Effects of intravascular administration of mesenchymal stromal cells derived from Wharton’s Jelly of the umbilical cord on systemic immunomodulation and neuroinflammation after traumatic brain injury (TRAUMACELL): study protocol for a multicentre randomised controlled trial

Abstract

Introduction

Traumatic brain injury (TBI) is one of the leading causes of death and disability worldwide. Treatments for TBI patients are limited and none has been shown to provide prolonged and long-term neuroprotective or neurorestorative effects. A growing body of evidence suggests a link between TBI-induced neuro-inflammation and neurodegenerative post-traumatic disorders. Consequently, new therapies triggering immunomodulation and promoting neurological recovery are the subject of major research efforts. We hypothesise that repeated intravenous treatment with mesenchymal stromal cells derived from Wharton’s Jelly of the umbilical cord-derived mesenchymal stromal cells ((WJ-UC-MSC) may be associated with a significant decrease of post-TBI neuroinflammation and improvement of neurological status.

Methods and analysis

The TRAUMACELL trial is a prospective, national multicentre, phase III, superiority, double-arm comparative randomised (1:1) double-blinded clinical trial. Among patients aged between 18–50, with a severe TBI defined by a Glasgow score less than 12 (within the first 48 hours) with brain traumatic lesion on CT Scan and needing intracranial pressure monitoring, with no other significant organ trauma (abbreviated injury scale<2) and unresponsive to verbal commands after 5 days of sedation discontinuation, 68 will be randomly allocated to receive either WJ-UC-MSC solution or placebo, with three intravenous injections 1 week apart. The primary outcome is the [¹⁸F]-DPA-714 signal intensity in corpus callosum measured by dynamic positron emission tomography (PET)-MRI at 6 months after the last injection, blinded to the randomisation arm, to evaluate the post-traumatic neuro-inflammation.

Ethics and dissemination

The TRAUMACELL trial has been approved by an independent ethics committee (CPP SUD EST II) and French Medicines Agency (2023-504415-33-00) for all study centres. Participant recruitment will be starting in September 2024. Results will be published in international peer-reviewed medical journals.

Trial registration number

NCT06146062, first posted 24 November 2023

Protocol version identifier

TRAUMACELL−V.2.0_20240102

Correspondence to Dr Stéphanie Sigaut; [email protected]

STRENGTHS AND LIMITATIONS OF THIS STUDY

• High-quality methodology using randomised controlled trial design.

• Translational approach with bench and bedside data collection.

• Risk of included patients’ death without neuroinflammation imaging evaluation at 6 months.

• Radioligand low binders will be included and randomised leading to patients without imaging results.

Introduction

Background and rationale

Traumatic brain injury (TBI) is one of the leading causes of death and disability worldwide. These patients are burdened by physical, cognitive and psychosocial deficits, leading to an important economic impact on society. Treatments for TBI patients are limited and none has been shown to provide long-term neuroprotective or neurorestorative effects.¹

In TBI, the primary damage occurs at the time of impact, then secondary brain insults arise through systemic or intracranial mechanisms that reduce cerebral blood flow. Later, tertiary long-term post-traumatic evolution is characterised by cerebral atrophy and white matter structural defects. An increasing number of experimental and clinical data show that the potential entry point in this third brain damage process is neuroinflammation.^2–4 Indeed, microglial cells, the phagocytic cells of the central nervous system, are activated following brain insult, migrate towards the lesion and release both neurotrophic and neurotoxic factors. This activation can last many years after TBI⁴ and thus contributes to the inflammatory process becoming chronic via epigenetic mechanisms. It has been demonstrated that persisting TBI-induced neuroinflammation is associated with poor outcomes⁵ and a growing body of evidence suggests a link between neuroinflammation and post-traumatic neurodegenerative disorders.⁶ Thus, the determinants of late neuroinflammation and their relation to the development of progressive chronic pathologies are key questions, since modulation of this process offers a promising therapeutic window for interventions aimed at improving the prognosis of TBI. Consequently, new therapies triggering immunomodulation and promoting neurological recovery are the subject of major research efforts. We assume that specific therapies to prevent the formation and worsening of tertiary lesions via immunomodulation during the acute phase, to limit brain damage and potential neurodegeneration, will improve the quality of life of patients.

In this context, mesenchymal cell-based therapies are currently investigated to treat various neurological disorders due to their ability to modulate neuroinflammation and to promote simultaneous neurogenesis, angiogenesis and neuroprotection. Indeed, several experimental studies have reported that human umbilical cord-derived mesenchymal stromal cells (MSC) have the ability to improve neurological outcomes and recovery in cerebral injury animal models, including TBI. Although MSC can be harvested from various tissue sources, there is compelling evidence that those from the Wharton’s jelly of the umbilical cord (WJ-UC-MSC) are endowed with the most robust anti-inflammatory and immunomodulatory properties.⁷ Furthermore, the composition of their secretome particularly qualifies them for repairing brain tissue by comparison with their counterparts derived from bone marrow or fat, because they preferentially secrete factors involved in neuroprotection, neurogenesis and angiogenesis.⁸ The ease of their procurement as well as their high proliferation potential, enabling the constitution of large-scale cell banks, further strengthen their attractivity.

Several experimental studies have reported that these cells have the ability to improve neurological outcomes and recovery in cerebral injury animal models, including TBI.^{9 10} In the clinics, WJ-UC-MSC have been tested in different diseases with a good safety record.^11–13 In the field of traumatic brain pathology, several phase I and II clinical studies evaluating the relevance and safety of MSC (mostly autologous bone marrow mononuclear cells) have reported encouraging results,^14–20 but intravenous injections of WJ-UC-MSC have yet to be investigated.

Aims and objectives

The primary aim of the TRAUMACELL study is to determine whether the iterative intravenous injections of WJ-UC-MSC, as compared with a placebo, reduce the post-traumatic neuroinflammation in corpus callosum by positron emission tomography (PET)–MRI (PET-MRI) at 6 months, in severe traumatic brain-injured patients unresponsive to simple verbal commands after 5 days of sedation discontinuation. We hypothesise that WJ-UC-MSC injected weekly for 3 weeks, starting maximum of 10 days after discontinuation of sedation, could decrease the neuroinflammation in corpus callosum (volume of interest, VOI) measured by dynamic PET-MRI at 6 months after the last injection. The PET-MRI uptake will be quantified using the (¹⁸F)-DPA-714 standard uptake value ratio (SUVr) by an investigator who is blinded to the randomisation arm.

The secondary aims of this trial are presented in table 1.

Table 1

Secondary outcomes and associated endpoints

Secondary objectives	Secondary outcomes
To evaluate the effect of iterative IV injections of WJ-UC-MSC at 6 months after the last injection on post-traumatic neuroinflammation measured by volume of interest approach (VOI) in other regions of the brain that is, pericontusional, grey and white matter, frontal, parietal, occipital, hippocampus, thalamus, mesencephalus, cerebellum, and global(18F)-DPA-714 signal intensity	(18F)-DPA-714 standard uptake value ratio (SUVr) in pericontusional, grey and white matter, frontal, parietal, occipital, hippocampus, thalamus, mesencephalus and cerebellum (volume of interest, VOI) measured by dynamic PET-MRI (90 min) at 6 months after the last injection and will be quantified blinded to the randomisation arm (same simultaneous PET and MR acquisitions as for the primary endpoint)
To evaluate the effect of iterative IV injections of WJ-UC-MSC at 6 months after the last injection on the radiological markers from PET-MRI	The regional fractional anisotropy (FA) and the mean diffusibility (MD) from DTI acquisition of PET-MRI (90 min) at 6 months after the last injection and quantified blindly to the randomisation arm (same simultaneous PET and MR acquisitions for the primary endpoint)
To evaluate the effect of iterative IV injections of WJ-UC-MSC at 6 months and at 1 year after the last injection on neurological assessment scales	Functional neurological outcome, assessed via two clinical evaluations, between 6 and 8 months and between 12 and 14 months after the last injection, done for all patients by a dedicated rehabilitation physician, including the Glasgow Outcome Scale-Extended for functional outcome and the MOCA scale for cognitive assessment
To evaluate the feasibility of iterative IV injections of MSC in severe traumatic brain-injured patients	Feasibility, evaluated by the number of treatments administered to the patient compared with the number of treatments planned
To evaluate the tolerability of iterative IV injections of MSC during injections, at short term, and at long term (6 months)	Short term tolerability, evaluated via: * Common Terminology Criteria for Adverse Events (CTCAE) and WHO toxicity scale, during injections and the 10 following days. Unacceptable toxicity is defined as a grade 3 or greater; * Full blood count, electrolyte test with creatinin, hepatic assessment, 24-hour urine protein test, donor-specific antibody testing within 48 hours after each injection. Long-term tolerability, evaluated via CTCAE and donor-specific antibody testing between 6 and 8 months after last injection
To explore biomarkers predictive of iterative IV injections of MSC' success on neuroinflammation (at 6 months after the last injection) and neurological outcome (at 6 months and at 1 year after the last injection)	Identification of biomarkers predictive of a good response, via analysis of immunomodulation at D1, D14, and M6, through: Cytokine profile Primary peripheral blood mononuclear cells (PBMC) profile Transcriptomics and regulatory epigenomics of circulating monocytes and lymphocytes at a single cell level Genome-wide single-nucleotide polymorphism (SNP) genotype
To analyse mechanisms of action, pharmacokinetics, and pharmacodynamics of WJ-UC-MSC in humans, to identify phenotypes involved in the immunomodulation and alloimmunisation induced by MSC administration	Evaluation of the early pharmacokinetics and impact of WJ-UC-MSC administration, on five blood samples done before the first injection, 48 hours (±1 day) after first, second and third injections, 72 hours (±1 day) after third injection, for a subgroup of 12 patients in each arm, by: A NGS and digital droplets (dd)-PCR approach Deep phenotyping of the main immune effector cell populations, performed on whole blood by flow cytometry. Cytokine and chemokine levels in plasma, measured with Luminex magnetic beads technology, using a 96-well Bio-plex Pro Human Cytokine 17-plex assay kit (Biorad laboratories). This is an ancillary study.

MOCA, Montréal Cognitive Assessment; PET, positron emission tomography; WHO, World Health Organization; WJ-UC-MSC, Wharton’s Jelly of the umbilical cord-derived mesenchymal stromal cells.

Methods and analysis

Design overview

The TRAUMACELL study is an investigator-initiated, national multicentre, phase III, superiority, parallel-group, double-blinded, comparative randomised controlled trial (RCT), in severe isolated TBI adult patients (defined as: Glasgow Coma Scale <12 within the 48 first hours+traumatic brain lesion on CT scan+need for intracranial pressure monitoring). Patients unresponsive to verbal commands 5 days after discontinuation of sedation, and for whom, after the usual clinical and paraclinical evaluation (including a quantitative MRI (qMRI), which will be used as baseline imaging for our study), there has been no decision to interrupt active treatment within 10 days of discontinuing sedation, will be allocated in a 1:1 ratio to WJ-UC-MSC (intervention group) or to placebo (control group). We report the study protocol according to the Standard Protocol Items: Recommendations for interventional Trials statement.²¹

Before inclusion and randomisation, a screening will be performed by the investigator or by a medical doctor representing the investigator to check the eligibility criteria at day 5 after discontinuation of sedation and during the next 5 days. If during this period and after the usual clinical evaluation and investigations, no decision to discontinue active treatment is documented, the patient will be included and simultaneously randomised after written informed consent is obtained from the patient’s next of kin. After randomisation (performed by the investigator or by a medical doctor representing the investigator) and before the first injection in both groups, blood samples will be performed for the predictive biomarkers objective, the ancillary study objective and also two specific assessments: donor-specific antigens (DSA) measurements for future allo-immunisation diagnostic and translocator protein (TSPO) rs6971 polymorphism genotype for identification of high, mixed and low-affinity binders (HAB, MAB and LAB).

WJ-UC-MSC suspension (2.10⁶ cells/kg in a NaCl 0.9%/Albumin 0.5% solution) or placebo (NaCl 0.9% solution) will be administered to the patients after the randomisation and the first blood collection. The experimental treatment and placebo injection will be repeated three times, 1 week apart. Blood collection will be repeated at the second and the third injections.

Neurointensive care unit management will follow the guidelines for severe TBI.^{22 23} Medical management and data collection will be identical between the two groups in all other aspects.

Two telephone consultations are scheduled at months 2 and 4 to check on each study participant’s health and to record any unexpected events. If a participant is unable to answer, their relative or doctor will be contacted.

At 6–8 months after the last injection, a PET-MRI will be performed in all randomised patients, except those in whom LAB were identified and who will have MRI only, and all patients (including those with LAB) will be evaluated clinically by a dedicated rehabilitation physician (GOS-E and MOCA scales). Blood sample collection for biobank (third point) (four tubes of 9 mL and one tube of 3 mL) and DSA measurement (one tube of 7 mL) will be collected.

At 12–14 months after the last injection, another clinical evaluation will be performed. A long-term follow-up will be organised in the context of usual care.

The trial design is summarised in table 2 and in figure 1.

Table 2

Timeline of the data collection points

Study period	Screening	Allocation D0	First injection	Second injection	Third injection	Follow-up phone call	PET-MRI first evaluation	Second evaluation
Timepoint	5D after sedation discontinuation	5 to 10D after sedation discontinuation	D0	D0+7D (±2)	D0+14D (±2)	M2 and M4	6 to 8M after the third injection	12 to 14M after the third injection
Enrolment
Eligibility screen	X
Next of kin written informed consent	X
Allocation		X
Pregnancy screening	X						X
Interventions
WJ-UC-MSC			X	X	X
Placebo			X	X	X
Assessment
Baseline variables
Demographics	X
Medical history	X
Clinical examination	X	X
qMRI	X
Clinical evaluation
Vital signs		X	X	X	X		X	X
Adverse events			X	X	X		X	X
Blood sample data*
TPSO screening		X
Safety tests (biochemistry, haematology)		X	X	X	X
Biological collection			X		X		X
Donor-specific antibodies (DSA)					X		X
Ancillary study			X	X	X
Outcome variables
(18F)-DPA-714 SUVr							X
Neurological clinical outcome MOCA and GOSE							X	X
Feasibility					X
CTCAE			X	X	X	X	X
WHO toxicity scale			X	X	X

CTCAE, Common Terminology Criteria for Adverse Events; GOSE, Glasgow outcome scale extended; MOCA, Montréal cognitive assessment; PET, positron emission tomography; SUVr, standard uptake value ratio; TSPO, Translocator protein; WHO, world health organization; WJ-UC-MSC, Wharton’s Jelly of the umbilical cord-derived mesenchymal stromal cells.

Figure 1. RCT flow diagram. D: day; DSA: Donnor Specific Antigen; M: month; MRI: Magnetic Resonance Imaging; PET: Positive emission Tomography; TBI: Traumatic Brain Injury.

Inclusions started on 23 September 2024. The inclusion period will be of 24 months. As participation period (treatment+follow up) is of 14 months, the total duration of the study will be 38 months.

Study setting and population

Participants will be prospectively recruited among patients hospitalised for isolated, severe TBI in three French neuro-intensive care units: Pitié Salpêtrière Hospital Neuro Intensive Care Unit, Beaujon Hospital Intensive Care Unit and Percy Military Hospital Intensive Care Unit.

Patients with isolated, severe TBI defined as: initial Glasgow Coma Scale <12 and presence of traumatic brain lesion on CT and need for ICP monitoring will be screened 5 days after sedation discontinuation if they are unresponsive to commands. Then, if after usual clinical evaluation and investigations (including a qMRI) performed during the 5 subsequent days, no withdrawal of life-sustaining measures (LSM) is made by the clinicians in charge, these patients will be included. These criteria were chosen to select patients that may benefit the most from immunomodulatory therapy: those with disorders of consciousness remaining after the acute phase of increased intracranial pressure and for whom it was decided to continue LSM. This last point is particularly important to check before inclusion to avoid ethical conflicts such as continuing futile LSM because of inclusion.

Considering that patients are not able to sign the informed consent form, the consent of a relative or support person will be required. Their family or proxy will be approached regarding participation in the study by the investigators or their nominated representatives after 5 days or more of discontinued sedation. Patients will be considered eligible for randomisation only if they fulfil all the inclusion criteria and none of the exclusion criteria apply, as defined in box 1.

Box 1

Eligibility criteria

Eligibility criteria

• Unresponsive to verbal commands after 5 days of sedation discontinuation, for whom, after usual clinical and paraclinical evaluation, there has been no decision to withdraw life-sustaining measures within 10 days after sedation discontinuation.

Inclusion criteria

• Demographic criteria: age 18–50,

• Severe TBI defined by:

  • Glasgow score <12 within the 48 first hours.

  • Brain traumatic lesion on CT scan.

  • Need for intracranial pressure monitoring.

• No other significant organ trauma (abbreviated injury scale— <2),

• Written consent signed by the patient’s next of kin.

Exclusion criteria

• History of disease or treatment impairing current or previous year immunity function (haematologic disease (leukaemia, myeloma), viral disease affecting immunity (like HIV), immunological treatment (corticoid, antirejection medication, anti-TNFα (tumor necrosis factor alpha), chemotherapy).

• History of severe neurological or psychiatric disease likely to alter neurological assessment,

• HTAP >grade III WHO.

• Ongoing uncontrolled infection with organ failure (septic shock, ARDS (acute respiratory distress syndrome) including those due to severe COVID-19. Patients with asymptomatic COVID-19 as incidental finding on admission are eligible.

• Platelets <100 000/µL, Hb <8 g/dL, lymphocytes count <1500/µL, neutrophils count <2500/µL, creatinine >100 μmol/L.

• Liver function abnormalities (bilirubin >2.5 mg/dL or transaminases >5× the ULN). Patients with Gilbert’s disease are eligible if liver function tests are normal apart from bilirubinaemia.

• Known HIV seropositivity.

• Neoplasia treated in the 3 years before screening.

• Bone marrow transplant recipient.

• History of transfusion reaction.

• Pregnancy.

• Contraindication for MRI and PET-MRI:

  • MR-incompatible pacemaker and defibrillator.

  • MR-incompatible prosthetic heart valve.

  • Metallic intraocular, intracerebral or intramedullary foreign bodies.

  • Implantable neurostimulation systems.

  • Cochlear implants/ear implant.

  • Metallic fragments such as bullets, shotgun pellets and metal shrapnel.

  • Cerebral artery aneurysm clips.

  • Ventriculoperitoneal shunt with metallic component generating significant artefacts on the MR sequence.

  • Catheters with metallic components (Swan-Ganz catheter).

  • Patient unable to remain supine and motionless during the duration of the examination,

• Participation in another interventional clinical trial of an investigational therapy within 30 days of consent.

• No affiliation to a social security regime.

• Vulnerable person according to article L1121-6 of the French public health code.

• Protected adult person.

Moreover, after inclusion, patient will be checked for TSPO rs6971 polymorphism genotype, to identify LAB. In this population, PET acquisition is compromised and thus these patients (estimated to be 10% of our population)²⁴ will be evaluated at 6 months with MRI and neurocognitive tests.

Interventions

Experimental group

Allogenic MSC derived from Wharton’s Jelly of the umbilical cord and expanded in vitro.

Final product is an MSC suspension (2.10⁶ cells/kg in 150 mL of NaCl 0.9%/human albumin 0.5% solution) conditioned aseptically and identified for intravenous administration three times 1 weeks apart after the randomisation.

MSC-based products are classified as advanced therapy medicinal products (ATMP) according to the European Regulation 1394/2007/EC. The experimental ATMP and the placebo solution will be manufactured by the MEARY Cell and Gene Therapy Center (Hôpital Saint-Louis, AP-HP) according to the Good Manufacturing Practices specific to ATMP (GMP-ATMP). To ensure the consistency of the cell product delivered to the treated patients and overcome the issue of variability in cord-derived MSCs, all the banked cells used in this trial originate from a single cord and thus the same batch.

In the literature, dose-effect analysis shows discording results: if some studies found a relationship between the number of cells injected and neurological recovery,²⁵ others do not. Thus, we choose a dose similar to the one used in most clinical trials.²⁶

In most of the trials, cells have been injected intravenously, which is justified by the presumed dual mechanism of action of MSC, both paracrine (rewiring of circulating and tissue-resident immune-inflammatory cell phenotype towards a reparative pattern) and endocrine (remote effect on the brain of these modified cells as they traffic in the bloodstream). Each injection will be done in 45 min, with continuous and non-invasive monitoring of arterial pressure, cardiac rate and oxygen saturation.

Finally, the ideal therapeutic window for MSC injections in our population is not clearly defined. Late injections (months or even years after TBI) seem to have little effect,^{16 27} and inhibition of immediate posttraumatic inflammatory response with injections in the acute phase does not appear as beneficial.^{28 29} Thus, our protocol plans the injections after the acute phase of increased intracranial pressure, when the patient is stabilised and withdrawal of sedation is ongoing, that is, around 15 days after TBI in most of the cases.

Control group

Placebo suspension (150 mL of NaCl 0.9% solution) prepared aseptically and identified for intravenous administration three times, 1 week apart, after the randomisation.

Outcomes

Primary outcome

The primary outcome is the [¹⁸F]-DPA-714 SUVr in corpus callosum (VOI) measured by dynamic PET-MRI (90 min) at 6 months after the last injection and will be quantified by an investigator who is blinded to the randomisation arm. We chose the [¹⁸F]-DPA-714 signal intensity in corpus callosum as the primary endpoint because if TBI is a heterogenous pathology when looking at cortical lesions, the corpus callosum shows much more homogeneous patterns, with 75% of TBI presenting anatomical lesions and 90% presenting fractional anisotropy (FA) modifications³⁰ in this VOI.

Simultaneous PET and MR acquisitions will be performed in the Department of Nuclear Medicine of Pitié-Salpêtrière Hospital with a PET/MR system SIGNA 3T on slots dedicated to research. The doses of [¹⁸F]-DPA-714 will be manufactured for this project by the Service Hospitalier Frederic Joliot (SHFJ, CEA) and delivered to Pitié-Salpêtrière Hospital.

A dynamic PET acquisition will start at the time of injection of 200 MBq of [¹⁸F]-DPA-714, in list mode, for up to 90 min. Dynamic data will be binned into 27 time frames (6×1 min, 7×2 min, 14×5 min) and reconstructed. MR sequences will include Dixon and Zero Time Echo sequences for PET attenuation correction, 3D T1 FSPGR, 3D FLAIR, DTI, susceptibility-weighted imaging (SWI).

PET using radioligands specific for monocyte/microglial activation has shown good sensitivity and specificity regarding cerebral inflammation evaluation.³¹ Among the several ligands tested, [¹⁸F]-DPA-714 is one of the most used because of pharmacokinetic advantages.³² When coupled with MRI, PET offers the possibility to quantify simultaneously the volumetric (anatomical region volume) and structural (regional FA and mean diffusion) insults as well as the intensity of global and localised microglial activation. Thus, PET-MRI is currently the most advanced technology to evaluate neuroinflammation, enabling regional analysis with the best spatial resolution. Its performance has been demonstrated in several animal models and in various human pathologies.^33–38

To ensure no or minimum loss to follow-up at this imaging evaluation, dedicated clinical research associate teams will arrange the follow-up and the scheduling of PET-MRI and clinical evaluation.

Secondary outcomes

See table 1 for the full list of secondary outcomes.

Randomisation and sequence generation

The randomisation will be performed using CleanWEB, an online centralised procedure service running 24 hours a day, 7 days a week. The randomisation sequence will be computer generated in advance by a statistician from the coordinating office. It will then be stratified by the centre.

Allocation concealment

The number of experimental units per block will be kept confidential to avoid prediction of future patient’s allocation. Only the independent statistician and the computer programmer who will implement the sequence assignment in the secure electronic case report form (eCRF) will have access to the randomisation list. Allocation concealment will be ensured, as CleanWeb services will not release the randomisation code until the patient has been recruited into the trial.

Blinding

WJ-UC-MSC and placebo will both be prepared by St Louis Hospital MEARY Cell and Gene Therapy Center. They will be conditioned, labelled and delivered the same way in order to maintain blinding. Neither the patients nor the medical staff will be aware of the randomisation arm. The rare, mild and non-specific potential side effects of WJ-UC-MSC will not compromise the blinding at individual level. The study statistician, also, will be blinded to the groups.

PET-MRI analysis

All neuroimaging analyses will be entrusted to engineers with expertise in MRI and PET from BioMaps, the CATI platform (https://cati-neuroimaging.com) and BRAINTALE (www.braintale.eu).

Quantification of post-TBI lesions with MRI

Radiological markers will be extracted from both the initial and the PET MRI. We will extract the regional values of FA, mean diffusibility, axial (L1) and radial (Lt) diffusibility from the DTI acquisition. The lesion load will also be estimated from FLAIR and SWI sequences, and atrophy from 3D T1-weighted images. These measurements will be made in the VOIs corpus callosum, pericontusional areas, grey and white matter, frontal, parietal and occipital lobes, hippocampus, thalamus, mesencephalus and cerebellum. We will analyse both the absolute lesion burden on the PET MRI and its progression between the initial qMRI and this later examination. Both clinical MRI and PET/MR are 3 Tesla General Electrics scanners and harmonisation of MR acquisition parameters will be monitored by the CATI.

Analysis of [18F]-DPA-714 PET images

[¹⁸F]-DPA-714 SUVr parametric maps will be calculated based on reference region, extracted using a supervised clustering algorithm from dynamic (90 min) PET scan. The latter technique has been developed to avoid a potential bias related to the a priori selection of the anatomical reference region (a region that is not affected by the disease). Supervised clustering algorithm (SVCA) is based on the creation of a set of predefined classes from a set of training images that help the algorithm to select only regions of low specific binding from the image. It has been successfully applied to other tracers such as [¹¹C]-PIB³⁹ or [¹¹C]-TMSX.⁴⁰ Recently, the SVCA has been validated for dynamic PET scan using [¹⁸F]-DPA-714.⁴¹ Parametric images (SUVr) will be then analysed with a pipeline developed by the CATI, allowing partial volume correction and VOI measurements on untransformed PET images. Neuroinflammation analysis will be performed using (1) the same regional volumes of interest as for MRI and (2) voxel-wise approach without regional a priori using SPM12.

Statistical considerations

Sample size calculation

Based on a previous prospective study using [¹⁸F]-DPA-714 SUVR in Alzheimer’s disease,³⁶ we extrapolated a 0.20 difference of SUVR between the placebo and experimental groups. With this difference and an SD of 0.25, we need to include 31 patients per group for a power of 90% and a type-I error of 0.05 to reach significance by performing a t-test. Given a randomised ratio of 1:1, 68 patients (31+3 additional patients per group in case of low-binder or death during the follow-up for the intention-to-treat analysis) will be included. Considering the deceased patients (expected rate less than 10%), the imputation rule will set the highest value of the signal intensity [¹⁸F]-DPA-714 SUVr regardless of the randomisation arm.

Statistical analysis

The analyses will follow the intention-to-treat principle.

The mean [¹⁸F]-DPA-714 SUVr in corpus callosum will be compared at 6 months between the two randomisation arms using a Student t-test or a Wilcoxon test, depending on final sample size and normality assessment. TSPO affinity status (LAB or MAB) will be considered as a covariable. The significant level for all statistical analyses will be a two-sided 5%. All statistical analyses will be performed using SAS software (SAS Institute, Cary, North Carolina) V.9.4 or later, or R software (R Foundation for Statistical Computing, Vienna, Austria. http://www.r-project.org/) V.4.0 or later.

All analyses will be conducted by a statistician according to a prespecified statistical analysis plan. A full statistical analysis plan has been written and is available in online supplemental material 1.

All analysis results will be reported according to the Consolidated Standards of Reporting Trials 2010 guidelines.⁴²

Data collection and management

Data collection will be done in electronic format, the statistical software CleanWeb for data entry will be used. The software will fulfil the regulatory requirements and security norms. Data will be handled according to the French law. All original records (including consent forms, reports of suspected unexpected serious adverse reactions and relevant correspondences) will be archived at trial sites for 15 years. The clean trial database file will be anonymised and maintained for 15 years.

We will collect data on primary and secondary endpoints as well as extensive blood samples for biocollection detailed in table 2.

The data of this study will be available on request from the corresponding author. The data will not be publicly available due to privacy and ethical restrictions. See ‘Legal obligations and approval’ for further details.

Patient and public involvement

Patients and public were not involved in any of the phases of the design of this study. Results of the trial will be made available to all participants via ClinicalTrials.gov as well as by email notification.

Trial status

The study started recruitment on 23 September 2024.

Ethics and dissemination

Legal obligations and approval

Sponsorship has been agreed by Assistance Publique—Hôpitaux de Paris (AP-HP, Clinical Research and Innovation Department) for this human research study. AP-HP has obtained the favourable opinion of the Comité de Protection des Personnes (CPP SUD EST II) and the agreement of French National Medicines Agency (Agence Nationale de Sécurité du Médicament et des Produits de Santé, ANSM) (2023-504415-33-00) for the study protocol (TRAUMACELL−V.1.3_20231004). The trial will be carried out in accordance with the Declaration of Helsinki and the Good Clinical Practice guidelines. Any substantial modification to the protocol must be sent to the sponsor for approval. Once approval has been received from the sponsor, it must also obtain approval from the CPP and ANSM before the amendment can be implemented. The information sheet and the consent form can be revised if necessary, particularly if there is a substantial amendment to the study or if adverse reactions occur. AP-HP is the owner of the data. The data cannot be used or disclosed to a third party without its prior permission.

Methods for obtaining information and consent from research participants

In accordance with Article L.1122-1-1 of the French Public Health Code, no research can be carried out on a person without his/her free and informed consent, obtained in writing after the person has been given the information specified in Article L.1122–1 of the said Code. When it is impossible for the person concerned to express their consent in writing, it may be attested by the trustworthy person, by a family member, or, failing that, by one of the close relatives of the person concerned, provided that this trustworthy person, family member or close relative is independent of the investigator and the sponsor.

The trustworthy person, a family member or a close relative will be given a reflection period of at least 1 hour between receiving oral and written information and being asked to sign the consent form for main and ancillary studies (see online supplemental material 2). The trustworthy person’s free and informed written consent will be obtained by the investigator, or by a doctor representing the investigator, before the patient is enrolled into the trial, during the screening visit. The information sheet and a copy of the consent form, signed and dated by the trustworthy person and by the investigator or the doctor representing the investigator, will be given to the individual prior to being enrolled into the trial. In addition, the investigator will specify in the research participant’s medical file the methods used for obtaining their consent as well as the methods used for providing information with a view to obtaining consent. Once the patient is able to consent, we will ask him to sign the pursuit consent form during a follow-up visit (see online supplemental material 2). The investigator will retain the original signed and dated consent forms.

Subjects may exit the study at any time and for any reason.

Data collection and quality control

The persons responsible for the quality control of clinical matters will take all necessary precautions to ensure the confidentiality of information relating to the study participants. These persons, as well as the investigators themselves, are bound by professional confidentiality. During or after the research, all data collected about the participants and sent to the sponsor by the investigators (or any other specialised collaborators) will be anonymised. Under no circumstances should the names, addresses and other protector identifiers of the subjects involved be shown.

In any case of premature withdrawals and exits, the investigator must document their reason(s) and try to collect primary endpoint, secondary endpoints and safety assessment, if the participant agrees. If a participant exits the study prematurely or withdraws consent, any data collected prior to the date of premature exit may still be used except if the participant refuses in writing.

To monitor compliance, all treatment packets will be stored after use for counting and auditing. All treatments (used or not) will be stored in the medical ward and sent to the site pharmacy at study end for destruction.

A data safety monitoring board (DSMB) has been established by the sponsor. The DSMB will operate in accordance with the sponsor’s procedures. The DSMB will work in an advisory capacity only and the sponsor retains all decision-making authority. This has been approved by the sponsor and the steering committee. The research data will be collected and monitored using an eCRF through CleanWEB Electronic Observation Book and will be centralised on a server hosted by the AP-HP Operations Department. This research is governed by the CNIL ‘Reference Method for processing personal data for clinical studies’ (MR-001, amended). AP-HP, the sponsor, has signed a declaration of compliance with this ‘Reference Method’.

Research staff of the sponsor will work with local investigators to obtain data that are as complete and as accurate as possible. An independent Clinical Research Associate appointed by the sponsor will be responsible for the proper running of the study, for collecting, documenting, recording and reporting all handwritten data, in accordance with the Standard Operating Procedures applied within the Clinical Research and Innovation Department of AP-HP. The investigators agree to accept the quality assurance audits carried out by the sponsor as well as the inspections carried out by the competent authorities. All data, documents and reports may be subjected to regulatory audits. These audits and inspections cannot be refused on the grounds of medical secrecy. An audit can be carried out at any time by independent individuals appointed by the sponsor. The aims of the audit are to ensure the quality of the study, the validity of the results and compliance with the legislation and regulations in force. The persons who manage and monitor the study agree to comply with the sponsor’s audit requirements. The audit may encompass all stages of the study, from the development of the protocol to the publication of the results and the storage of the data used or produced as part of the study. Sponsor is responsible for access to the study database and will oversee the intrastudy data sharing process. All principal investigators will be given access to the cleaned data sets. Project data sets will be housed by the sponsor, and all data sets will be password-protected. Project principal investigators will have direct access to their own site’s data sets and will have access to other sites data by request.

Safety considerations

The investigator can temporarily or permanently withdraw a subject from the study for any safety reason or if it is in the subject’s best interests.

The investigator or the treating doctor may request unblinding for any reason s/he considers essential.

According to article R.1123–49 of the French Public Health Code, the investigator must notify the sponsor without delay on the day when the investigator becomes aware of any serious adverse event, which occurs during the trial, related to the studied treatment or not, except those which are listed below as not requiring a notification without delay.

Other events, judged as being ‘medically significant’, require the investigator to notify the sponsor without delay (clinical or biological events that may suggest toxicity or require an increased monitoring of the subjects exposed):

During study treatment infusion

Presence of a clinical presentation compatible with transfusion incompatibility or a transfusion-related infection (urticaria, bronchospasm, etc.)

Within 6 hours following study treatment infusion

• Occurrence of haemodynamic instability requiring the initiation of norepinephrine ≥2 mg/hour; or decrease in blood pressure requiring an increase in the dose of norepinephrine ≥2 mg/hour compared with its initial value.

• Occurrence of ventricular tachycardia, ventricular fibrillation or cardiac arrest.

• Occurrence of severe hypoxemia (PaO2/FiO2) ≤100 mm Hg or ≥50% of its value before infusion.

Within 24 hours following study treatment infusion

• Any cardiac arrest.

Within study treatment period (J0–J30)

• Occurrence of pulmonary embolism.

• Worsening of arterial oxygenation with hypoxaemia unexplained by a ventilation-acquired pneumonia.

• Any clinical event perceived by the clinician as unusual in nature, frequency, incidence or severity in relation to the clinical course of TBI.

The following adverse events, related to the surgery and/or to a pre-existing illness or condition, are simply recorded in the eCRF and do not require the investigator to notify the sponsor without delay. A CRF extraction of these adverse events will be undertaken every 6 months.

Normal and natural course of the condition

• Agitation.

• Delirium.

• Nosocomial infections.

• Pressure wounds.

• Gastrointestinal bleeding.

Special circumstances

• Hospitalisation for a pre-existing illness or condition.

• Hospitalisation for a medical or surgical treatment scheduled prior to the study.

• Admission for social or administrative reasons.

The protocol of administration of the MSC (or placebo) entails three injections 1 week apart, based on the tolerance profile observed in the STROMA-COV study (Umbilical cord-derived mesenchymal stromal cell therapy in SARS-CoV-2-associated acute respiratory distress syndrome).⁴³ In this trial, which has included 47 patients, no treatment-related serious adverse events (particularly those which could have been related to alloimmunisation) have been observed after WJ-UC-MSC injections. Indeed, WJ-UC-MSC weakly express HLA-ABC antigens and do not express HLA-DR nor costimulation molecules, while their expression of HLA-G is stronger than the one of bone marrow-derived MSC.⁴⁴ These characteristics and previous experiences lead to consider them as lowly immunogenic, even in an inflammatory environment.⁴⁵ Thus, extending the interval between MSC injections to give enough time for tracking putative donor-specific antibodies has been deemed unnecessary, and no immunosuppressive concomitant treatment will be administered to the patients included. Measurements of DSA will be performed to monitor safety at the population level, not individually.

To date, [¹⁸F]-DPA-714 has been administered to more than 500 subjects without adverse events. Dosimetry of [¹⁸F]-DPA-714 is extremely low due to the short half-life of fluorine-18 (109.8 min). Radiation exposure (effective dose) is estimated at 0.021 mSv/MBq, that is, 4.2 mSv for an injected dose of 200 MBq. The brain [¹⁸F]-DPA-714 PET scan conducted on hybrid PET-MRI systems entails an overall irradiation due to the injection of the radiotracer of 4.2 mSv (millisievert), which is significantly less than a normal whole-body PET scan used in cancer assessment (25 mSv) and is equivalent to the average annual radiation dose for someone living in France (3.5 mSv/year). There is no CT scan associated with PET-MRI acquisition. There are no side effects reported in the literature for such exposures in adults.

The most common adverse effects described with the radiopharmaceuticals are pain or haematoma at the injection site. No severe effect has been recorded.

Adverse events potentially related to treatments prescribed as part of the care provided during the study follow-up

The mortality rate in severe TBI after sedation discontinuation will be expected at 10%.⁴⁶ If there is any imbalance between the randomisation groups or the mortality rate is higher than expected, affecting the safety of trial subjects and which requires the sponsor to take urgent safety measures, the French National Agency for Medication will be informed about the emerging safety issue without delay.

Trials oversight committees

Two oversight committees have been established to oversee the conduct of this trial, the Steering Committee and Scientific Committee, the composition of each is listed at the end of this paper.

Publication plan

Scientific presentations and reports corresponding to the study will be written under the responsibility of the coordinating investigator of the study with the agreement of the principal investigators and the methodologist. The coauthors of the report and the publications will be the investigators and clinicians involved, on a pro rata basis of their contribution in the study, as well as the biostatistician and associated researchers. Rules on publication will follow international recommendations.⁴⁷

We would like to thank Dr Era Soukhin (Auckland City Hospital, NZ) for kindly proofreading the manuscript.

Ethics statements

Patient consent for publication

Not applicable.

Footnote

X @stephsig

Contributors SS contributed to the conception and design of the research protocol, assisted by CC, CT, A-MF, IC and VD, under the supervision of PM. AJ, MB, PG, M-OH, AM, AR, MB, DG, AC and JL provided critical input pertaining to the design of the trial interventions and procedures. SS and CC wrote the first draft of the protocol and this manuscript. CT and IC designed and wrote the statistical analysis plan. All authors critically revised and modified the protocol and the article. They all approved the final version to be published. SS is the guarantor of the content of the manuscript.

Funding The TRAUMACELL trial is is funded by a grant from AAP ANR/DGOS 2021 and by the Keep Fighting Foundation. The funders had no role in the trial design, trial conduct, data handling, data analysis or writing and publication of the manuscript.

Competing interests None declared.

Patient and public involvement Patients and/or the public were not involved in the design, or conduct, or reporting, or dissemination plans of this research.

Provenance and peer review Not commissioned; externally peer-reviewed.

Supplemental material This content has been supplied by the author(s). It has not been vetted by BMJ Publishing Group Limited (BMJ) and may not have been peer-reviewed. Any opinions or recommendations discussed are solely those of the author(s) and are not endorsed by BMJ. BMJ disclaims all liability and responsibility arising from any reliance placed on the content. Where the content includes any translated material, BMJ does not warrant the accuracy and reliability of the translations (including but not limited to local regulations, clinical guidelines, terminology, drug names and drug dosages), and is not responsible for any error and/or omissions arising from translation and adaptation or otherwise.

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