Repair of Acute Respiratory Distress Syndrome in COVID-19 by Stromal Cells (REALIST-COVID Trial): A Multicenter, Randomized, Controlled Clinical Trial.

Rationale: Mesenchymal stromal cells (MSCs) may modulate inflammation, promoting repair in coronavirus disease (COVID-19)-related acute respiratory distress syndrome (ARDS). Objectives: We investigated the safety and efficacy of ORBCEL-C (CD362 [cluster of differentiation 362]-enriched, umbilical cord-derived MSCs) in COVID-19-related ARDS. Methods: In this multicenter, randomized, double-blind, allocation-concealed, placebo-controlled trial (NCT03042143), patients with moderate to severe COVID-19-related ARDS were randomized to receive ORBCEL-C (400 million cells) or placebo (Plasma-Lyte 148). The primary safety and efficacy outcomes were the incidence of serious adverse events and oxygenation index at Day 7, respectively. Secondary outcomes included respiratory compliance, driving pressure, PaO2:FiO2 ratio, and Sequential Organ Failure Assessment score. Clinical outcomes relating to duration of ventilation, lengths of ICU and hospital stays, and mortality were collected. Long-term follow-up included diagnosis of interstitial lung disease at 1 year and significant medical events and mortality at 2 years. Transcriptomic analysis was performed on whole blood at Days 0, 4, and 7. Measurements and Main Results: Sixty participants were recruited (final analysis: n = 30 received ORBCEL-C, n = 29 received placebo; 1 participant in the placebo group withdrew consent). Six serious adverse events occurred in the ORBCEL-C group and three in the placebo group (risk ratio, 2.9 [95% confidence interval, 0.6-13.2]; P = 0.25). Day 7 mean (SD) oxygenation index did not differ (ORBCEL-C, 98.3 [57.2] cm H2O/kPa; placebo, 96.6 [67.3] cm H2O/kPa). There were no differences in secondary surrogate outcomes or in mortality at Day 28, Day 90, 1 year, or 2 years. There was no difference in the prevalence of interstitial lung disease at 1 year or significant medical events up to 2 years. ORBCEL-C modulated the peripheral blood transcriptome. Conclusion: ORBCEL-C MSCs were safe in subjects with moderate to severe COVID-19-related ARDS but did not improve surrogates of pulmonary organ dysfunction.

Repair of acute respiratory distress syndrome by stromal cell administration (REALIST): a structured study protocol for an open-label dose-escalation phase 1 trial followed by a randomised, triple-blind, allocation concealed, placebo-controlled phase 2 trial.

BACKGROUND: Mesenchymal stromal cells (MSCs) may be of benefit in ARDS due to immunomodulatory and reparative properties. This trial investigates a novel CD362 enriched umbilical cord derived MSC product (REALIST ORBCEL-C), produced to Good Manufacturing Practice standards, in patients with moderate to severe ARDS due to COVID-19 and ARDS due to other causes. METHODS: Phase 1 is a multicentre open-label dose-escalation pilot trial. Patients will receive a single infusion of REALIST ORBCEL-C (100 × 106 cells, 200 × 106 cells or 400 × 106 cells) in a 3 + 3 design. Phase 2 is a multicentre randomised, triple blind, allocation concealed placebo-controlled trial. Two cohorts of patients, with ARDS due to COVID-19 or ARDS due to other causes, will be recruited and randomised 1:1 to receive either a single infusion of REALIST ORBCEL-C (400 × 106 cells or maximal tolerated dose in phase 1) or placebo. Planned recruitment to each cohort is 60 patients. The primary safety outcome is the incidence of serious adverse events. The primary efficacy outcome is oxygenation index at day 7. The trial will be reported according to the Consolidated Standards for Reporting Trials (CONSORT 2010) statement. DISCUSSION: The development and manufacture of an advanced therapy medicinal product to Good Manufacturing Practice standards within NHS infrastructure are discussed, including challenges encountered during the early stages of trial set up. The rationale to include a separate cohort of patients with ARDS due to COVID-19 in phase 2 of the trial is outlined. TRIAL REGISTRATION: ClinicalTrials.gov NCT03042143. Registered on 3 February 2017. EudraCT Number 2017-000584-33.

Repair of acute respiratory distress syndrome by stromal cell administration (REALIST) trial: A phase 1 trial.

BACKGROUND: Mesenchymal stromal cells (MSCs) may be of benefit in acute respiratory distress syndrome (ARDS) due to immunomodulatory, reparative, and antimicrobial actions. ORBCEL-C is a population of CD362 enriched umbilical cord-derived MSCs. The REALIST phase 1 trial investigated the safety and feasibility of ORBCEL-C in patients with moderate to severe ARDS. METHODS: REALIST phase 1 was an open label, dose escalation trial in which cohorts of mechanically ventilated patients with moderate to severe ARDS received increasing doses (100, 200 or 400 × 106 cells) of a single intravenous infusion of ORBCEL-C in a 3 + 3 design. The primary safety outcome was the incidence of serious adverse events. Dose limiting toxicity was defined as a serious adverse reaction within seven days. Trial registration clinicaltrials.gov NCT03042143. FINDINGS: Nine patients were recruited between the 7th January 2019 and 14th January 2020. Study drug administration was well tolerated and no dose limiting toxicity was reported in any of the three cohorts. Eight adverse events were reported for four patients. Pyrexia within 24 h of study drug administration was reported in two patients as pre-specified adverse events. A further two adverse events (non-sustained ventricular tachycardia and deranged liver enzymes), were reported as adverse reactions. Four serious adverse events were reported (colonic perforation, gastric perforation, bradycardia and myocarditis) but none were deemed related to administration of ORBCEL-C. At day 28 no patients had died in cohort one (100 × 106), three patients had died in cohort two (200 × 106) and one patient had died in cohort three (400 × 106). Overall day 28 mortality was 44% (n = 4/9). INTERPRETATION: A single intravenous infusion of ORBCEL-C was well tolerated in patients with moderate to severe ARDS. No dose limiting toxicity was reported up to 400 × 106 cells.

Repair of Acute Respiratory Distress Syndrome by Stromal Cell Administration in COVID-19 (REALIST-COVID-19): A structured summary of a study protocol for a randomised, controlled trial.

OBJECTIVES: The primary objective of the study is to assess the safety of a single intravenous infusion of Mesenchymal Stromal Cells (MSCs) in patients with Acute Respiratory Distress Syndrome (ARDS) due to COVID-19. Secondary objectives are to determine the effects of MSCs on important clinical outcomes, as described below. TRIAL DESIGN: REALIST COVID 19 is a randomised, placebo-controlled, triple blinded trial. PARTICIPANTS: The study will be conducted in Intensive Care Units in hospitals across the United Kingdom. Patients with moderate to severe ARDS as defined by the Berlin definition, receiving invasive mechanical ventilation and with a diagnosis of COVID-19 based on clinical diagnosis or PCR test will be eligible. Patients will be excluded for the following reasons: more than 72 hours from the onset of ARDS; age < 16 years; patient known to be pregnant; major trauma in previous 5 days; presence of any active malignancy (other than non-melanoma skin cancer); WHO Class III or IV pulmonary hypertension; venous thromboembolism currently receiving anti-coagulation or within the past 3 months; patient receiving extracorporeal life support; severe chronic liver disease (Child-Pugh > 12); Do Not Attempt Resuscitation order in place; treatment withdrawal imminent within 24 hours; prisoners; declined consent; non-English speaking patients or those who do not adequately understand verbal or written information unless an interpreter is available; previously enrolled in the REALIST trial. INTERVENTION AND COMPARATOR: Intervention: Allogeneic donor CD362 enriched human umbilical cord derived mesenchymal stromal cells (REALIST ORBCEL-C) supplied as sterile, single-use cryopreserved cell suspension of a fixed dose of 400 x106 cells in 40ml volume, to be diluted in Plasma-Lyte 148 to a total volume of 200mls for administration. Comparator (placebo): Plasma-Lyte 148 Solution for Infusion (200mls). The cellular product (REALIST ORBCEL-C) was developed and patented by Orbsen Therapeutics. MAIN OUTCOMES: The primary safety outcome is the incidence of serious adverse events. The primary efficacy outcome is Oxygenation Index (OI) at day 7. Secondary outcomes include: OI at days 4 and 14; respiratory compliance, driving pressure and PaO2/FiO2 ratio (PF ratio) at days 4, 7 and 14; Sequential Organ Failure Assessment (SOFA) score at days 4, 7 and 14; extubation and reintubation; ventilation free days at day 28; duration of mechanical ventilation; length of ICU and hospital stay; 28-day and 90-day mortality. RANDOMISATION: After obtaining informed consent, patients will be randomised via a centralised automated 24-hour telephone or web-based randomisation system (CHaRT, Centre for Healthcare Randomised Trials, University of Aberdeen). Randomisation will be stratified by recruitment centre and by vasopressor use and patients will be allocated to REALIST ORBCEL-C or placebo control in a 1:1 ratio. BLINDING (MASKING): The investigator, treating physician, other members of the site research team and participants will be blinded. The cell therapy facility and clinical trials pharmacist will be unblinded to facilitate intervention and placebo preparation. The unblinded individuals will keep the treatment information confidential. The infusion bag will be masked at the time of preparation and will be administered via a masked infusion set. NUMBERS TO BE RANDOMISED (SAMPLE SIZE): A sample size of 60 patients with 30 patients randomised to the intervention and 30 to the control group. If possible, recruitment will continue beyond 60 patients to provide more accurate and definitive trial results. The total number of patients recruited will depend on the pandemic and be guided by the data monitoring and ethics committee (DMEC). TRIAL STATUS: REALIST Phase 1 completed in January 2020 prior to the COVID-19 pandemic. This was an open label dose escalation study of REALIST ORBCEL-C in patients with ARDS. The COVID-19 pandemic emerged as REALIST Phase 2 was planned to commence and the investigator team decided to repurpose the Phase 2 trial as a COVID-19 specific trial. This decision was discussed and approved by the Trial Steering Committee (TSC) and DMEC. Submissions were made to the Research Ethics Committee (REC) and MHRA to amend the protocol to a COVID-19 specific patient population and the protocol amendment was accepted by the REC on 27th March 2020 and MHRA on 30th March 2020 respectively. Other protocol changes in this amendment included an increase in the time of onset of ARDS from 48 to 72 hours, inclusion of clinical outcomes as secondary outcomes, the provision of an option for telephone consent, an indicative sample size and provision to continue recruitment beyond this indicative sample size. The current protocol in use is version 4.0 23.03.2020 (Additional file 1). Urgent Public Health status was awarded by the NIHR on 2 April 2020 and the trial opened to recruitment and recruited the first participant the same day. At the time of publication the trial was open to recruitment at 5 sites across the UK (Belfast Health and Social Care Trust, King's College London, Guys and St Thomas' Hospital London, Birmingham Heartlands Hospital and the Queen Elizabeth Hospital Birmingham) and 12 patients have been recruited across these sites. Additional sites are planned to open and appropriate approvals for these are being obtained. It is estimated recruitment will continue for 6 months. TRIAL REGISTRATION: ClinicalTrials.gov NCT03042143 (Registered 3 Feb 2017). EudraCT 2017-000585-33 (Registered 28 Nov 2017). FULL PROTOCOL: The full protocol (version 4.0 23.03.2020) is attached as an additional file, accessible from the Trials website (Additional file 1). In the interest of expediting dissemination of this material, the familiar formatting has been eliminated; this Letter serves as a summary of the key elements of the full protocol. The study protocol has been reported in accordance with the Standard Protocol Items: Recommendations for Clinical Interventional Trials (SPIRIT) guidelines (Additional file 2).

Liver homing of clinical grade Tregs after therapeutic infusion in patients with autoimmune hepatitis.

Autoimmune hepatitis (AIH) is an immune-mediated disease with no curative treatment. Regulatory T cell (Treg) therapy is potentially curative in AIH given the critical role of Tregs in preventing autoimmunity. To work effectively, adoptively transferred Tregs must migrate to and survive within the inflamed liver. We conducted a proof-of-concept study aiming to assess the safety and liver-homing properties of good manufacturing practice (GMP)-grade autologous Tregs in patients with AIH. METHODS: Autologous polyclonal GMP-grade Tregs were isolated using leukapheresis and CliniMACS, labelled with indium tropolonate and re-infused intravenously to 4 patients with AIH. GMP-Treg homing to the liver was investigated with longitudinal gamma camera and SPECT-CT scanning. GMP-Treg immunophenotype, function and immunometabolic state were assessed during the study. RESULTS: We observed that the isolated Tregs were suppressive and expressed CXCR3, a chemokine receptor involved in recruitment into the inflamed liver, as well as Treg functional markers CD39, CTLA-4 and the transcription factor Foxp3. Serial gamma camera and SPECT-CT imaging demonstrated that 22-44% of infused Tregs homed to and were retained in the livers of patients with autoimmune hepatitis for up to 72 h. The infused cells did not localise to any off-target organs other than the spleen and bone marrow. GMP-Tregs were metabolically competent and there were no infusion reactions or high-grade adverse effects after Treg infusion. CONCLUSION: Our novel findings suggest that the liver is a good target organ for Treg cellular therapy, supporting the development of clinical trials to test efficacy in autoimmune hepatitis and other autoimmune liver diseases. LAY SUMMARY: Autoimmune liver diseases occur when the body's immune cells target their own liver cells. Regulatory T cells (Tregs) prevent autoimmunity, thus they are a potential therapy for autoimmune liver diseases. In patients with autoimmune hepatitis, Treg infusion is safe, with nearly a quarter of infused Tregs homing to the liver and suppressing tissue-damaging effector T cells. Thus, Tregs are a potentially curative immune cell therapy for early autoimmune liver diseases.

Granulocyte colony-stimulating factor and autologous CD133-positive stem-cell therapy in liver cirrhosis (REALISTIC): an open-label, randomised, controlled phase 2 trial.

BACKGROUND: Results of small-scale studies have suggested that stem-cell therapy is safe and effective in patients with liver cirrhosis, but no adequately powered randomised controlled trials have been done. We assessed the safety and efficacy of granulocyte colony-stimulating factor (G-CSF) and haemopoietic stem-cell infusions in patients with liver cirrhosis. METHODS: This multicentre, open-label, randomised, controlled phase 2 trial was done in three UK hospitals and recruited patients with compensated liver cirrhosis and MELD scores of 11·0-15·5. Patients were randomly assigned (1:1:1) to receive standard care (control), treatment with subcutaneous G-CSF (lenograstim) 15 µg/kg for 5 days, or treatment with G-CSF for 5 days followed by leukapheresis and intravenous infusion of three doses of CD133-positive haemopoietic stem cells (0·2 × 106 cells per kg per infusion). Randomisation was done by Cancer Research UK Clinical Trials Unit staff with a minimisation algorithm that stratified by trial site and cause of liver disease. The coprimary outcomes were improvement in severity of liver disease (change in MELD) at 3 months and the trend of change in MELD score over time. Analyses were done in the modified intention-to-treat population, which included all patients who received at least one day of treatment. Safety was assessed on the basis of the treatment received. This trial was registered at Current Controlled Trials on Nov 18, 2009; ISRCTN, number 91288089; and the European Clinical Trials Database, number 2009-010335-41. FINDINGS: Between May 18, 2010, and Feb 26, 2015, 27 patients were randomly assigned to the standard care, 26 to the G-CSF group, and 28 to the G-CSF plus stem-cell infusion group. Median change in MELD from day 0 to 90 was -0·5 (IQR -1·5 to 1·1) in the standard care group, -0·5 (-1·7 to 0·5) in the G-CSF group, and -0·5 (-1·3 to 1·0) in the G-CSF plus stem-cell infusion group. We found no evidence of differences between the treatment groups and control group in the trends of MELD change over time (p=0·55 for the G-CSF group vs standard care and p=0·75 for the G-CSF plus stem-cell infusion group vs standard care). Serious adverse events were more frequent the in G-CSF and stem-cell infusion group (12 [43%] patients) than in the G-CSF (three [11%] patients) and standard care (three [12%] patients) groups. The most common serious adverse events were ascites (two patients in the G-CSF group and two patients in the G-CSF plus stem-cell infusion group, one of whom was admitted to hospital with ascites twice), sepsis (four patients in the G-CSF plus stem-cell infusion group), and encephalopathy (three patients in the G-CSF plus stem-cell infusion group, one of whom was admitted to hospital with encephalopathy twice). Three patients died, including one in the standard care group (variceal bleed) and two in the G-CSF and stem-cell infusion group (one myocardial infarction and one progressive liver disease). INTERPRETATION: G-CSF with or without haemopoietic stem-cell infusion did not improve liver dysfunction or fibrosis and might be associated with increased frequency of adverse events compared with standard care. FUNDING: National Institute of Health Research, The Sir Jules Thorn Charitable Trust.

Giardia genotypes in pigs in Western Australia: prevalence and association with diarrhea.

Little is known of the prevalence of Giardia species and genotypes in pre- and post-weaned domestic pigs. In the present study, a total of 297 pig faecal samples were screened for the presence of Giardia by PCR and genotyped. An overall prevalence of 31.1% (90/289) (25.8, 36.5 CI) was detected. Giardia was detected in 17% (23/123) (11.8-25.6 CI) of pre-weaned piglet faecal samples and 41% (64/156) (33.3-48.7 CI) post-weaned faecal samples analysed. Sequence analysis identified assemblage A and E in pre- and post-weaned pigs. Assemblage F was identified in one post-weaned pig. Assemblage E was the most prevalent assemblage detected.

Identification of zoonotic Giardia genotypes in marsupials in Australia.

A total of 421 fecal samples from a variety of captive and wild marsupial hosts in Western Australia, Victoria and South Australia were screened for the presence of Giardia species/genotypes using PCR and sequence analysis of a fragment of the 18S rRNA gene. Giardia spp. were identified in 13.4% (28/209) of samples from captive marsupials and 13.7% (29/212) of samples from wild marsupials. Sequence analysis at the 18S locus identified the zoonotic Giardia duodenalis Genotypes A and B in both captive and wild marsupials. Eight isolates were typed as genotype B3 and B4 at the gdh locus, although 7/8 were typed as genotype A at the 18S rRNA locus. The possible reasons for this discordance are discussed. This is the first report of genotype B and only the second report of genotype A in marsupials. As some of the genotype B isolates were identical to human-derived Giardia gdh sequences, these results suggest that marsupials in catchments may pose a public health risk and therefore warrant further investigation.

T-cell depletion with Campath-1H "in the bag" for matched related allogeneic peripheral blood stem cell transplantation is associated with reduced graft-versus-host disease, rapid immune constitution and improved survival.

We studied the outcome of 24 peripheral blood stem cell (PBSC) graft recipients, who were T-cell depleted (TCD) with either 20 mg (n = 14) or 10 mg (n = 10) Campath-1H in vitro, in comparison with a retrospective cohort of 23 unmanipulated (UM) PBSC recipients. While the neutrophil engraftment was similar, the platelet engraftment occurred earlier in the TCD group (d 11 vs 14). The incidence of acute and chronic graft-versus-host-disease (GVHD) was 8.7% and 4.4% in the TCD group, respectively, compared with 47.7% and 56.3% in UM group (P < 0.001). In the TCD group, 5/6 chronic myeloid leukaemia (CML) and 4/18 non-CML patients relapsed (vs 0/6 and 3/17 in UM group, P = 0.06). All four molecular or cytogenetic relapses of CML were disease-free survivors following donor lymphocyte infusion. There was no difference in the incidence of serious infection between the TCD and UM groups and the lymphocyte recovery at 100 d was comparable. In the TCD cohort, the lymphocyte recovery was quicker in the 10 mg Campath-1H group. The non-relapse mortality (19.1%vs 66.3%) and 3 year survival (73.1 vs 19.2) were improved in the TCD group (P = 0.05). Thus elimination of late mortalities related to chronic GVHD and a rapid immune reconstitution, limiting either infection or relapse related deaths, contributed to an improved outcome following T-cell depletion with Campath-1H "in the bag".