1Centre for Human Drug Research (CHDR), Leiden, Netherlands 2Leiden University Medical Center, Leiden, Netherlands 3Leiden Academic Centre for Drug Research, Universiteit Leiden, Leiden, Netherlands 4Poolbeg Pharma Limited, London, United Kingdom 5Department of Experimental and Translational Medicine, University College London Research, London, United Kingdom
Background and aim: POLB 001 is an oral p38 mitogen-activated protein kinase (MAPK) inhibitor in development for the prevention of cancer immunotherapy-induced cytokine release syndrome (CRS). It has previously been shown to be well tolerated and capable of decreasing ex vivo lipopolysaccharide (LPS)-induced tumour necrosis factor (TNF) secretion in a phase 1 first-in-human trial. This study aimed to evaluate the anti-inflammatory effects of POLB 001 following in vivo LPS administration in healthy volunteers.
Methods: Participants received POLB 001 at doses of 30, 70, or 150 mg, or placebo, twice daily for seven consecutive days and were challenged locally with intradermal (ID) LPS on day 4 and systemically with intravenous (IV) LPS on day 6. Following ID LPS administration, skin perfusion and erythema were measured, and skin suction blisters were created to collect blister fluid containing infiltrating immune cells and extracellular fluid. Following IV LPS administration, circulating cytokine levels, leukocyte counts, leukocyte p38 MAPK phosphorylation levels, and vital signs were measured.
Results: POLB 001 was well tolerated. It reduced the ID LPS-driven immune cell attraction and cytokine responses measured in blister fluid. The suppression of immune cell recruitment was most pronounced in neutrophils (72.4%–81.5%, p = 0.0091), classical monocytes (68.4%–73.6%, p = 0.0036), CD3+ T cells (56.4%–65.9%, p = 0.0047), and myeloid dendritic cells (59%–64.4%, p = 0.0174). The suppression of cytokine responses was most pronounced for TNF (35.3%–65.1%, p = 0.0099). Overall, POLB 001 did not substantially modulate the intradermal LPS-driven increase in local erythema and perfusion. POLB 001 significantly reduced the IV LPS-driven increase in interleukin (IL)-6, IL-8, and TNF (37.7%–80.7%, all p < 0.0003), p38 MAPK phosphorylation levels in target cells (16.7%–60.9%, all p < 0.0001), and heart rate increase (4–9.3 bpm, p < 0.0001).
Conclusion: POLB 001 was safe and well-tolerated. Pharmacodynamic findings confirm that POLB 001 inhibits LPS-induced local and systemic inflammation in vivo through inhibition of p38 MAPK.
Clinical trial registration: https://onderzoekmetmensen.nl/en/trial/51741, identifier NL81214.056.22.
Mitogen-activated protein kinases (MAPKs) are components of signalling cascades that initiate inflammatory cellular responses when activated by extracellular stimuli (1). Several subgroups of MAPKs have been identified, including p38 MAPK. Triggers for p38 MAPK activation include lipopolysaccharide (LPS) and other bacterial products, ultraviolet irradiation, hypoxia, and inflammatory cytokines (1, 2). Activation of the p38 signalling pathway plays an important role in the production of various inflammatory mediators, such as tumour necrosis factor (TNF) (3, 4). Inhibition of the p38 MAPK signalling pathway results in reduced production of these inflammatory mediators.
POLB 001 (previously known as Org 48775-0) is a potent and highly selective p38α/β MAPK inhibitor in development for cancer immunotherapy-induced cytokine release syndrome (CRS). POLB 001 has shown efficacy in several pharmacological disease models, including humanised mouse models of CRS (5). It was well tolerated under both fed and fasted conditions in a previous phase 1 trial in healthy male volunteers. In that first-in-human single ascending dose (SAD) study and a multiple ascending dosing (MAD) study, POLB 001 was administered as a single dose up to 600 mg and as multiple doses twice daily for six consecutive days, up to 150 mg. Using peripheral whole blood samples from treated participants, POLB 001 was shown to effectively decrease LPS-induced TNF release (6).
While previous results demonstrate that POLB 001 decreases LPS-induced TNF release from circulating leukocytes ex vivo, the effects of POLB 001 on in vivo inflammatory responses were unknown. Ex vivo refers to the evaluation of drug activity in a test tube, with the immune trigger added to the biospecimen obtained from drug-exposed participants. In contrast, in vivo refers to the evaluation of drug activity in humans, where drug-treated participants were exposed to the immune trigger. The aim of this phase 1 study was to determine the inhibitory effects of POLB 001 on the inflammatory response following local and systemic inflammation in healthy volunteers, upon intradermal (ID) and intravenous (IV) LPS administration. A multimodal pharmacodynamic evaluation approach was employed, comprising clinical endpoints, imaging-based endpoints, local tissue response endpoints, and blood-based endpoints.
This was a randomised, double-blind, placebo-controlled, parallel-group, MAD study in healthy volunteers. The study was executed at the Centre for Human Drug Research, the Netherlands, between August 2022 and December 2022. It adhered to ethical principles derived from international guidelines, including the Declaration of Helsinki, as well as all applicable ICH, GCP, and Dutch Medical Research Involving Human Subjects Act guidelines. The study protocol was registered in the EudraCT database (number 2022-001458-48). The protocol (registered at Toetsingonline, number NL81214.056.22) was approved by a medical ethics committee (Stichting Beoordeling Ethiek Biomedisch Onderzoek, Assen, the Netherlands) prior to the start of the clinical phase. All participants provided written informed consent before any study-related activities were performed.
The study aimed to recruit 36 healthy volunteers. Volunteers were eligible if they were men, had Fitzpatrick skin type I–III, and were aged between 18 and 55 years. Male volunteers were excluded if any of the following criteria applied: previous participation in a systemic (IV or inhaled) LPS challenge study within a year before the first study day, a current or recurrent clinically significant skin condition at the intradermal LPS target areas (including tattoos), or a known immunodeficiency.
Participants received oral doses of POLB 001 (30 mg—low dose, 70 mg—intermediate dose, 150 mg—high dose) or a matching placebo twice daily for seven consecutive days (days 1 to 7) (Figure 1). Participants were scheduled to take the study medication either at the study site or at home. Drug intake at home was monitored using an electronic Patient Reported Outcomes (ePRO) application. All participants received the ID challenge on day 4 and the IV LPS challenge on day 6 upon admission to the clinical unit. Safety monitoring was performed throughout the study course for all participants. Blood samples for PK serum analysis were collected predose and repeatedly on days 1, 4, and 6, and were analysed at the Ardena Bioanalytical Laboratory (Assen, the Netherlands).
Figure 1. Overview of the study design. POLB 001 or a matching placebo was administered twice daily for seven consecutive days. On day 4, participants received four intradermal injections of LPS. The local response was evaluated at specified timepoints by using different imaging techniques and skin blister fluid analysis. On day 6, participants received a single dose of intravenous LPS, with pre- and posthydration. Endpoints included different blood assessments and vital signs. The asterisk indicates PK on day 1 only for the highest dose group. PK, pharmacokinetics; LPS, lipopolysaccharide; MSD, Meso Scale Discovery; CRP, C-reactive protein. Created with Biorender.com.
Both the study medication and LPS (for both ID and IV administration) were formulated by the pharmacy at the Leiden University Medical Centre, Leiden, the Netherlands. POLB 001 was administered as an oral suspension of 30, 70, or 150 mg POLB 001 powder in 20 mL SyrSpend® Cherry SF PH4 Vehicle. For each dose level, a matching placebo was prepared. Participants were instructed to fast for 8 h prior to every morning dose and for 1 h prior to each evening dose. Purified LPS, prepared from Escherichia coli O113:H10:K negative (US Standard Reference Endotoxin), was used as a challenge agent. The LPS batch was manufactured in the United States (USA) by List Biological Laboratories (Campbell, California). Each participant received four ID LPS injections on the volar forearms, each consisting of 5 ng LPS in 0.05 mL of 0.9% sodium chloride (NaCl) solution (a total of 20 ng per participant). A different LPS injection site was used for each blister timepoint. Participants also received an IV infusion of glucose (2.5% w/v) and NaCl (0.45% w/v) solution, starting 2 h prior to LPS administration and continuing for 6 h afterward. IV LPS consisted of 1 ng/kg LPS in 10 mL of 0.9% NaCl solution, administered as a 2-min infusion (7).
For the evaluation of ex vivo pharmacodynamic effects, LPS was added as an immune trigger to test tubes containing whole blood obtained from drug-exposed participants. Baseline whole blood samples were collected on days 1 and 6 prior to the IV LPS challenge. Blood was diluted (1:1) with RPMI 1640 supplemented with 25 mM HEPES and l-glutamine, and stimulated with LPS (2.5 ng/mL). Samples were incubated for 24 h at 37 °C (5% CO2), and the supernatant was stored. Secreted interleukin (IL)-1β, IL-6, IL-8, and TNF were analysed from the supernatant using a Meso Scale Discovery (MSD) multiplex immunoassay (Ardena, Assen, the Netherlands).
The skin was imaged pre-LPS and at 3, 9, and 24 h afterward. Perfusion was evaluated using laser speckle contrast imaging (LSCI) (PeriCam PSI System, Perimed Jäfälla, Sweden), and erythema was assessed with multispectral imaging using Antera 3D (Miravex, Dublin, Ireland). The performance and analysis of these noninvasive measurements were standardised for all participants.
After skin imaging was performed, 10 mm suction blisters were generated over the LPS-injected skin areas, as previously reported (8). Three suction blisters were created per participant: at 3, 9, and 24 h after the ID LPS injections.
Blister fluid was collected in a plate containing 50 μL of 3% cold sodium citrate (Sigma‐Aldrich, St. Louis, MO, USA) in PBS (Gibco, Thermo Fischer Scientific, Waltham, MA, USA). Levels of IL-1β, IL-6, IL-8, and TNF in blister fluid were analysed by MSD (Ardena, Assen, the Netherlands). Leukocytes in blister fluid were immunophenotyped by flow cytometry (for antibodies, see Supplementary Table S1). The gating strategy for blister leukocytes was selected using a peripheral blood sample collected before LPS administration (see Supplementary Figure S1). Flow cytometry measurements were done in the MACSQuant Analyzer 16 (Miltenyi Biotec GmbH, Bergisch Gladbach, Germany), and data were analysed with Flowlogic 8.7 (Inivai Technologies, Mentone, VIC, Australia) at the Centre for Human Drug Research (CHDR) laboratory (Leiden, the Netherlands).
Blood samples for p38 MAPK phosphorylation analyses were collected prior to IV LPS administration and at 1, 1.5, and 2 h afterward. Within 1 h of collection, sodium heparinised whole blood was fixed, and red blood cell lysis was performed simultaneously. Cells were permeabilised and stained with anti-CD14 (REA 599) and p38 MAPK antibody (RUO 36/p38 [pT180/pY182]) in the same test tube (see Supplementary Table S1). Live cells were gated from the forward scatter (FSC) vs. side scatter (SSC) leukocyte population, and doublets were removed by gating for FSC-Height (FSC-H) vs. FSC-Area (FSC-A) (Supplementary Figure S2). Monocytes were identified as CD14+. Phosphorylation levels of p38 MAPK in whole blood and in CD14+ cells were expressed as a percentage and median fluorescence intensity (MFI). Analysis was performed at the laboratory of CHDR (Leiden, the Netherlands).
Vital sign measurements, including heart rate, blood pressure, and temperature, were measured to evaluate the clinical response to IV LPS.
Blood was collected from each participant prior to IV LPS administration and 1–4 h (every hour), 6 h, and 9 h after IV LPS administration for analysis of circulating cytokine levels by MSD: IL-1β, IL-6, IL-8, IL-10, and TNF (Ardena, Assen, the Netherlands).
Blood samples for leukocyte differential analysis were collected prior to the IV LPS administration and at 3, 6, and 24 h after LPS administration. Blood samples for C-reactive protein (CRP) analysis were collected prior to the IV LPS challenge and 6, 9, and 24 h after administration. All blood samples were analysed at the laboratory of the Leiden University Medical Center (“Klinische chemie en laboratoriumgeneeskunde” LUMC, Leiden, the Netherlands).
A sample size of nine per treatment group was deemed sufficient based on historical data (8–10). This sample size provides at least 80% power to detect a 50% decrease in LPS-induced systemic and dermal responses using a two-sided significance level of 0.05. Repeatedly measured data were analysed with a mixed model analysis of covariance (ANCOVA) with treatment, time, and treatment-by-time as fixed factors, subject as a random factor, and the (average) baseline measurement as a covariate. Treatment effects were calculated for each parameter over the time period from baseline up to 24 h, except for p38 MAPK phosphorylation (up to 2 h postdose), ex vivo analysis (baseline day 1 vs. day 6 pre-IV LPS), and circulating cytokines (up to 9 h postdose). The following comparisons were calculated for both the ID and IV LPS challenges (analysed separately): POLB 001 at 30 mg vs. pooled placebo, POLB 001 at 70 mg vs. pooled placebo, and POLB 001 at 150 mg vs. pooled placebo. Comparisons were reported with estimated differences, 95% confidence intervals (CI), and p-values. All calculations were performed using SAS for Windows V9.4 (SAS Institute Inc., Cary, NC, USA).
Sixty-three participants were screened for this study. Of these, 41 participants, including reserve participants, were enrolled. Thirty-six participants were treated in three cohorts of 12 participants each (nine active vs. three placebo).
All 36 participants were men, and 94.4% were White. The mean ± SD age was 28.6 years ± 10.1 years. Further baseline characteristics are presented in Table 1.
Table 1. Baseline demographic characteristics.
Administration of POLB 001 for seven consecutive days was well tolerated. No severe adverse events (SAEs) occurred following dosing. No meaningful relationships between POLB 001 dose and the incidence of adverse events (AEs) were observed. No clinically relevant changes in ECG parameters, blood chemistry, or haematology—apart from CRP and leukocyte differential levels described as PD endpoints and an increase in liver enzymes in one participant—were observed.
A full list of AEs can be found in Supplementary Table S2. Headache was the most commonly reported AE overall and in the POLB 001 treatment groups. Dizziness, fatigue, and chills were additional commonly reported AEs in all treatment groups. Only three participants who received POLB 001 reported events considered probably related to the study drug. Most AEs were possibly or unrelated to POLB 001 and could be related to the LPS challenges.
POLB 001 showed dose-proportional PK for Cmax and exposure over 12 h (AUC12 h), in line with the previous study (6).
LPS-induced IL-1β, IL-6, IL-8, and TNF release ex vivo was decreased by POLB 001 in the day 6 samples compared to baseline. Percentages of inhibition by POLB 001 ranged from 38.8% to 42.2% for IL-1β, 32.1% to 52.2% for IL-6, 52.1% to 54.7% for IL-8, and 53% to 59.8% for TNF (Supplementary Figure S3).
After administration of ID LPS, a gradual increase in skin erythema and perfusion was observed up to 24 h (Figure 2). POLB 001 at 30 mg slightly but significantly decreased LPS-driven erythema compared to placebo (change: − 0.6782 Antera® skin colour a* units [redness; p < 0.05]). However, the POLB 001 at 70 and 150 mg dose groups showed no significant effect compared to placebo (change: − 0.5495 and 0.0068 Antera® skin colour a* units with p = 0.1040 and p = 0.9831, respectively). Consistent with this, POLB 001 did not significantly reduce the LPS-driven increase in skin perfusion compared to placebo.
Figure 2. Quantification of the inflammatory skin response following the intradermal LPS challenge by (A) multispectral imaging (erythema) and (B) laser speckle contrast imaging (LSCI; perfusion). In (A), POLB 001–30 mg vs. placebo showed p < 0.05, whereas other contrasts showed p > 0.05. In (B), POLB 001 vs. placebo, all p > 0.05. Data are expressed as means with SD. AU, arbitrary units; ID, intradermal; LPS, lipopolysaccharide.
In the absence of POLB 001, ID LPS administration led to an increase in neutrophils and classical monocytes, peaking at 9 h, and an increase in myeloid dendritic cells (mDCs), plasmacytoid DCs (pDCs), CD3+ T cells, CD4+ T cells, CD8+ T cells, nonclassical monocytes, intermediate monocytes, B cells, natural killer (NK) cells, and total immune cells, peaking at 24 h in blister fluid (Figure 3, Supplementary Figure S4). For cytokines, a rapid response of IL-8 and TNF was observed, peaking at 3 h, whereas a more delayed response was observed for IL-1β and IL-6, peaking at 9 h (Figure 4).
Figure 3. Overview of skin blister fluid analysis by flow cytometry following intradermal LPS administration: (A) CD3+ T cells, (B) CD4+ T cells, (C) CD8+ T cells, (D) mDCs, (E) classical monocytes, and (F) neutrophils. In (A, B), POLB 001 vs. placebo, all p < 0.01. In (C), POLB at 30 mg vs. placebo showed p > 0.05, whereas POLB 001 at 70 and 150 mg vs. placebo showed p < 0.01. In (D), POLB 001 vs. placebo, all p < 0.05. In (E), POLB 001 vs. placebo, all p < 0.01. In (F), POLB 001 vs. placebo, all p < 0.05. Data are expressed as means with SD. ID, intradermal; LPS, lipopolysaccharide; mDCs, myeloid dendritic cells; SD, standard deviation.
Figure 4. Overview of cytokine responses in skin blister fluid following intradermal LPS administration: (A) IL-1β response, (B) TNF response, (C) IL-8 response, and (D) IL-6 response. In (A), POLB 150 mg vs. placebo showed p < 0.02, whereas other contrasts showed p > 0.05. In (B), POLB 150 mg vs. placebo showed p < 0.02, whereas other contrasts showed p > 0.05. In (C), POLB 001 vs. placebo, all p > 0.05. In (D), due to too many values above the ULOQ, the requirement for analysis was not met. Data are expressed as means with SD. ID, intradermal; LPS, lipopolysaccharide; TNF, tumour necrosis factor; IL, interleukin.
Treatment with POLB 001 resulted in a decrease in the number of CD3+ T cells, CD4+ T cells, mDCs, classical monocytes, neutrophils, and total immune cells in blister fluid compared with placebo treatment (Figures 3A, B, D–F, Supplementary Figure S4A). This inhibition was dose-dependent for CD3+ T cells, classical monocytes, and total immune cells, with percentages of suppression ranging from 56.4% to 65.9% (all dose groups p < 0.01 vs. placebo) for CD3+ T cells, 68.4% to 73.6% (all dose groups p < 0.01 vs. placebo) for classical monocytes, and 65.5% to 70.9% (all dose groups p < 0.01 vs. placebo) for total immune cells. For mDCs, CD4+ T cells, and neutrophils, percentages of inhibition compared with placebo were comparable across the three dose levels: 59.0% to 64.4% for mDCs (all dose groups p < 0.05 vs. placebo), 62.8% to 66.2% for CD4+ T cells (all dose groups p < 0.01 vs. placebo), and 72.4% to 81.5% for neutrophils (all dose groups p < 0.05 vs. placebo). Additionally, CD8+ T cells and NK cells were reduced by POLB 001 treatment (Figure 3C, Supplementary Figure S4B), with significant inhibition observed for POLB 001 at 70 and 150 mg compared to placebo (64.1% and 66.6% for CD8+ T cells, 61.9% and 63.0% for NK cells, respectively; all dose groups p < 0.01 vs. placebo). For pDCs, nonclassical monocytes, intermediate monocytes, and B cells, there were too many zero values to allow formal analysis, mainly due to (i) a biologically delayed response to ID LPS by particular cell types, (ii) overall low cell counts, and/or (iii) substantially decreased infiltration induced by POLB 001. However, graphical presentations of nonclassical monocytes, intermediate monocytes, and B cells showed an apparent inhibitory effect of POLB 001 compared to placebo, which appeared to be dose-dependent for nonclassical and intermediate monocytes (Supplementary Figures S4C–E). Graphical presentation of pDCs also showed an apparent inhibitory effect with the two highest dose groups (POLB 001 at 70 and 150 mg) at 24 h compared to the placebo group (Supplementary Figure S4F).
POLB 001 at 150 mg significantly decreased LPS-driven IL-1β (47.3%, p < 0.02 vs. placebo) and TNF (65.1%, p < 0.001 vs. placebo) responses in blister fluid compared to placebo (Figures 4A, B). No significant effect of POLB 001 compared to placebo was observed for IL-8 (Figure 4C). For IL-6, the requirements for analysis were not met due to too many values above the upper limit of quantification (ULOQ), mostly in the placebo group (Figure 4D).
IV LPS administration resulted in the phosphorylation of p38 MAPK in whole blood and CD14+ monocytes, peaking at 1 h in the placebo group (Figure 5 for CD14+ monocytes; whole blood data not shown).
Figure 5. Overview of p38 MAPK phosphorylation in CD14+ monocytes expressed as (A) MFI and (B) percentage. In (A), POLB 001 vs. placebo, all p < 0.02. In (B), POLB 001 vs. placebo, all p < 0.05. Data are expressed as means with SD. IV, intravenous; LPS, lipopolysaccharide; MAPK, mitogen-activated protein kinase; MFI, mean fluorescence intensity.
The level of p38 MAPK phosphorylation in whole blood upon IV LPS challenge was significantly lower in all participants receiving POLB 001, with an inhibition range of 16.7%–28.1% (all dose groups p < 0.01 vs. placebo), although no clear dose response was observed. LPS-induced p38 MAPK phosphorylation in CD14+ monocytes was significantly and dose-dependently decreased by POLB 001, with an inhibition range of 32.0%–60.9% (all dose groups p < 0.02 vs. placebo) compared to the placebo group.
Heart rate (HR) increased in all participants as expected following IV LPS, peaking at 3–4 h post-LPS (Figure 6A). The increase in HR was 4.0–9.3 beats per minute (bpm) lower in participants receiving POLB 001 compared to placebo (p < 0.05 for POLB 001 at 30 mg vs. placebo and p < 0.001 for the two highest dose groups vs. placebo). Maximal inhibition was reached for POLB 001 at 70 mg. An increase in body temperature following IV LPS was also observed in all participants (Figure 6B), whereas minimal mean changes in blood pressure were noted (Figures 6C, D). POLB 001 had no statistically significant effect on LPS-driven changes in body temperature or blood pressure.
Figure 6. Overview of vital sign measurements during the IV LPS challenge: (A) heart rate, (B) temperature, (C) systolic blood pressure, and (D) diastolic blood pressure. In (A), POLB 001 vs. placebo, all p < 0.05. In (B–D), POLB 001 vs. placebo, all p > 0.05. Data are expressed as means with SD. IV, intravenous; LPS, lipopolysaccharide; bpm, beats per minute; °C, degrees Celsius.
In the placebo group, IV LPS caused a rapid increase in IL-6, IL-8, TNF, IL-10, and IL-1β in plasma, peaking 1–4 h after administration (Figures 7A–E). An increase in CRP levels was also observed, peaking at 24 h (Figure 7F).
Figure 7. Overview of the cytokine and CRP response in plasma during the IV LPS challenge: (A) IL-6, (B) IL-8, (C) TNF, (D) IL-10, (E) IL-1β, and (F) CRP. In (A–C), POLB 001 vs. placebo, all p < 0.05. In (D), POLB 001 at 30 mg vs. placebo showed p > 0.05, whereas POLB 001 at 70 and 150 mg vs. placebo showed p < 0.001. In (E), due to levels below the LLOQ, the requirements for analysis were not met. In (F), POLB 001–30 at mg vs. placebo showed p > 0.05, whereas POLB 001 at 70 and 150 mg vs. placebo showed p < 0.05. Data are expressed as means with SD. IV, intravenous; LPS, lipopolysaccharide; TNF, tumour necrosis factor; IL, interleukin; CRP, C-reactive protein.
Treatment with POLB 001 resulted in a significant, dose-dependent reduction in the LPS-driven IL-6 response, with an inhibition range of 37.7%–63.5% compared with placebo (all dose groups p < 0.05 vs. placebo). A significant reduction in the LPS-driven increase in IL-8 and TNF in plasma was observed in all POLB 001 treatment groups compared with the placebo group (all dose groups p < 0.05 vs. placebo). Maximal inhibition appeared to be reached at a dose of 70 mg, with 80.7% for IL-8 and 73.5% for TNF. For IL-10, a significant inhibition was observed for POLB 001 at 70 mg (62.4%) and 150 mg (62.7%) compared with placebo (both p < 0.001 vs. placebo). IL-1β levels in plasma were below the LLOQ, and the data distribution was not normal or log-normal; therefore, the requirements for analysis were not met.
POLB 001 treatment dose-dependently decreased the LPS-driven CRP response, reaching statistical significance for the intermediate dose group, with a decrease of 33.1%, and for the highest dose group, with a decrease of 33.3%, compared with the placebo group (p < 0.05 vs. placebo).
Administration of IV LPS resulted in a transient increase in neutrophils and a biphasic response in monocytes (an initial decrease at 3 h followed by an increase at 6 h), as observed in placebo participants (Supplementary Figure S5). Moreover, a mild decrease in lymphocytes, eosinophils, and basophils was observed following IV LPS administration (data not shown).
The LPS-driven drop in monocytes was suppressed by POLB 001 in all treatment groups compared to the placebo group (30.9%–52.4%, all dose groups p < 0.05 vs. placebo). This effect was not dose-dependent. Neutrophils following IV LPS administration were significantly higher in the intermediate POLB 001 dose group compared to the placebo group (22.5%, p < 0.05). For eosinophils, basophils, and lymphocytes, no significant effect of POLB 001 compared with placebo was observed.
The main goal of this randomised controlled trial was to evaluate the effect of POLB 001 on local and systemic inflammatory responses following in vivo LPS administration in healthy volunteers. In addition, the safety, tolerability, and PK of POLB 001 were assessed. The doses of POLB 001 administered in this study matched those tested in the previous MAD study (30, 70, and 150 mg twice daily).
POLB 001 was well tolerated, and no treatment-related SAEs or death occurred. The safety profile observed in this study was similar to that reported in the phase 1 study (6), with headache, chills, and dizziness being the most commonly reported AEs. No trends in AEs were observed for POLB 001. As other p38 MAPK inhibitors have been associated with liver enzyme elevations, special attention was paid to increases in liver enzymes, particularly Alanine aminotransferase (ALAT). One participant who received the highest dose of POLB 001 had out-of-range ALAT levels, which resolved by day 21.
POLB 001 decreased ex vivo TNF secretion, consistent with previously published results (6). Blood samples for ex vivo analysis in this study were collected at baseline and on day 6, before administration of the next POLB 001 dose. Compared to the same timepoint (before the next dose was taken) in the previous study, the percentages of inhibition were similar (± 50%–60%).
Administration of ID and IV LPS in vivo resulted in consistent, well-detectable inflammatory responses, as expected and in line with previous studies (7, 8, 10, 11). POLB 001 had no meaningful effects on skin erythema or local perfusion after ID LPS administration. However, POLB 001 significantly decreased the attraction of CD3+ T cells, classical monocytes, mDCs, T-helper cells, and neutrophils in blister fluid at all dose levels tested. Local LPS-induced increases in IL-1β and TNF were suppressed by the highest dose of POLB 001.
Furthermore, the effect of POLB 001 on systemic inflammatory responses was evaluated using an IV LPS challenge. LPS-driven p38 MAPK phosphorylation in CD14+ monocytes was lower in all POLB 001 treatment groups compared with the placebo group, showing a clear dose-dependent effect. Additionally, POLB 001 treatment resulted in lower levels of circulating cytokines (IL-6, IL-8, IL-10, and TNF) and reduced CRP levels following the IV LPS challenge.
The p38 MAPK inhibitors RWJ-67657 and BIRB 796 BS were evaluated in healthy volunteers following an IV LPS dose of 4 ng/kg (1, 12–14). Consistent with our study, both inhibitors suppressed the LPS-induced increase of TNF, IL-6, and IL-8. In addition, both reduced the LPS-induced increase in temperature, and RWJ-67657 also reduced the LPS-induced increase in blood pressure and modulated leukocyte functions. Greater fluctuations in vital signs were observed in these studies compared with our study, which can be attributed to the fourfold higher LPS dose. PF-03715455, PH-797804, and AZD7624 were also shown to inhibit plasma cytokines and neutrophilic airway inflammation following an inhaled LPS challenge in healthy volunteers (15, 16).
Over 20 other p38 MAPK inhibitors have been developed and evaluated in clinical trials across different patient populations. Several have shown (early) evidence of clinical utility, including neflamapimod, pexmetinib, and PH-797804. Neflamapimod demonstrated multiple positive effects in patients with mild Alzheimer’s disease (AD) and in patients with dementia with Lewy bodies and will be further investigated in phase 2b clinical trials for both indications (17, 18). In combination with nivolumab and/or ipilimumab, pexmetinib elicited disease control in a number of PD-(L)1-refractory patients with advanced solid tumours in a phase 1b study (19). PH-797804 demonstrated improvements in lung function and dyspnoea in a phase II study in patients with moderate to severe chronic obstructive pulmonary disease (COPD) (20). Other p38 MAPK inhibitors, such as dilmapimod, ralimetinib, losmapimod, BMS-582949, VX-702, pamapimod, and LY3007113, were evaluated in patients at risk for acute respiratory distress syndrome (ARDS), patients with neuropathic pain, (advanced) cancer, arterial inflammation, or rheumatoid arthritis (RA), but were less successful, or their clinical development was not progressed (21–33).
POLB 001 is in clinical development for the prevention of cancer-immunotherapy-induced CRS. CRS is a common adverse event following CAR T-cell therapies and bispecific antibody therapies. It is characterised by fever, tachypnoea, headache, tachycardia, hypotension, rash, and/or hypoxia caused by the release of cytokines. CRS can progress rapidly, with potentially life-threatening effects, and requires close observation, management, and early intervention. The significant risk of CRS is one of several systemic barriers to wider access to CAR T-cell and T-cell-engaging bispecific antibodies, increasing the cost of treatment and limiting access by requiring a period of inpatient care or close monitoring following treatment. Existing treatment options have been investigated in a prophylactic setting but are less effective at preventing grade ≥ 2 CRS, which requires prompt intervention.
In conclusion, POLB 001 was shown to be safe and effective in decreasing local and systemic LPS-induced inflammation in healthy male volunteers. Moreover, this study provides a solid basis for continuing investigations of POLB 001 in patients with inflammatory conditions.
Raw data from the current study are available from the corresponding author upon reasonable request.
The studies involving humans were approved by Stichting Beoordeling Ethiek Biomedisch Onderzoek, Assen, the Netherlands. The studies were conducted in accordance with the local legislation and institutional requirements. The participants provided their written informed consent to participate in this study.
Dd: Writing – original draft. MJ: Writing – original draft. DP: Writing – review & editing. Lv: Writing – review & editing. EK: Writing – review & editing. MO: Writing – review & editing. JS: Writing – review & editing. PM: Writing – review & editing. LT: Writing – review & editing. AB: Writing – review & editing. LM: Writing – review & editing. KM: Writing – review & editing. MS: Writing – review & editing. DG: Writing – review & editing. NK: Writing – review & editing. MM: Writing – original draft.
The author(s) declared that financial support was received for this work and/or its publication. This study was funded by Poolbeg Pharma Limited. POLB 001 was provided by Poolbeg Pharma limited.
Authors JS, PM, LT, AB, LM, KM, and MS were employed by company Poolbeg Pharma Limited. DG received funds for scientific advice by Poolbeg Pharma Limited.
The remaining author(s) declared that this work was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.
The author MM declared that they were an editorial board member of Frontiers, at the time of submission. This had no impact on the peer review process and the final decision.
The author(s) declared that generative AI was not used in the creation of this manuscript.
Any alternative text (alt text) provided alongside figures in this article has been generated by Frontiers with the support of artificial intelligence and reasonable efforts have been made to ensure accuracy, including review by the authors wherever possible. If you identify any issues, please contact us.
All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.
The Supplementary Material for this article can be found online at: https://www.frontiersin.org/articles/10.3389/fimmu.2025.1684307/full#supplementary-material
Supplementary Figure 1 | Gating strategy of immunophenotyping of blister cells. Expression of cell counts of the different cell subsets present in blister cell fluid are defined in a gating strategy for a whole blood sample (Bsgating). Events were gated in FSC vs. SSC dot plot and debris were excluded. Cells were gated on FSC-Area (A) vs. FSC-Height (H) dot plot to eliminate doublets. To define cell populations, first, live (PI-) CD45+ cells are gated. From the leukocyte gate, T cells are identified as CD3+ (total), and further subdivided into CD4+ (T helper, Th) and CD8+ (T cytotoxic, Tc). Next, total HLA-DR+ cells are gated. From HLA-DR+ gating, B cells are CD19+. From the CD19- downstream gate monocyte subsets are further characterised as: CD14+CD16- (classical), CD14+CD16+ (intermediate) and CD14-CD16+ (non-classical). From the HLA-DR+, CD14- CD16- cells, dendritic cells are defined as CD11c+ for myeloid DCs (mDCs) and CD123+ for plasmacytoid DCs (pDCs). Neutrophils are identified as SSChi, CD16+, CD66b+. Natural killer (NK) cells are identified from total leukocytes as CD56+. FSC, forward scatter; SSC, side scatter.
Supplementary Figure 2 | Gating strategy for determining phosphorylation levels of p38 MAPK in ex-vivo LPS challenged whole blood. Events were gated in FSC vs. SSC dot plot and debris were excluded. Cells were gated on FSC-Area (A) vs. FSC-Height (H) dot plot to eliminate doublets. To define cell populations, first, live cells (DAPI, VioBlue -) are gated. From the leukocyte gate, monocytes are identified as CD14+. Phosphorylation levels of p38 MAPK in all cells - whole blood (live cell gating) and monocytes (CD14+) are expressed as percentage (%) and median fluorescence intensity (MFI) of p38 MAPK (PE).
Supplementary Figure 3 | Ex vivo cytokine release measured in Day 6 samples compared to baseline for all treatment groups: (A) IL-1β, (B) IL-6, (C) IL-8, and (D) TNF. TNF, tumour necrosis factor; IL, interleukin.
Supplementary Figure 4 | Overview of skin blister fluid analysis by flow cytometry following intradermal LPS administration: (A) Total immune cells, (B) NK cells, (C) non-classical monocytes, (D) intermediate monocytes, (E) B cells, and (F) pDCs. Data are expressed as means with SD. ID, intradermal; LPS, lipopolysaccharide; NK, natural killer; pDCs, plasmacytoid dendritic cells.
Supplementary Figure 5 | Circulating leukocyte subset levels during the IV LPS challenge: (A) neutrophils and (B) monocytes. Data are expressed as means with SD. IV, intravenous; LPS, lipopolysaccharide.
Supplementary Table 1 | Antibodies used for immunophenotyping of blister cells and to measure phosphorylation levels of p38 MAPK.
Supplementary Table 2 | Treatment-emergent adverse events by preferred term by treatment group (including adverse events reported during the intradermal and intravenous LPS challenge).
1. Branger J, van den Blink B, Weijer S, Madwed J, Bos CL, Gupta A, et al. Anti-inflammatory effects of a p38 mitogen-activated protein kinase inhibitor during human endotoxemia1. J Immunol. (2002) 168:4070–7. doi: 10.4049/jimmunol.168.8.4070
PubMed Abstract | Crossref Full Text | Google Scholar
2. Yang Y, Kim SC, Yu T, Yi YS, Rhee MH, Sung GH, et al. Functional roles of p38 mitogen-activated protein kinase in macrophage-mediated inflammatory responses. Mediators Inflamm. (2014) 2014:352371. doi: 10.1155/2014/352371
PubMed Abstract | Crossref Full Text | Google Scholar
3. Zarubin T and Han J. Activation and signaling of the p38 MAP kinase pathway. Cell Res. (2005) 15:11–8. doi: 10.1038/sj.cr.7290257
PubMed Abstract | Crossref Full Text | Google Scholar
4. Zhang J, Shen B, and Lin A. Novel strategies for inhibition of the p38 MAPK pathway. Trends Pharmacol Sci. (2007) 28:286–95. Available online at: https://www-sciencedirect-com.libproxy1.nus.edu.sg/science/article/pii/S0165614707000879 (Accessed April 25, 2025).
Google Scholar
5. Searle E, Tremble L, Popat R, de Bruin D, Moerland M, and Buckley B. Polb 001, an oral broad-spectrum anti-inflammatory with the potential to prevent cytokine release syndrome (CRS). Blood. (2023) 142:2093. doi: 10.1182/blood-2023-189062
Crossref Full Text | Google Scholar
6. Moerland M, Kales AJ, Broekhuizen K, Nässander U, Nelissen R, de Kam ML, et al. Proof of pharmacology of Org 48775-0, a p38 MAP kinase inhibitor, in healthy volunteers. Br J Clin Pharmacol. (2021) 87:2321–32. doi: 10.1111/bcp.14655
PubMed Abstract | Crossref Full Text | Google Scholar
7. Dillingh MR, van Poelgeest EP, Malone KE, Kemper EM, Stroes ESG, Moerland M, et al. Characterization of inflammation and immune cell modulation induced by low-dose LPS administration to healthy volunteers. J Inflammation. (2014) 11:28. doi: 10.1186/s12950-014-0028-1
Crossref Full Text | Google Scholar
8. Buters TP, Hameeteman PW, Jansen IME, van Hindevoort FC, Ten Voorde W, Florencia E, et al. Intradermal lipopolysaccharide challenge as an acute in vivo inflammatory model in healthy volunteers. Br J Clin Pharmacol. (2022) 88:680–90. doi: 10.1111/bcp.14999
PubMed Abstract | Crossref Full Text | Google Scholar
9. van Poelgeest EP, Dillingh MR, de Kam M, Malone KE, Kemper M, Stroes ESG, et al. Characterization of immune cell, endothelial, and renal responses upon experimental human endotoxemia. J Pharmacol Toxicol Methods. (2018) 89:39–46. Available online at: https://www-sciencedirect-com.libproxy1.nus.edu.sg/science/article/pii/S1056871917300795 (Accessed January 09, 2025).
PubMed Abstract | Google Scholar
10. Buters TP, Hameeteman PW, Jansen IME, van Hindevoort FC, Ten Voorde W, Grievink HW, et al. Clinical, cellular, and molecular effects of corticosteroids on the response to intradermal lipopolysaccharide administration in healthy volunteers. Clin Pharmacol Ther. (2022) 111:964–71. doi: 10.1002/cpt.2516
PubMed Abstract | Crossref Full Text | Google Scholar
11. Monnet E, Lapeyre G, van Poelgeest E, Jacqmin P, de Graaf K, Reijers J, et al. Evidence of NI-0101 pharmacological activity, an anti-TLR4 antibody, in a randomized phase I dose escalation study in healthy volunteers receiving LPS. Clin Pharmacol Ther. (2017) 101:200–8. doi: 10.1002/cpt.522
PubMed Abstract | Crossref Full Text | Google Scholar
12. Fijen JW, Zijlstra JG, De Boer P, Spanjersberg R, Tervaert JW, van der Werf TS, et al. Suppression of the clinical and cytokine response to endotoxin by RWJ-67657, a p38 mitogen-activated protein-kinase inhibitor, in healthy human volunteers. Clin Exp Immunol. (2001) 124:16–20. doi: 10.1046/j.1365-2249.2001.01485.x
PubMed Abstract | Crossref Full Text | Google Scholar
13. Faas MM, Moes H, Fijen JW, Muller Kobold AC, Tulleken JE, and Zijlstra JG. Monocyte intracellular cytokine production during human endotoxaemia with or without a second in vitro LPS challenge: effect of RWJ-67657, a p38 MAP-kinase inhibitor, on LPS-hyporesponsiveness. Clin Exp Immunol. (2002) 127:337–43. doi: 10.1046/j.1365-2249.2002.01765.x
PubMed Abstract | Crossref Full Text | Google Scholar
14. Fijen JW, Tulleken JE, Kobold ACM, de Boer P, van der Werf TS, Ligtenberg JJM, et al. Inhibition of p38 mitogen-activated protein kinase: dose-dependent suppression of leukocyte and endothelial response after endotoxin challenge in humans. Crit Care Med. (2002) 30:841–5. doi: 10.1097/00003246-200204000-00021
PubMed Abstract | Crossref Full Text | Google Scholar
15. Singh D, Siew L, Christensen J, Plumb J, Clarke GW, Greenaway S, et al. Oral and inhaled p38 MAPK inhibitors: effects on inhaled LPS challenge in healthy subjects. Eur J Clin Pharmacol. (2015) 71:1175–84. doi: 10.1007/s00228-015-1920-1
PubMed Abstract | Crossref Full Text | Google Scholar
16. Pehrson R, Hegelund-Myrbäck T, Cunoosamy D, Asimus S, Jansson P, Patel N, et al. AZD7624, an Inhaled p38 Inhibitor, Demonstrates Local Lung Inhibition of LPS-Induced TNFα with Minimal Systemic Exposure. J Pharmacol Exp Ther. (2018) 365:567–72. Available online at: https://www-sciencedirect-com.libproxy1.nus.edu.sg/science/article/pii/S0022356524267884 (Accessed July 31, 2025).
PubMed Abstract | Google Scholar
17. Prins ND, Harrison JE, Chu HM, Blackburn K, Alam JJ, and Scheltens P. A phase 2 double-blind placebo-controlled 24-week treatment clinical study of the p38 alpha kinase inhibitor neflamapimod in mild Alzheimer’s disease. Alzheimers Res Ther. (2021) 13:106. doi: 10.1186/s13195-021-00843-2
PubMed Abstract | Crossref Full Text | Google Scholar
18. Jiang Y, Alam JJ, Gomperts SN, Maruff P, Lemstra AW, Germann UA, et al. Preclinical and randomized clinical evaluation of the p38α kinase inhibitor neflamapimod for basal forebrain cholinergic degeneration. Nat Commun. (2022) 13:5308. doi: 10.1038/s41467-022-32944-3
PubMed Abstract | Crossref Full Text | Google Scholar
19. Augustin R, Poklepovic A, Gao X, Gooding W, Brodeur S, Urban J, et al. 588 Phase Ib study of the p38 inhibitor ARRY-614 with nivolumab, ipilimumab or nivolumab+ipilimumab in advanced solid tumors. J Immunother Cancer. (2022) 10. Available online at: https://jitcsite-bmj.vercel.app/content/10/Suppl_2/A615 (Accessed July 31, 2025).
Google Scholar
20. MacNee W, Allan RJ, Jones I, De Salvo MC, and Tan LF. Efficacy and safety of the oral p38 inhibitor PH-797804 in chronic obstructive pulmonary disease: a randomised clinical trial. Thorax. (2013) 68:738 LP – 74. Available online at: http://thorax.bmj.com.libproxy1.nus.edu.sg/content/68/8/738.abstract (Accessed July 23, 2025).
PubMed Abstract | Google Scholar
21. Patnaik A, Haluska P, Tolcher AW, Erlichman C, Papadopoulos KP, Lensing JL, et al. A first-in-human phase I study of the oral p38 MAPK inhibitor, ralimetinib (LY2228820 dimesylate), in patients with advanced cancer. Clin Cancer Res an Off J Am Assoc Cancer Res. (2016) 22:1095–102. doi: 10.1158/1078-0432.CCR-15-1718
PubMed Abstract | Crossref Full Text | Google Scholar
22. Vergote I, Heitz F, Buderath P, Powell M, Sehouli J, Lee CM, et al. A randomized, double-blind, placebo-controlled phase 1b/2 study of ralimetinib, a p38 MAPK inhibitor, plus gemcitabine and carboplatin versus gemcitabine and carboplatin for women with recurrent platinum-sensitive ovarian cancer. Gynecol Oncol. (2020) 156:23–31. doi: 10.1016/j.ygyno.2019.11.006
PubMed Abstract | Crossref Full Text | Google Scholar
23. Biau J, Thivat E, Chautard E, Stefan D, Boone M, Chauffert B, et al. Phase 1 trial of ralimetinib (LY2228820) with radiotherapy plus concomitant temozolomide in the treatment of newly diagnosed glioblastoma. Radiother Oncol J Eur Soc Ther Radiol Oncol. (2021) 154:227–34. doi: 10.1016/j.radonc.2020.09.036
PubMed Abstract | Crossref Full Text | Google Scholar
24. Bhattacharjee D, Bakar J, Chitnis SP, Sausville EL, Ashtekar KD, Mendelson BE, et al. Inhibition of a lower potency target drives the anticancer activity of a clinical p38 inhibitor. Cell Chem Biol. (2023) 30:1211–22.e5. Available online at: https://www-sciencedirect-com.libproxy1.nus.edu.sg/science/article/pii/S245194562300332X (Accessed October 30, 2024).
PubMed Abstract | Google Scholar
25. Singh D, Smyth L, Borrill Z, Sweeney L, and Tal-Singer R. A randomized, placebo-controlled study of the effects of the p38 MAPK inhibitor SB-681323 on blood biomarkers of inflammation in COPD patients. J Clin Pharmacol. (2010) 50:94–100. doi: 10.1177/0091270009347873
PubMed Abstract | Crossref Full Text | Google Scholar
26. Betts JC, Mayer RJ, Tal-Singer R, Warnock L, Clayton C, Bates S, et al. Gene expression changes caused by the p38 MAPK inhibitor dilmapimod in COPD patients: analysis of blood and sputum samples from a randomized, placebo-controlled clinical trial. Pharmacol Res Perspect. (2015) 3:e00094. doi: 10.1002/prp2.94
PubMed Abstract | Crossref Full Text | Google Scholar
27. Christie JD, Vaslef S, Chang PK, May AK, Gunn SR, Yang S, et al. A Randomized Dose-Escalation Study of the Safety and Anti-Inflammatory Activity of the p38 Mitogen-Activated Protein Kinase Inhibitor Dilmapimod in Severe Trauma Subjects at Risk for Acute Respiratory Distress Syndrome. Crit Care Med. (2015) 43:1859–69. Available online at: https://journals-lww-com.libproxy1.nus.edu.sg/ccmjournal/fulltext/2015/09000/a_randomized_dose_escalation_study_of_the_safety.11.aspx (Accessed July 03, 2024).
PubMed Abstract | Google Scholar
28. Anand P, Shenoy R, Palmer JE, Baines AJ, Lai RYK, Robertson J, et al. Clinical trial of the p38 MAP kinase inhibitor dilmapimod in neuropathic pain following nerve injury. Eur J Pain. (2011) 15:1040–8. doi: 10.1016/j.ejpain.2011.04.005
PubMed Abstract | Crossref Full Text | Google Scholar
29. Fisk M, Cheriyan J, Mohan D, Forman J, Mäki-Petäjä KM, McEniery CM, et al. The p38 mitogen activated protein kinase inhibitor losmapimod in chronic obstructive pulmonary disease patients with systemic inflammation, stratified by fibrinogen: A randomised double-blind placebo-controlled trial. PloS One. (2018) 13:e0194197. doi: 10.1371/journal.pone.0194197
PubMed Abstract | Crossref Full Text | Google Scholar
30. Emami H, Vucic E, Subramanian S, Abdelbaky A, Fayad ZA, Du S, et al. The effect of BMS-582949, a P38 mitogen-activated protein kinase (P38 MAPK) inhibitor on arterial inflammation: a multicenter FDG-PET trial. Atherosclerosis. (2015) 240:490–6. doi: 10.1016/j.atherosclerosis.2015.03.039
PubMed Abstract | Crossref Full Text | Google Scholar
31. Damjanov N, Kauffman RS, and Spencer-Green GT. Efficacy, pharmacodynamics, and safety of VX-702, a novel p38 MAPK inhibitor, in rheumatoid arthritis: results of two randomized, double-blind, placebo-controlled clinical studies. Arthritis Rheum. (2009) 60:1232–41. doi: 10.1002/art.24485
PubMed Abstract | Crossref Full Text | Google Scholar
32. Cohen SB, Cheng TT, Chindalore V, Damjanov N, Burgos-Vargas R, Delora P, et al. Evaluation of the efficacy and safety of pamapimod, a p38 MAP kinase inhibitor, in a double-blind, methotrexate-controlled study of patients with active rheumatoid arthritis. Arthritis Rheum. (2009) 60:335–44. doi: 10.1002/art.24266
PubMed Abstract | Crossref Full Text | Google Scholar
33. Goldman JW, Rosen LS, Tolcher AW, Papadopoulos K, Beeram M, Shi P, et al. Phase 1 and pharmacokinetic study of LY3007113, a p38 MAPK inhibitor, in patients with advanced cancer. Invest New Drugs. (2018) 36:629–37. doi: 10.1007/s10637-017-0532-2
PubMed Abstract | Crossref Full Text | Google Scholar
Keywords: Cytokine release syndrome (CRS), in vivo, inflammation, LPS, p38 MAPK
Citation: de Bruin DT, Jansen MAA, Pereira DR, van Schijndel L, Klaassen ES, Otto ME, Skillington J, Maguire P, Tremble L, Bell A, Maher L, Mihara K, Sumeray M, Gilroy DW, Klarenbeek NB and Moerland M (2026) POLB 001, a p38 MAPK inhibitor, decreases local and systemic inflammatory responses following in vivo LPS administration in healthy volunteers: a randomised, double-blind, placebo-controlled study. Front. Immunol. 16:1684307. doi: 10.3389/fimmu.2025.1684307
Received: 12 August 2025; Accepted: 30 December 2025; Revised: 15 December 2025;Published: 23 January 2026.
Edited by:
Reviewed by:
Copyright © 2026 de Bruin, Jansen, Pereira, van Schijndel, Klaassen, Otto, Skillington, Maguire, Tremble, Bell, Maher, Mihara, Sumeray, Gilroy, Klarenbeek and Moerland. This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.
*Correspondence: Matthijs Moerland, mmoerland@chdr.nl
