autologous bone marrow mesenchymal stromal cells and omental maturation of the decellularized esophagus. Methods Eighteen pigs divided into 4 experimental groups according to mesenchymal stromal cells recellularization and omental maturation underwent a 5-cm long circumferential replacement of the thoracic esophagus. Turbo green florescent protein labelling was used for in vivo mesenchymal stromal cells tracking. The graft area was covered by a stent for 3 months. Clinical and histologic outcomes were analyzed over a 6-month period. Results The median follow-up was 112 days [5; 205]. Two animals died during the first postoperative month, 2 experienced an anastomotic leakage, 13 experienced a graft area stenosis following stent migration of which 3 were sacrificed as initially planned after successful endoscopic treatment. The stent could be removed in 2 animals: the graft area showed a continuous mucosa without stenosis. After 3 months, the graft area showed a tissue specific regeneration with a mature epithelium and muscular cells. Clinical and histologic results were similar across experimental groups. Conclusion Circumferential esophageal replacement by a decellularized esophagus was feasible and allowed tissue remodeling toward an esophageal phenotype. We could not demonstrate any benefit provided by the omental maturation of the decellularized esophagus nor its recellularization with mesenchymal stromal cells.

Background Mesenchymal stem/stromal cells (MSC) have immunomodulatory properties, studied in a wide range of diseases. Validated quality controls must confirm this activity in the context of clinical trials. This study presents a method’s validation, assessing MSC’s ability to inhibit lymphocyte proliferation, according to the ICH Q2 standard. Methods MSC were co-cultured with CellTrace™ Violet-labeled Peripheral blood mononuclear cells (PBMC) coming from a bank of ten donors, at seven different ratios for 7 days. Cell trace violet PBMC bank was validated in parallel. Flow cytometry analysis was used to obtain the division percentage of T cells. The percentage of inhibition of lymphocyte proliferation by MSC, for each ratio X, was calculated using the formula: Ratio × percentage of inhibition = (control percentage of division—ratio × percentage of division)/control percentage of division. The inhibition percentage of lymphocyte proliferation function of co-culture ratios was represented in a line graph. The corresponding area under the curve was calculated, representing MSC’s ability to inhibit lymphocyte proliferation. Results Two cell trace violet PBMC banks were compared for bank validation. When compared using four different MSC samples coming each from a different donor, their area under the curve did not show any statistical differences and were correlated. Moreover, the stability of one cell trace violet PBMC bank was confirmed up to 509 days of storage. Analytical parameters were investigated for method validation. Analysis of repeatability and reproducibility respectively showed a standard deviation of 6.1% and 4.6%. The assay was robust regarding PBMC, as no statistical differences were found between inhibitory activities when testing three adjacent concentrations of PBMC. Still, attention is needed on MSC quantity as it can influence results. Linearity was evaluated: the percentage of inhibition of lymphocyte proliferation function of co-culture ratios was linear on the exploited range. Finally, the assay measurement range allowed to differentiate MSC presenting different inhibition activities. Conclusion This quantification method displayed low analytical variability and no inter-bank variability of PBMC. However, MSC quantification should be checked before co-culture to reduce variability. Therefore, it could be used for the qualification of MSC batches’ immunomodulatory activity.

International audience; Myelomeningocele (MMC) is a spinal cord congenital defect that leads to paraplegia, bladder incontinence and bowel dysfunction. A randomized human trial demonstrated that in utero surgical repair of the MMC defect improves lower limb motor function. However, functional recovery remains incomplete. Stem cell therapy has recently generated great interest in the field of prenatal repair of MMC. In this systematic review we attempt to provide an overview of the current application of stem cells in different animal models of MMC. Publications were retrieved from PubMed and Cochrane Library databases. This process yielded twenty-two studies for inclusion in this review, experimenting five different types of stem cells: human embryonic stem cells, neural stem cells, induced pluripotent stem cells, human amniotic fluid stem cells, and mesenchymal stem cells (MSCs). Rodents and ovine were the two major species used for animal model studies. The source, the aims, and the main results were analyzed. Stem cell therapy appears to be a promising candidate for prenatal repair of MMC, especially MSCs. Further explorations in ovine and rodent models, reporting clinical and functional results, are necessary before an application in humans.