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Introduction: Cigarette affordability, the price of tobacco relative to consumer income, is a key determinant of tobacco consumption. This study examined trends over 12 years in individualised factory-made cigarette affordability in the Netherlands, and whether these trends differed by sex, age, and education.Methods: Data from 10 waves (2008-2020) of the International Tobacco Control (ITC) Netherlands Surveys were used to estimate individualised affordability, measured as the percentage of income required to buy 100 cigarette packs (Relative Income Price, RIP), using self-reported prices and income. The higher the RIP, the less affordable cigarettes are. Generalised estimating equation regression models assessed trends in individualised affordability over time and by sex, age, and education.Results: Affordability decreased significantly between 2008 and 2020, with RIP increasing from 1.89% (2008) to 2.64% (2020) (p≤.001), Except for 2008-2010, no significant year-on-year changes in affordability were found. Lower affordability was found among subgroups who have a lower income level: females (versus males), 18-24 and 25-39 year olds (versus 55 and over) and low or moderate educated individuals (versus high educated). Interactions between wave and education (p=.007) were found, but not with sex (p=.653) or age (p=.295). A decreasing linear trend in affordability was found for moderately (p=.041) and high-educated (p=.025), but not for low-educated individuals (p=.149).Conclusions: Cigarettes in the Netherlands have become less affordable between 2008 and 2020, yet this was mostly due to the decrease in affordability between 2008 and 2010. There is a need for more significant increases in tax to further decrease affordability.Implications: Our findings suggest that cigarettes have become less affordable in the Netherlands between 2008 and 2020. But, this appears to be the result of a steep decrease in affordability between 2008 and 2010. Affordability was lower among groups who have on average lower incomes (females, young adults, low- and moderate educated individuals), and differences in trends across education levels could be explained by per capita income changes. Our individualised measure indicated lower affordability than published aggregate affordability estimations. Future tax increases should be large enough to result in a lower affordability. [ABSTRACT FROM AUTHOR]

Purpose: Immunotherapies have shown limited responses in patients with advanced pancreatic cancer. Recently, we reported that dendritic cell (DC)-based immunotherapy induced T-cell responses against pancreatic cancer antigens. The primary objective of this study was to determine the efficacy of DC-based immunotherapy to prevent recurrence of disease., Methods: This was a single-center, open-label, single-arm, combined phase I/II trial. The primary end point was the 2-year recurrence-free survival (RFS) rate. A 2-year RFS rate of ≥60% was defined as a clinically meaningful improvement. We included patients with pancreatic cancer after resection and completion of standard-of-care (SOC) treatment without recurrent disease on cross-sectional imaging. Patients were treated with autologous DCs pulsed with an allogeneic mesothelioma tumor cell lysate, comprising antigens also expressed in pancreatic ductal adenocarcinoma., Results: Thirty-eight patients were included in the analysis of the primary end point (47% male, 53% female). The median age was 62 years (IQR, 55-68). Twenty-eight patients (74%) received five DC vaccinations and completed the study protocol. Three patients (8%) received four vaccinations, and seven patients (16%) received three vaccinations. After a median follow-up of 25.5 months, 26 patients (68%) had not developed recurrence of disease. The estimated 2-year RFS was 64%. Vaccination led to the enrichment of circulating activated CD4+ T cells and the detection of treatment-induced immune responses in vitro. T-cell receptor-sequencing analyses of a resected solitary lung metastasis showed influx of vaccine-specific T cells., Conclusion: This study reached its primary end point of a 2-year RFS rate of ≥60% following pancreatectomy after SOC treatment and adjuvant DC-based immunotherapy in patients with pancreatic cancer. These results warrant a future randomized trial.

Purpose: Amid the need for new approaches to improve survival in pancreatic ductal adenocarcinoma (PDAC), immune-based therapies have garnered interest. Rintatolimod, a Toll-like receptor 3 (TLR-3) agonist, is a potential candidate due to its dual impact on restraining PDAC cell functions and boosting the antitumor immune response. This study investigates the effect of TLR-3 activation through rintatolimod on the peripheral immune landscape of patients with advanced PDAC., Experimental Design: Paired blood samples of 30 patients with advanced PDAC, collected at baseline and after 12 rintatolimod intravenous infusions, underwent comprehensive transcriptomic NanoString and proteomic flow cytometry profiling. The impact of rintatolimod and immunologic factors on survival outcomes was assessed through univariate Cox proportional hazards models., Results: Rintatolimod treatment enhances peripheral immune activity at the transcriptomic and proteomic levels, particularly involving type 1 conventional dendritic cells (cDC1) and T cells. Post-rintatolimod, the increased peripheral abundance of BTLA+ XCR1+ cDC1s and CD4+SELL+ T cells correlated with improved clinical outcomes. Patients with stable disease exhibited pronouncedDCand T-cell activation gene overexpression. Notably, the expression of immune checkpoints PD-L1 and PD-L2 decreased post-rintatolimod across all patients. However, those with progressive disease showed increased expression of genes encoding IDO1 and PD-1., Conclusions: This study presents compelling evidence of the immune-stimulatory properties linked to TLR-3 activation through rintatolimod. Rintatolimod may break immunologic tolerance by enhancing antitumor immunity through DC-mediated Th-cell responses. Furthermore, our findings lay the groundwork for investigating the potential synergy between TLR-3 activation and immune checkpoint inhibitor therapy to improve therapeutic outcomes. See related commentary by Martínez-Riaño et al., p. 3355., (©2024 American Association for Cancer Research.)

Background: Dendritic cell immunotherapy has proven to be safe and induces an immune response in humans. We aimed to establish the efficacy of dendritic cells loaded with allogeneic tumour cell lysate (MesoPher, Amphera BV, 's-Hertogenbosch, Netherlands) as maintenance therapy in patients with pleural mesothelioma., Methods: In this open-label, randomised, phase 2/3 study, patients with histologically confirmed unresectable pleural mesothelioma, aged 18 years or older, with an Eastern Cooperative Oncology Group performance status score of 0-1, and non-progressing disease after four to six cycles of standard chemotherapy (with pemetrexed 500 mg/m 2 plus platinum [cisplatin 75 mg/m 2 or carboplatin area under the curve of 5]) were recruited from four centres in Belgium, France, and The Netherlands. Participants were randomly assigned (1:1), using block randomisation (block size of 4), stratified by centre and histology (epithelioid vs other), to MesoPher treatment plus best supportive care or best supportive care alone. Patients received up to a maximum of five MesoPher infusions, with treatment administered on days 1, 15, and 29, and weeks 18 and 30. At each timepoint, participants received an injection of 25 × 10 6 dendritic cells (two-thirds of the dendritic cells were administered intravenously and a third were injected intradermally). Best supportive care was per local institutional standards. The primary endpoint was overall survival, assessed in all participants randomly assigned to treatment (full analysis set) and safety assessed in all randomly assigned participants, and who underwent leukapheresis if they were in the MesoPher group. This study is registered with ClinicalTrials.gov, NCT03610360, and is closed for accrual., Findings: Between June 21, 2018, and June 10, 2021, 176 patients were screened and randomly assigned to the MesoPher group (n=88) or best supportive care alone group (n=88). One participant in the MesoPher group did not undergo leukapheresis. Mean age was 68 years (SD 8), 149 (85%) of 176 were male, 27 (15%) were female, 173 (98%) were White, two were Asian (1%), and one (1%) was other race. As of data cutoff (June 24, 2023), after a median follow up of 15·1 months (IQR 9·5-22·4), median overall survival was 16·8 months (95% CI 12·4-20·3; 61 [69%] of 88 died) in the MesoPher group and 18·3 months (14·3-21·9; 59 [67%] of 88 died) in the best supportive care group (hazard ratio 1·10 [95% CI 0·77-1·57]; log-rank p=0·62). The most common grade 3-4 treatment-emergent adverse events were chest pain (three [3%] of 87 in the MesoPher group vs two [2%] of 88 in the best supportive care group), dyspnoea (none vs two [2%]), anaemia (two [2%] vs none), nausea (none vs two [2%]), and pneumonia (none vs two [2%]). No deaths due to treatment-emergent adverse events were recorded. Treatment-related adverse events consisted of infusion-related reactions (fever, chills, and fatigue), which occurred in 64 (74%) of 87 patients in the MesoPher group, and injection-site reactions (itch, erythema, and induration), which occurred in 73 (84%) patients, and all were grade 1-2 in severity. No deaths were determined to be treatment related., Interpretation: MesoPher did not show improvement in overall survival in patients with pleural mesothelioma. Immune checkpoint therapy is now standard of care in pleural mesothelioma. Further randomised studies are needed of combinations of MesoPher and immune checkpoint therapy, which might increase efficacy without adding major toxicities., Funding: Amphera BV and EU HORIZON., Competing Interests: Declaration of interests JGA reports grants, personal fees, research support, and stock ownership from Amphera and personal fees from Bristol-Myers Squibb, Eli-Lilly, MSD, CureVac, and Novocure. JGA has a patent for a tumour cell lysate licensed, a patent for a combination immunotherapy licensed, and a patent for a biomarker for immunotherapy licensed. RC reports personal fees from Janssen, MSD, Spectrum, and Librerium. PB reports receiving consulting fees for advisorships from AstraZeneca and Bristol Myers Squibb. AS reports personal fees and non-financial support from AstraZeneca, Bristol Myers Squibb, and MSD, of which the principal investigator fees directly went to their institution (CHRU Lille). DF reports personal fees from Bayer, Boehringer Ingelheim, Bristol Myers Squibb, and AstraZeneca; personal fees and travel bursary from RS Oncology; and research funding from Astex Pharmaceuticals, Bergen Bio, Iovance, Owkin, GSK, MSD Oncology, Roche UK, Clovis Oncology, and Aldeyra. KB reports personal fees from Amphera. RK reports that he received fees from Amphera BV for statistical services related to the study. RBer reports personal fees from Boehringer Ingelheim, Otsuka, Amgen, Lilly, GSK, EISAI, and Gilead; grants from Astra Zeneca, Roche, andNovartis; and grants and personal fees from MSD. RM and IE received consulting fees from Amphera BV and are stock owner of Amphera BV. All other authors declare no competing interests., (Copyright © 2024 Elsevier Ltd. All rights reserved, including those for text and data mining, AI training, and similar technologies.)

Objectives: Female-specific issues, such as breast injuries, pelvic floor dysfunctions, saddle sores, and menstrual symptoms, can significantly impact female athletes. This study examined the prevalence, perceived impact on performance, and role of the support staff in addressing and managing these issues in elite athletes based at a multisport training centre., Design: Anonymous online questionnaire., Methods: A total of 180 female athletes from various sport disciplines at the Dutch Olympic Training Centre were invited to participate in the study. An anonymous, 39-item questionnaire was developed and hosted on Qualtrics collecting data on respondents' characteristics, breast injuries, pelvic floor dysfunctions, saddle sores, menstrual symptoms, contraceptive use, and communication about these issues., Results: A total of 105 valid responses were analysed. The findings revealed that athletes regularly experience breast injuries (15.2 %), urinary incontinence (29.5 %), other pelvic floor problems (21.0 %), saddle sores (65.7 %), and menstrual cycle-related symptoms, with negative performance effects. Communication with support staff remained limited, with only 23.8 % of athletes discussing any of these issues, often due to concerns about understanding, shame, or a lack of opportunity for easy communication., Conclusions: The findings underscore the crucial role of creating a supportive environment, promoting early intervention, and utilising multisport centres to comprehensively address these concerns, emphasising the need for open communication, education, and support to enhance female athletes' performance and well-being., Competing Interests: Declaration of interest statement The authors have no competing interest to declare., (Copyright © 2024 The Authors. Published by Elsevier Ltd.. All rights reserved.)

The prevalence of comorbidities in individuals with neurodevelopmental disorders (NDDs) is not well understood, yet these are important for accurate diagnosis and prognosis in routine care and for characterizing the clinical spectrum of NDD syndromes. We thus developed PhenomAD-NDD, an aggregated database containing the comorbid phenotypic data of 51,227 individuals with NDD, all harmonized into Human Phenotype Ontology (HPO), with in total 3,054 unique HPO terms. We demonstrate that almost all congenital anomalies are more prevalent in the NDD population than in the general population, and the NDD baseline prevalence allows for an approximation of the enrichment of symptoms. For example, such analyses of 33 genetic NDDs show that 32% of enriched phenotypes are currently not reported in the clinical synopsis in the Online Mendelian Inheritance in Man (OMIM). PhenomAD-NDD is open to all via a visualization online tool and allows us to determine the enrichment of symptoms in NDD., (© 2024. The Author(s), under exclusive licence to Springer Nature America, Inc.)

Recently, OTULIN haploinsufficiency was linked to enhanced susceptibility to Staphylococcus aureus infections accompanied by local necrosis and systemic inflammation. The pathogenesis observed in haploinsufficient patients differs from the hyperinflammation seen in classical OTULIN-related autoinflammatory syndrome (ORAS) patients and is characterized by increased susceptibility of dermal fibroblasts to S. aureus alpha toxin-inflicted cytotoxic damage. Immunological abnormalities were not observed in OTULIN haploinsufficient patients, suggesting a non-hematopoietic basis. In this research report, we investigated an Otulin +/- mouse model after in vivo provocation with lipopolysaccharide (LPS) to explore the potential role of hematopoietic-driven inflammation in OTULIN haploinsufficiency. We observed a hyperinflammatory signature in LPS-provoked Otulin +/- mice, which was driven by CD64 + monocytes and macrophages. Bone marrow-derived macrophages (BMDMs) of Otulin +/- mice demonstrated higher proinflammatory cytokine secretion after in vitro stimulation with LPS or polyinosinic:polycytidylic acid (Poly(I:C)). Our experiments in full and mixed bone marrow chimeric mice suggest that, in contrast to humans, the observed inflammation was mainly driven by the hematopoietic compartment with cell-extrinsic effects likely contributing to inflammatory outcomes. Using an OTULIN haploinsufficient mouse model, we validated the role of OTULIN in the regulation of environmentally directed inflammation., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2024 Staels, Bücken, De Vuyst, Willemsen, Van Nieuwenhove, Gerbaux, Neumann, Malviya, Van Meerbeeck, Haughton, Seldeslachts, Gouwy, Martinod, Vande Velde, Proost, Yshii, Schlenner, Schrijvers, Liston and Humblet-Baron.)

Introduction: While secondhand smoke exposure in outdoor spaces has been investigated before, no data on outdoor secondhand smoke exposure have been collected in the Netherlands. Such data could help policymakers gain support for smoke-free outdoor public spaces., Methods: Between May and November 2021, we visited 25 outdoor locations across the Netherlands. At each location, we conducted four measurements with smokers and one measurement without smokers. During each measurement, we counted the number of smokers present and we rated tobacco smell intensity on a five-point scale. Airborne nicotine and 3-ethenylpyridine (3-EP) data were collected through active sampling on thermal desorption tubes. The contents of these tubes were later analyzed using gas chromatography-mass spectrometry. Using linear mixed models, we investigated the association between levels of nicotine and the presence of smokers, the number of smokers, and the intensity of tobacco smell. We also investigated these association with levels of 3-EP., Results: Nicotine levels were higher when smokers were present (B=1.40; 95% CI: 0.69-2.11, p<0.001). For each additional smoker present, we measured higher levels of nicotine (B=0.23; 95% CI: 0.10-0.37, p=0.001). When the smell of tobacco smoke was noted to be stronger by the researchers, higher levels of nicotine were measured through sampling (B=0.85; 95% CI: 0.44-1.26, p<0.001). We found similar results for 3-EP levels., Conclusions: This study showed that both nicotine and 3-EP are useful in quantifying levels of secondhand smoke in various outdoor locations. The level of nicotine exposure outdoors was positively associated with the number of smokers nearby. The intensity of the tobacco smell was also related to nicotine exposure: the stronger the smell of tobacco smoke, the more nicotine was measured in the air., Competing Interests: The authors have completed and submitted the ICMJE Form for Disclosure of Potential Conflicts of Interest. The authors declare that they have no competing interests, financial or otherwise, related to the current work. All authors report that since the initial planning of the work J. Bommelé received a joint grant from the Dutch Heart Foundation, Lung Foundation Netherlands, Dutch Cancer Society, Dutch Diabetes Research Foundation and Netherlands Thrombosis Foundation (2.1.19.001), and a grant from the Netherlands Ministry of Health, Welfare and Sport (329845)., (© 2024 Bommelé J. et al.)

Sabin Inactivated Poliovirus Vaccine (sIPV) has become one of the preferred vaccination options for the last step in the Poliovirus eradication program. Sequencing of poliovirus samples is needed during the manufacturing of poliovirus vaccines to assure the safety and immunogenicity of these vaccines. Next-generation sequencing analysis is the current costly and time-consuming gold standard for monitoring the manufacturing processes. We developed a low-cost and quick, highly sensitive, and allele-specific locked nucleic acid-probe-based reverse transcription quantitative PCR alternative that can accurately detect mutations in poliovirus vaccine samples during process development, scaling up, and release. Using the frequently in vitro occurring and viral replication-impacting VP1-E 295 K mutation as a showcase, we show that this technology can accurately detect E 295 K mutations in poliovirus 2 samples to similar levels as NGS. The qPCR technology was developed employing a synthetic dsDNA fragment-based standard curve containing mixes of E 295 K-WT (wildtype) and Mut (mutant) synthetic dsDNA fragments ranging from 1 × 10 7 copies/µL to 1 × 10 2 copies/µL to achieve a linear correlation with R 2  > 0.999, and PCR efficiencies of 95-105 %. Individual standard concentration levels achieved accuracies of ≥92 % (average 96 %) and precisions of ≤17 % (average 3.3 %) RSD. Specificity of locked nucleic acid (LNA)-probes was confirmed in the presence and absence of co-mutations in the probe-binding region. Application of the developed assay to Sabin Poliovirus type 2 production run samples, illustrated a linear relationship with an R 2 of 0.994, and an average accuracy of 97.2 % of the variant (allele)-specific AS LNA qPCR result, compared to NGS. The assay showed good sensitivity for poliovirus samples, containing E 295 K mutation levels between 0 % and 95 % (quantification range). In conclusion, the developed AS LNA qPCR presents a valuable low-cost, and fast tool, suitable for the process development and quality control of polio vaccines., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Author(s). Published by Elsevier Ltd.. All rights reserved.)

Background: FOLFIRINOX chemotherapy has improved outcomes for pancreatic cancer patients, but poor long-term survival outcomes and high toxicity remain challenges. This study investigates the impact of FOLFIRINOX on plasma proteins and peripheral immune cells to guide immune-based combination therapies and, ideally, to identify a potential biomarker to predict early disease progression during FOLFIRINOX., Methods: Blood samples were collected from 86 pancreatic cancer patients before and two weeks after the first FOLFIRINOX cycle and subjected to comprehensive immune cell and proteome profiling. Principal Component Analysis and Linear Mixed Effect Regression models were used for data analysis. FOLFIRINOX efficacy was radiologically evaluated after the fourth cycle., Results: One cycle of FOLFIRINOX diminished tumour-cell-related pathways and enhanced pathways related to immune activation, illustrated by an increase in pro-inflammatory IL-18, IL-15, and TNFRSF4. Similarly, FOLFIRINOX promoted the activation of CD4 + and CD8 + T cells, the proliferation of NK(T), and the activation of antigen-presenting cells. Furthermore, high pre-treatment levels of VEGFA and PRDX3 and an elevation in FCRL3 levels after one cycle predicted early progression under FOLFIRINOX. Finally, patients with progressive disease exhibited high levels of inhibitory markers on B cells and CD8 + T cells, while responding patients exhibited high levels of activation markers on CD4 + and CD8 + T cell subsets., Conclusion: FOLFIRINOX has immunomodulatory effects, providing a foundation for clinical trials exploring immune-based combination therapies that harness the immune system to treat pancreatic cancer. In addition, several plasma proteins hold potential as circulating predictive biomarkers for early prediction of FOLFIRINOX response in patients with pancreatic cancer., Competing Interests: Declaration of Competing Interest J.G.J.V. Aerts reports personal fees and nonfinancial support from MSD, personal fees from BMS, Boehringer Ingelheim, Amphera, Eli‐Lilly, Takeda, Bayer, Roche, Astra Zeneca outside the submitted work; in addition, J.G.J.V. Aerts holds ownership interest (including patients) in Amphera BV and is a consultant/advisory board member for Amphera. The other authors declare that they have no competing interests., (Copyright © 2023 The Authors. Published by Elsevier Ltd.. All rights reserved.)

The DNA polymerase δ complex (PolD), comprising catalytic subunit POLD1 and accessory subunits POLD2, POLD3, and POLD4, is essential for DNA synthesis and is central to genome integrity. We identified, by whole exome sequencing, a homozygous missense mutation (c.1118A > C; p.K373T) in POLD3 in a patient with Omenn syndrome. The patient exhibited severely decreased numbers of naïve T cells associated with a restricted T-cell receptor repertoire and a defect in the early stages of TCR recombination. The patient received hematopoietic stem cell transplantation at age 6 months. He manifested progressive neurological regression and ultimately died at age 4 years. We performed molecular and functional analysis of the mutant POLD3 and assessed cell cycle progression as well as replication-associated DNA damage. Patient fibroblasts showed a marked defect in S-phase entry and an enhanced number of double-stranded DNA break-associated foci despite normal expression levels of PolD components. The cell cycle defect was rescued by transduction with WT POLD3. This study validates autosomal recessive POLD3 deficiency as a novel cause of profound T-cell deficiency and Omenn syndrome., (© 2023. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Pancreatic ductal adenocarcinoma (PDAC) remains one of the deadliest solid tumors and is resistant to immunotherapy. B cells play an essential role in PDAC progression and immune responses, both locally and systemically. Moreover, increasing evidence suggests that microbial compositions inside the tumor, as well as in the oral cavity and the gut, are important factors in shaping the PDAC immune landscape. However, the gut-associated lymphoid tissue (GALT) has not previously been explored in PDAC patients. In this study, we analyzed healthy vermiform appendix (VA) from 20 patients with PDAC and 32 patients with colon diseases by gene expression immune profiling, flow cytometry analysis, and microbiome sequencing. We show that the VA GALT of PDAC patients exhibits markers of increased inflammation and cytotoxic cell activity. In contrast, B cell function is decreased in PDAC VA GALT based on gene expression profiling; B cells express significantly fewer MHC class II surface receptors, whereas plasma cells express the immune checkpoint molecule HLA-G. Additionally, the vermiform appendix microbiome of PDAC patients is enriched with Klebsiella pneumoniae , Bifidobacterium animalis , and Adlercreutzia equolifaciens , while certain commensals are depleted. Our findings may suggest impaired B cell function within the GALT of PDAC patients, which could potentially be linked to microbial dysbiosis. Additional investigations are imperative to validate our observations and explore these potential targets of future therapies., Competing Interests: Author AO was employed by the company Pathan BV. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Vietsch, Latifi, Verheij, van der Oost, de Wilde, Haen, van den Boom, Koerkamp, Doornebosch, van Verschuer, Ooms, Mohammad, Willemsen, Aerts, Krog, de Miranda, van den Bosch, Mueller, Katsikis and van Eijck.)

Purpose: FOXP3 deficiency results in severe multisystem autoimmunity in both mice and humans, driven by the absence of functional regulatory T cells. Patients typically present with early and severe autoimmune polyendocrinopathy, dermatitis, and severe inflammation of the gut, leading to villous atrophy and ultimately malabsorption, wasting, and failure to thrive. In the absence of successful treatment, FOXP3-deficient patients usually die within the first 2 years of life. Hematopoietic stem cell transplantation provides a curative option but first requires adequate control over the inflammatory condition. Due to the rarity of the condition, no clinical trials have been conducted, with widely unstandardized therapeutic approaches. We sought to compare the efficacy of lead therapeutic candidates rapamycin, anti-CD4 antibody, and CTLA4-Ig in controlling the physiological and immunological manifestations of Foxp3 deficiency in mice., Method: We generated Foxp3-deficient mice and an appropriate clinical scoring system to enable direct comparison of lead therapeutic candidates rapamycin, nondepleting anti-CD4 antibody, and CTLA4-Ig., Results: We found distinct immunosuppressive profiles induced by each treatment, leading to unique protective combinations over distinct clinical manifestations. CTLA4-Ig provided superior breadth of protective outcomes, including highly efficient protection during the transplantation process., Conclusion: These results highlight the mechanistic diversity of pathogenic pathways initiated by regulatory T cell loss and suggest CTLA4-Ig as a potentially superior therapeutic option for FOXP3-deficient patients., (© 2023. The Author(s).)

Background: Malignant peritoneal mesothelioma (MPM) is an aggressive malignancy with a poor prognosis. Cytoreductive surgery (CRS) and hyperthermic intraperitoneal chemotherapy (HIPEC) improves survival outcomes, but recurrence rates remain high. Dendritic cell-based immunotherapy (DCBI) showed promising results in patients with pleural mesothelioma. The primary aim of this trial was to determine feasibility of adjuvant DCBI after CRS-HIPEC., Methods: This open-label, single-center, phase II clinical trial, performed in the Erasmus MC Cancer Institute Rotterdam, the Netherlands, included patients with epithelioid MPM. 4-6 weeks before CRS-HIPEC leukapheresis was performed. 8-10 weeks after surgery, DCBI was administered three times biweekly. Feasibility was defined as administration of at least three adjuvant vaccinations in 75% of patients. Comprehensive immune cell profiling was performed on peripheral blood samples prior to and during treatment., Results: All patients who received CRS-HIPEC (n=16) were successfully treated with adjuvant DCBI. No severe toxicity related to DCBI was observed. Median progression-free survival (PFS) was 12 months (IQR 5-23) and median overall survival was not reached. DCBI was associated with increased proliferation of circulating natural killer cells and CD4+ T-helper (Th) cells. Co-stimulatory molecules, including ICOS, HLA-DR, and CD28 were upregulated predominantly on memory or proliferating Th-cells and minimally on CD8+ cytotoxic T-lymphocytes (CTLs) after treatment. However, an increase in CD8+ terminally differentiated effector memory (Temra) cells positively correlated with PFS, whereas co-expression of ICOS and Ki67 on CTLs trended towards a positive correlation., Conclusions: Adjuvant DCBI after CRS-HIPEC in patients with MPM was feasible and safe, and showed promising survival outcomes. DCBI had an immune modulatory effect on lymphoid cells and induced memory T-cell activation. Moreover, an increase of CD8+ Temra cells was more pronounced in patients with longer PFS. These data provide rationale for future combination treatment strategies., Trial Registration Number: NTR7060; Dutch Trial Register (NTR)., Competing Interests: Competing interests: JGJVA declares consulting fees from MSD, BMS, Takeda, Amphera, AstraZeneca and Eli-Lilly, is a stock owner of Amphera and has intellectual property rights in Amphera and Pamgene., (© Author(s) (or their employer(s)) 2023. Re-use permitted under CC BY-NC. No commercial re-use. See rights and permissions. Published by BMJ.)

Background: Severe congenital neutropenia presents with recurrent infections early in life as a result of arrested granulopoiesis. Multiple genetic defects are known to block granulocyte differentiation; however, a genetic cause remains unknown in approximately 40% of cases., Objective: We aimed to characterize a patient with severe congenital neutropenia and syndromic features without a genetic diagnosis., Methods: Whole exome sequencing results were validated using flow cytometry, Western blotting, coimmunoprecipitation, quantitative PCR, cell cycle and proliferation analysis of lymphocytes and fibroblasts and granulocytic differentiation of primary CD34 + and HL-60 cells., Results: We identified a homozygous missense mutation in DBF4 in a patient with mild extra-uterine growth retardation, facial dysmorphism and severe congenital neutropenia. DBF4 is the regulatory subunit of the CDC7 kinase, together known as DBF4-dependent kinase (DDK), the complex essential for DNA replication initiation. The DBF4 variant demonstrated impaired ability to bind CDC7, resulting in decreased DDK-mediated phosphorylation, defective S-phase entry and progression and impaired differentiation of granulocytes associated with activation of the p53-p21 pathway. The introduction of wild-type DBF4 into patient CD34 + cells rescued the promyelocyte differentiation arrest., Conclusion: Hypomorphic DBF4 mutation causes autosomal-recessive severe congenital neutropenia with syndromic features., (Copyright © 2023 The Authors. Published by Elsevier Inc. All rights reserved.)

Invasive group A streptococcal (iGAS) disease cases increased in the first half of 2022 in the Netherlands, with a remarkably high proportion of emm 4 isolates. Whole-genome sequence analysis of 66 emm 4 isolates, 40 isolates from the pre-coronavirus disease 2019 (COVID-19) pandemic period 2009-2019 and 26 contemporary isolates from 2022, identified a novel Streptococcus pyogenes lineage (M4 NL22 ), which accounted for 85 % of emm 4 iGAS cases in 2022. Surprisingly, we detected few isolates of the emm 4 hypervirulent clone, which has replaced nearly all other emm 4 in the USA and the UK. M4 NL22 displayed genetic differences compared to other emm 4 strains, although these were of unclear biological significance. In publicly available data, we identified a single Norwegian isolate belonging to M4 NL22 , which was sampled after the isolates from this study, possibly suggesting export of M4 NL22 to Norway. In conclusion, our study identified a novel S. pyogenes emm 4 lineage underlying an increase of iGAS disease in early 2022 in the Netherlands and the results have been promptly communicated to public health officials.

Inborn errors of immunity are a heterogeneous group of monogenic immunologic disorders caused by mutations in genes with critical roles in the development, maintenance, or function of the immune system. The genetic basis is frequently a mutation in a gene with restricted expression and/or function in immune cells, leading to an immune disorder. Several classes of inborn errors of immunity, however, result from mutation in genes that are ubiquitously expressed. Despite the genes participating in cellular processes conserved between cell types, immune cells are disproportionally affected, leading to inborn errors of immunity. Mutations in DNA replication, DNA repair, or DNA damage response factors can result in monogenic human disease, some of which are classified as inborn errors of immunity. Genetic defects in the DNA repair machinery are a well-known cause of T - B - NK + severe combined immunodeficiency. An emerging class of inborn errors of immunity is those caused by mutations in DNA replication factors. Considerable heterogeneity exists within the DNA replication-associated inborn errors of immunity, with diverse immunologic defects and clinical manifestations observed. These differences are suggestive for differential sensitivity of certain leukocyte subsets to deficiencies in specific DNA replication factors. Here, we provide an overview of DNA replication-associated inborn errors of immunity and discuss the emerging mechanistic insights that can explain the observed immunologic heterogeneity., (Copyright © 2022 The Authors. Published by Elsevier Inc. All rights reserved.)

Calcium signaling is essential for lymphocyte activation, with genetic disruptions of store-operated calcium (Ca 2+ ) entry resulting in severe immunodeficiency. The inositol 1,4,5-trisphosphate receptor (IP 3 R), a homo- or heterotetramer of the IP 3 R1-3 isoforms, amplifies lymphocyte signaling by releasing Ca 2+ from endoplasmic reticulum stores following antigen stimulation. Although knockout of all IP 3 R isoforms in mice causes immunodeficiency, the seeming redundancy of the isoforms is thought to explain the absence of variants in human immunodeficiency. In this study, we identified compound heterozygous variants of ITPR3 (a gene encoding IP 3 R subtype 3) in two unrelated Caucasian patients presenting with immunodeficiency. To determine whether ITPR3 variants act in a nonredundant manner and disrupt human immune responses, we characterized the Ca 2+ signaling capacity, the lymphocyte response, and the clinical phenotype of these patients. We observed disrupted Ca 2+ signaling in patient-derived fibroblasts and immune cells, with abnormal proliferation and activation responses following T-cell receptor stimulation. Reconstitution of IP 3 R3 in IP 3 R knockout cell lines led to the identification of variants as functional hypomorphs that showed reduced ability to discriminate between homeostatic and induced states, validating a genotype-phenotype link. These results demonstrate a functional link between defective endoplasmic reticulum Ca 2+ channels and immunodeficiency and identify IP 3 Rs as diagnostic targets for patients with specific inborn errors of immunity. These results also extend the known cause of Ca 2+ -associated immunodeficiency from store-operated entry to impaired Ca 2+ mobilization from the endoplasmic reticulum, revealing a broad sensitivity of lymphocytes to genetic defects in Ca 2+ signaling., (© 2022. The Author(s).)

Purpose: Mendelian susceptibility to mycobacterial disease (MSMD) is caused by inborn errors of IFN-γ immunity. The most frequent genetic defects are found in IL12 or a subunit of its receptor. IL23R deficiency in MSMD has only been reported once, in two pediatric patients from the same kindred with isolated disseminated Bacille Calmette-Guérin disease. We evaluated the impact of a homozygous stop mutation in IL23R (R381X), identified by whole exome sequencing, in an adult patient with disseminated non-tuberculous mycobacterial disease., Methods: We performed functional validation of the R381X mutation by evaluating IL23R expression and IL-23 signaling (STAT3 phosphorylation, IFN-γ production) in primary cells (PBMCs, EBV-B cells) and cell lines (HeLa) with or without back-complementation of wild-type IL23R., Results: We report on a 48-year-old male with disseminated non-tuberculous mycobacterial disease. We identified and characterized a homozygous loss-of-function stop mutation underlying IL23R deficiency, resulting in near absent expression of membrane bound IL23R. IL23R deficiency was characterized by impaired IL-23-mediated IFN-γ secretion in CD4 + , CD8 + T, and mucosal-associated invariant T (MAIT) cells, and low frequencies of circulating Th17 (CD3 + CD45RA - CCR4 + CXCR3 - RORγT + ), Th1* (CD45RA - CCR4 - CXCR3 + RORγT + ), and MAIT (CD3 + CD8 + Vα7.2 + CD161 + ) cells. Although the patient did not have a history of recurrent fungal infections, impaired Th17 differentiation and blunted IL-23-mediated IL-17 secretion in PBMCs were observed., Conclusion: We demonstrate that impaired IL-23 immunity caused by a homozygous R381X mutation in IL23R underlies MSMD, corroborating earlier findings with a homozygous p.C115Y IL23R mutation. Our report further supports a model of redundant contribution of IL-23- to IL-17-mediated anti-fungal immunity.1., (© 2022. The Author(s).)

Background: Patients with locally advanced pancreatic cancer (LAPC) are treated with chemotherapy. In selected cases, stereotactic body radiotherapy (SBRT) can be added to the regimen. We hypothesized that adding an adjuvant containing a heat-killed mycobacterium (IMM-101) to SBRT may lead to beneficial immuno-modulatory effects, thereby improving survival. This study aims to investigate the safety of adding IMM-101 to SBRT and to investigate the immuno-modulatory effects of the combination treatment in the peripheral blood of LAPC patients., Methods: LAPC patients were treated with SBRT (40 Gy) and six intradermal vaccinations of one milligram IMM-101. The primary endpoint was an observed toxicity rate of grade 4 or higher. Targeted gene-expression profiling and multicolor flow cytometry were performed for longitudinal immune-monitoring of the peripheral blood., Results: Twenty patients received study treatment. No treatment-related adverse events of grade 4 or higher occurred. SBRT/IMM-101 treatment induced a transient decrease in different lymphocyte subsets and an increase in CD14+CD16-CD11b+HLA-DR low myeloid-derived suppressor cells. Importantly, treatment significantly increased activated ICOS+, HLA-DR+ and Ki67+PD1+ T and NK cell frequencies. This was not accompanied by increased levels of most inhibitory markers, such as TIM-3 and LAG-3., Conclusions: Combination therapy with SBRT and a heat-killed mycobacterium vaccine was safe and had an immune-stimulatory effect.

NFKB1 haploinsufficiengcy was first described in 2015 in three families with common variable immunodeficiency (CVID), presenting heterogeneously with symptoms of increased infectious susceptibility, skin lesions, malignant lymphoproliferation and autoimmunity. The described mutations all led to a rapid degradation of the mutant protein, resulting in a p50 haploinsufficient state. Since then, more than 50 other mutations have been reported, located throughout different domains of NFKB1 with the majority situated in the N-terminal Rel homology domain (RHD). The clinical spectrum has also expanded with possible disease manifestations in almost any organ system. In silico prediction tools are often used to estimate the pathogenicity of NFKB1 variants but to prove causality between disease and genetic findings, further downstream functional validation is required. In this report, we studied 2 families with CVID and two novel variants in NFKB1 (c.1638-2A>G and c.787G>C). Both mutations affected mRNA and/or protein expression of NFKB1 and resulted in excessive NLRP3 inflammasome activation in patient macrophages and upregulated interferon stimulated gene expression. Protein-protein interaction analysis demonstrated a loss of interaction with NFKB1 interaction partners for the p.V263L mutation. In conclusion, we proved pathogenicity of two novel variants in NFKB1 in two families with CVID characterized by variable and incomplete penetrance., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Staels, De Keukeleere, Kinnunen, Keskitalo, Lorenzetti, Vanmeert, Prezzemolo, Pasciuto, Lescrinier, Bossuyt, Gerbaux, Willemsen, Neumann, Van Loo, Corveleyn, Willekens, Stalmans, Meyts, Liston, Humblet-Baron, Seppänen, Varjosalo and Schrijvers.)

Purpose: Lurbinectedin is a promising new drug being investigated in pre-treated patients with small cell lung cancer (SCLC) or malignant pleural mesothelioma (MPM). Its clinical activity in the real-world setting has not been investigated yet., Patients and Methods: Clinical data of patients with SCLC and MPM who were treated with lurbinectedin were prospectively collected. Comprehensive immune cell profiling by flow cytometry was performed on screening and treating peripheral blood samples., Results: A total of 95 patients (43 SCLC and 52 MPM) were treated, mostly as ≥3-line of therapy. In the SCLC cohort, a median progression-free survival (mPFS) was 1.5 months (95% CI: 1.4-3.0), and median overall survival was 7.0 months (95% CI: 4.7-not reached). Objective radiological response and disease control rate after 12 weeks were 16% and 28%, respectively. In the MPM cohort, median progression-free survival was 2.8 months (95% CI: 1.4-4.2), and median overall survival was 7.2 months (95% CI: 5.9-not reached). Disease control rate after 12 weeks was 29%, whereas no partial responses were registered. No new safety signals were observed. Lurbinectedin treatment was significantly associated with the depletion of circulating classical monocytes, which correlated with a better PFS in patients with SCLC. Lurbinectedin increased the proliferation of CD4 + and CD8 + T cells (SCLC) and natural killer and natural killer T cells (SCLC and MPM) and altered co-stimulatory and co-inhibitory receptor expression on circulating lymphocytes., Conclusion: Lurbinectedin has a manageable safety profile and shows clinical activity in pre-treated patients with SCLC and MPM. Its immune-modulatory functions make lurbinectedin a potential platform for immunotherapy combinations., Competing Interests: Conflict of interest statement The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: The authors declare no relevant conflict of interest related to the published manuscript. Relevant financial activities outside the submitted work: DD reports receiving speakers fee from BMS, Roche, Pfizer, and Novartis. LC is granted by ESMO with an ESMO Translational Research Fellowship. Any views, opinions, findings, conclusions, or recommendations expressed in this material are those solely of the author(s) and do not necessarily reflect those of ESMO. RC reports consulting for Roche, MSD, Boehringer Ingelheim and receiving speaker's fee from BMS, Roche, Pfizer, Boehringer Ingelheim, Novartis. AD reports receiving grants from Bristol-Myers Squibb, AbbVie, and Amgen; and other fees from Roche, Eli Lilly, Boehringer Ingelheim, AstraZeneca, Bristol-Myers Squibb, Amgen, Novartis, Merck Sharp & Dohme, Takeda, and PharmaMar outside of the submitted work. JGJVA reports receiving commercial research grants from Amphera, Eli-Lilly and Roche, holds ownership interest (including patents) in Amphera BV, and is a consultant/advisory board member for Amphera, Boehringer Ingelheim, Bristol-Myers Squibb, Eli-Lilly, MSD, Takeda, Bayer, Astra Zeneca and Roche. The other authors have no conflicts of interest to declare., (Copyright © 2022 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Background: Cytochrome P450 4F3 (CYP4F3) is an ω-hydroxylase that oxidizes leukotriene B4 (LTB4), prostaglandins, and fatty acid epoxides. LTB4 is synthesized by leukocytes and acts as a chemoattractant for neutrophils, making it an essential component of the innate immune system. Recently, involvement of the LTB4 pathway was reported in various immunological disorders such as asthma, arthritis, and inflammatory bowel disease. We report a 26-year-old female with a complex immune phenotype, mainly marked by exhaustion, muscle weakness, and inflammation-related conditions. The molecular cause is unknown, and symptoms have been aggravating over the years., Methods: Whole exome sequencing was performed and validated; flow cytometry and enzyme-linked immunosorbent assay were used to describe patient's phenotype. Function and impact of the mutation were investigated using molecular analysis: co-immunoprecipitation, western blot, and enzyme-linked immunosorbent assay. Capillary electrophoresis with ultraviolet detection was used to detect LTB4 and its metabolite and in silico modelling provided structural information., Results: We present the first report of a patient with a heterozygous de novo missense mutation c.C1123 > G;p.L375V in CYP4F3 that severely impairs its activity by 50% (P < 0.0001), leading to reduced metabolization of the pro-inflammatory LTB4. Systemic LTB4 levels (1034.0 ± 75.9 pg/mL) are significantly increased compared with healthy subjects (305.6 ± 57.0 pg/mL, P < 0.001), and immune phenotyping shows increased total CD19+ CD27- naive B cells (25%) and decreased total CD19+ CD27+ IgD- switched memory B cells (19%). The mutant CYP4F3 protein is stable and binding with its electron donors POR and Cytb5 is unaffected (P > 0.9 for both co-immunoprecipitation with POR and Cytb5). In silico modelling of CYP4F3 in complex with POR and Cytb5 suggests that the loss of catalytic activity of the mutant CYP4F3 is explained by a disruption of an α-helix that is crucial for the electron shuffling between the electron carriers and CYP4F3. Interestingly, zileuton still inhibits ex vivo LTB4 production in patient's whole blood to 2% of control (P < 0.0001), while montelukast and fluticasone do not (99% and 114% of control, respectively)., Conclusions: A point mutation in the catalytic domain of CYP4F3 is associated with high leukotriene B4 plasma levels and features of a more naive adaptive immune response. Our data provide evidence for the pathogenicity of the CYP4F3 variant as a cause for the observed clinical features in the patient. Inhibitors of the LTB4 pathway such as zileuton show promising effects in blocking LTB4 production and might be used as a future treatment strategy., (© 2022 The Authors. Journal of Cachexia, Sarcopenia and Muscle published by John Wiley & Sons Ltd on behalf of Society on Sarcopenia, Cachexia and Wasting Disorders.)