Homozygous variant in TKFC abolishing triokinase activities is associated with isolated immunodeficiency.

BACKGROUND: Triokinase and FMN cyclase (TKFC) is a bifunctional enzyme involved in fructose metabolism. Triokinase catalyses the phosphorylation of fructose-derived glyceraldehyde (GA) and exogenous dihydroxyacetone (DHA), while FMN cyclase generates cyclic FMN. TKFC regulates the antiviral immune response by interacting with IFIH1 (MDA5). Previously reported pathogenic variants in TKFC are associated with either a multisystemic disease or isolated hypotrichosis with loose anagen hairs. METHODS: Whole-exome sequencing identified a homozygous novel variant in TKFC (c.1624G>A; p.Gly542Arg) in an individual with a complex primary immunodeficiency disorder. The variant was characterised using enzymatic assays and yeast studies of mutant recombinant proteins. RESULTS: The individual presented with chronic active Epstein-Barr virus disease and multiple bacterial and viral infections. Clinical investigations revealed hypogammaglobulinaemia, near absent natural killer cells and decreased memory B cells. Enzymatic assays showed that this variant displayed defective DHA and GA kinase activity while maintaining FMN cyclase activity. An allogenic bone marrow transplantation corrected the patient's immunodeficiency. CONCLUSION: Our report suggests that TKFC may have a role in the immunological system. The pathological features associated with this variant are possibly linked with DHA/GA kinase inactivation through a yet an unknown mechanism. This report thus adds a possible new pathway of immunometabolism to explore further.

Allogeneic hematopoietic cell transplantation is effective for p47phox chronic granulomatous disease: A Primary Immune Deficiency Treatment Consortium study.

BACKGROUND: P47phox (neutrophil cytosolic factor-1) deficiency is the most common cause of autosomal recessive chronic granulomatous disease (CGD) and is considered to be associated with a milder clinical phenotype. Allogeneic hematopoietic cell transplantation (HCT) for p47phox CGD is not well-described. OBJECTIVES: We sought to study HCT for p47phox CGD in North America. METHODS: Thirty patients with p47phox CGD who received allogeneic HCT at Primary Immune Deficiency Treatment Consortium centers since 1995 were included. RESULTS: Residual oxidative activity was present in 66.7% of patients. In the year before HCT, there were 0.38 CGD-related infections per person-years. Inflammatory diseases, predominantly of the lungs and bowel, occurred in 36.7% of the patients. The median age at HCT was 9.1 years (range 1.5-23.6 years). Most HCTs (90%) were performed after using reduced intensity/toxicity conditioning. HCT sources were HLA-matched (40%) and -mismatched (10%) related donors or HLA-matched (36.7%) and -mismatched (13.3%) unrelated donors. CGD-related infections after HCT decreased significantly to 0.06 per person-years (P = .038). The frequency of inflammatory bowel disease and the use of steroids also decreased. The cumulative incidence of graft failure and second HCT was 17.9%. The 2-year overall and event-free survival were 92.3% and 82.1%, respectively, while at 5 years they were 85.7% and 77.0%, respectively. In the surviving patients evaluated, ≥95% donor myeloid chimerism at 1 and 2 years after HCT was 93.8% and 87.5%, respectively. CONCLUSIONS: Patients with p47phox CGD suffer from a significant disease burden that can be effectively alleviated by HCT. Similar to other forms of CGD, HCT should be considered for patients with p47phox CGD.

Outcomes following treatment for ADA-deficient severe combined immunodeficiency: a report from the PIDTC.

Adenosine deaminase (ADA) deficiency causes ∼13% of cases of severe combined immune deficiency (SCID). Treatments include enzyme replacement therapy (ERT), hematopoietic cell transplant (HCT), and gene therapy (GT). We evaluated 131 patients with ADA-SCID diagnosed between 1982 and 2017 who were enrolled in the Primary Immune Deficiency Treatment Consortium SCID studies. Baseline clinical, immunologic, genetic characteristics, and treatment outcomes were analyzed. First definitive cellular therapy (FDCT) included 56 receiving HCT without preceding ERT (HCT); 31 HCT preceded by ERT (ERT-HCT); and 33 GT preceded by ERT (ERT-GT). Five-year event-free survival (EFS, alive, no need for further ERT or cellular therapy) was 49.5% (HCT), 73% (ERT-HCT), and 75.3% (ERT-GT; P < .01). Overall survival (OS) at 5 years after FDCT was 72.5% (HCT), 79.6% (ERT-HCT), and 100% (ERT-GT; P = .01). Five-year OS was superior for patients undergoing HCT at <3.5 months of age (91.6% vs 68% if ≥3.5 months, P = .02). Active infection at the time of HCT (regardless of ERT) decreased 5-year EFS (33.1% vs 68.2%, P < .01) and OS (64.7% vs 82.3%, P = .02). Five-year EFS (90.5%) and OS (100%) were best for matched sibling and matched family donors (MSD/MFD). For patients treated after the year 2000 and without active infection at the time of FDCT, no difference in 5-year EFS or OS was found between HCT using a variety of transplant approaches and ERT-GT. This suggests alternative donor HCT may be considered when MSD/MFD HCT and GT are not available, particularly when newborn screening identifies patients with ADA-SCID soon after birth and before the onset of infections. This trial was registered at www.clinicaltrials.gov as #NCT01186913 and #NCT01346150.

The diagnosis of severe combined immunodeficiency (SCID): The Primary Immune Deficiency Treatment Consortium (PIDTC) 2022 Definitions.

Severe combined immunodeficiency (SCID) results from defects in the differentiation of hematopoietic stem cells into mature T lymphocytes, with additional lymphoid lineages affected in particular genotypes. In 2014, the Primary Immune Deficiency Treatment Consortium published criteria for diagnosing SCID, which are now revised to incorporate contemporary approaches. Patients with typical SCID must have less than 0.05 × 109 autologous T cells/L on repetitive testing, with either pathogenic variant(s) in a SCID-associated gene, very low/undetectable T-cell receptor excision circles or less than 20% of CD4 T cells expressing naive markers, and/or transplacental maternally engrafted T cells. Patients with less profoundly impaired autologous T-cell differentiation are designated as having leaky/atypical SCID, with 2 or more of these: low T-cell numbers, oligoclonal T cells, low T-cell receptor excision circles, and less than 20% of CD4 T cells expressing naive markers. These patients must also have either pathogenic variant(s) in a SCID-associated gene or reduced T-cell proliferation to certain mitogens. Omenn syndrome requires a generalized erythematous rash, absent transplacentally acquired maternal engraftment, and 2 or more of these: eosinophilia, elevated IgE, lymphadenopathy, hepatosplenomegaly. Thymic stromal defects and other causes of secondary T-cell deficiency are excluded from the definition of SCID. Application of these revised Primary Immune Deficiency Treatment Consortium 2022 Definitions permits precise categorization of patients with T-cell defects but does not imply a preferred treatment strategy.

Aberrant T-cell exhaustion in severe combined immunodeficiency survivors with poor T-cell reconstitution after transplantation.

BACKGROUND: Severe combined immunodeficiency (SCID) comprises rare inherited disorders of immunity that require definitive treatment through hematopoietic cell transplantation (HCT) or gene therapy for survival. Despite successes of allogeneic HCT, many SCID patients experience incomplete immune reconstitution, persistent T-cell lymphopenia, and poor long-term outcomes. OBJECTIVE: We hypothesized that CD4+ T-cell lymphopenia could be associated with a state of T-cell exhaustion in previously transplanted SCID patients. METHODS: We analyzed markers of exhaustion in blood samples from 61 SCID patients at a median of 10.4 years after HCT. RESULTS: Compared to post-HCT SCID patients with normal CD4+ T-cell counts, those with poor T-cell reconstitution showed lower frequency of naive CD45RA+/CCR7+ T cells, recent thymic emigrants, and TCR excision circles. They also had a restricted TCR repertoire, increased expression of inhibitory receptors (PD-1, 2B4, CD160, BTLA, CTLA-4), and increased activation markers (HLA-DR, perforin) on their total and naive CD8+ T cells, suggesting T-cell exhaustion and aberrant activation, respectively. The exhaustion score of CD8+ T cells was inversely correlated with CD4+ T-cell count, recent thymic emigrants, TCR excision circles, and TCR diversity. Exhaustion scores were higher among recipients of unconditioned HCT, especially when further in time from HCT. Patients with fewer CD4+ T cells showed a transcriptional signature of exhaustion. CONCLUSIONS: Recipients of unconditioned HCT for SCID may develop late post-HCT T-cell exhaustion as a result of diminished production of T-lineage cells. Elevated expression of inhibitory receptors on their T cells may be a biomarker of poor long-term T-cell reconstitution.

The diagnosis of severe combined immunodeficiency: Implementation of the PIDTC 2022 Definitions.

BACKGROUND: Shearer et al in 2014 articulated well-defined criteria for the diagnosis and classification of severe combined immunodeficiency (SCID) as part of the Primary Immune Deficiency Treatment Consortium's (PIDTC's) prospective and retrospective studies of SCID. OBJECTIVE: Because of the advent of newborn screening for SCID and expanded availability of genetic sequencing, revision of the PIDTC 2014 Criteria was needed. METHODS: We developed and tested updated PIDTC 2022 SCID Definitions by analyzing 379 patients proposed for prospective enrollment into Protocol 6901, focusing on the ability to distinguish patients with various SCID subtypes. RESULTS: According to PIDTC 2022 Definitions, 18 of 353 patients eligible per 2014 Criteria were considered not to have SCID, whereas 11 of 26 patients ineligible per 2014 Criteria were determined to have SCID. Of note, very low numbers of autologous T cells (<0.05 × 109/L) characterized typical SCID under the 2022 Definitions. Pathogenic variant(s) in SCID-associated genes was identified in 93% of patients, with 7 genes (IL2RG, RAG1, ADA, IL7R, DCLRE1C, JAK3, and RAG2) accounting for 89% of typical SCID. Three genotypes (RAG1, ADA, and RMRP) accounted for 57% of cases of leaky/atypical SCID; there were 13 other rare genotypes. Patients with leaky/atypical SCID were more likely to be diagnosed at more than age 1 year than those with typical SCID lacking maternal T cells: 20% versus 1% (P < .001). Although repeat testing proved important, an initial CD3 T-cell count of less than 0.05 × 109/L differentiated cases of typical SCID lacking maternal cells from leaky/atypical SCID: 97% versus 7% (P < .001). CONCLUSIONS: The PIDTC 2022 Definitions describe SCID and its subtypes more precisely than before, facilitating analyses of SCID characteristics and outcomes.

Intestinal microbiome and metabolome signatures in patients with chronic granulomatous disease.

BACKGROUND: Chronic granulomatous disease (CGD) is caused by defects in any 1 of the 6 subunits forming the nicotinamide adenine dinucleotide phosphate oxidase complex 2 (NOX2), leading to severely reduced or absent phagocyte-derived reactive oxygen species production. Almost 50% of patients with CGD have inflammatory bowel disease (CGD-IBD). While conventional IBD therapies can treat CGD-IBD, their benefits must be weighed against the risk of infection. Understanding the impact of NOX2 defects on the intestinal microbiota may lead to the identification of novel CGD-IBD treatments. OBJECTIVE: We sought to identify microbiome and metabolome signatures that can distinguish individuals with CGD and CGD-IBD. METHODS: We conducted a cross-sectional observational study of 79 patients with CGD, 8 pathogenic variant carriers, and 19 healthy controls followed at the National Institutes of Health Clinical Center. We profiled the intestinal microbiome (amplicon sequencing) and stool metabolome, and validated our findings in a second cohort of 36 patients with CGD recruited through the Primary Immune Deficiency Treatment Consortium. RESULTS: We identified distinct intestinal microbiome and metabolome profiles in patients with CGD compared to healthy individuals. We observed enrichment for Erysipelatoclostridium spp, Sellimonas spp, and Lachnoclostridium spp in CGD stool samples. Despite differences in bacterial alpha and beta diversity between the 2 cohorts, several taxa correlated significantly between both cohorts. We further demonstrated that patients with CGD-IBD have a distinct microbiome and metabolome profile compared to patients without CGD-IBD. CONCLUSION: Intestinal microbiome and metabolome signatures distinguished patients with CGD and CGD-IBD, and identified potential biomarkers and therapeutic targets.

Real-World Effectiveness of Nirmatrelvir/Ritonavir on Coronavirus Disease 2019-Associated Hospitalization Prevention: A Population-based Cohort Study in the Province of Quebec, Canada.

BACKGROUND: Nirmatrelvir/ritonavir has shown to reduce COVID-19 hospitalization and death before Omicron, but updated real-world evidence studies are needed. This study aimed to assess whether nirmatrelvir/ritonavir reduces the risk of COVID-19-associated hospitalization among high-risk outpatients. METHODS: A retrospective cohort study of outpatients with SARS-CoV-2 between March 15 and 15 October 2022, using data from the Quebec clinico-administrative databases. Outpatients treated with nirmatrelvir/ritonavir were compared with infected ones not receiving nirmatrelvir/ritonavir using propensity-score matching. Relative risk (RR) of COVID-19-associated hospitalization within 30 days was assessed using a Poisson regression. RESULTS: A total of 8402 treated outpatients were matched to controls. Regardless of vaccination status, nirmatrelvir/ritonavir treatment was associated with a 69% reduced RR of hospitalization (RR: .31; 95% CI: .28; .36; number needed to treat [NNT] = 13). The effect was more pronounced in outpatients with incomplete primary vaccination (RR: .04; 95% CI: .03; .06; NNT = 8), while no benefit was found in those with a complete primary vaccination (RR: .93; 95% CI: .78; 1.08). Subgroups analysis among high-risk outpatients with a complete primary vaccination showed that nirmatrelvir/ritonavir treatment was associated with a significant decrease in the RR of hospitalization in severely immunocompromised outpatients (RR: .66; 95% CI: .50; .89; NNT = 16) and in high-risk outpatients aged ≥70 years (RR: .50; 95% CI: .34; .74; NNT = 10) when the last dose of the vaccine was received at least 6 months ago. CONCLUSIONS: Nirmatrelvir/ritonavir reduces the risk of COVID-19-associated hospitalization among incompletely vaccinated high-risk outpatients and among some subgroups of completely vaccinated high-risk outpatients.

PEGylated Recombinant Adenosine Deaminase Maintains Detoxification and Lymphocyte Counts in Patients with ADA-SCID.

PURPOSE: Metabolic detoxification with enzyme replacement therapy (ERT) promotes immune recovery in patients with adenosine deaminase (ADA)-deficient severe combined immunodeficiency (ADA-SCID). Elapegademase is a PEGylated recombinant bovine ADA ERT developed to replace the now-discontinued bovine-derived pegademase. This study was a 1-way crossover from pegademase to elapegademase in 7 patients with ADA-SCID to assess efficacy and safety outcomes for elapegademase. METHODS: After once-weekly pegademase dosage was adjusted to achieve therapeutic metabolic detoxification and trough ADA activity, patients transitioned to a bioequivalent dose of elapegademase. Maintenance of metabolic detoxification and adequate ADA activity were evaluated periodically. RESULTS: One patient withdrew after 2 doses of an early elapegademase formulation due to injection-site pain caused by EDTA. The 6 remaining patients completed 71-216 weeks of elapegademase therapy with a formulation that did not contain EDTA. In these patients, elapegademase improved ADA activity compared with pegademase and maintained metabolic detoxification. Total lymphocyte counts increased for all completer patients from between 1.2- and 2.1-fold at the end of study compared with baseline. Elapegademase had a comparable safety profile to pegademase; no patient developed a severe infectious complication. Three patients had transient, non-neutralizing antibodies to pegademase, elapegademase, and/or polyethylene glycol ≤ 47 weeks of treatment without effect on trough plasma ADA activity or trough erythrocyte deoxyadenosine nucleotide levels. CONCLUSION: Elapegademase was safe, well tolerated, achieved stable trough plasma ADA activity with weekly dosing, was effective in maintaining metabolic detoxification, and was associated with maintenance or improvements in lymphocyte counts compared with pegademase therapy in patients with ADA-SCID.

Clinical Practice Guidelines for the Immunological Management of Chromosome 22q11.2 Deletion Syndrome and Other Defects in Thymic Development.

Current practices vary widely regarding the immunological work-up and management of patients affected with defects in thymic development (DTD), which include chromosome 22q11.2 microdeletion syndrome (22q11.2del) and other causes of DiGeorge syndrome (DGS) and coloboma, heart defect, atresia choanae, retardation of growth and development, genital hypoplasia, ear anomalies/deafness (CHARGE) syndrome. Practice variations affect the initial and subsequent assessment of immune function, the terminology used to describe the condition and immune status, the accepted criteria for recommending live vaccines, and how often follow-up is needed based on the degree of immune compromise. The lack of consensus and widely varying practices highlight the need to establish updated immunological clinical practice guidelines. These guideline recommendations provide a comprehensive review for immunologists and other clinicians who manage immune aspects of this group of disorders.

A decade of progress in juvenile idiopathic athritis treatments and outcomes in Canada: results from ReACCh-Out and the CAPRI registry.

OBJECTIVE: To assess changes in juvenile idiopathic arthritis (JIA) treatments and outcomes in Canada, comparing a 2005-2010 and a 2017-2021 inception cohorts. METHODS: Patients enrolled within three months of diagnosis in the Research in Arthritis in Canadian Children Emphasizing Outcomes (ReACCh-Out) and the Canadian Alliance of Pediatric Rheumatology Investigators Registry (CAPRI) cohorts were included. Cumulative incidences of drug starts and outcome attainment within 70 weeks of diagnosis were compared with Kaplan Meier survival analysis and multivariable Cox regression. RESULTS: The 2005-2010 and 2017-2021 cohorts included 1128 and 721 patients, respectively. JIA category distribution and baseline clinical juvenile idiopathic arthritis disease activity (cJADAS10) scores at enrolment were comparable. By 70 weeks, 6% of patients (95% CI 5, 7) in the 2005-2010 and 26% (23, 30) in the 2017-2021 cohort had started a biologic DMARD (bDMARD), and 43% (40, 47) and 60% (56, 64) had started a conventional DMARD (cDMARD), respectively. Outcome attainment was 64% (61, 67) and 83% (80, 86) for Inactive disease (Wallace criteria), 69% (66, 72) and 84% (81, 87) for minimally active disease (cJADAS10 criteria), 57% (54, 61) and 63% (59, 68) for pain control (<1/10), and 52% (47, 56) and 54% (48, 60) for a good health-related quality of life. CONCLUSION: Although baseline disease characteristics were comparable in the 2005-2010 and 2017-2021 cohorts, cDMARD and bDMARD use increased with a concurrent increase in minimally active and inactive disease. Improvements in parent and patient reported outcomes were smaller than improvements in disease activity.

Genome-wide CRISPR screens identify ferroptosis as a novel therapeutic vulnerability in acute lymphoblastic leukemia.

Acute lymphoblastic leukemia (ALL) is the most frequent cancer diagnosed in children. Despite the great progress achieved over the last 40 years, with cure rates now exceeding 85%, refractory or relapsed ALL still exhibit a dismal prognosis. This poor outcome reflects the lack of treatment options specifically targeting relapsed or refractory ALL. In order to address this gap, we performed whole-genome CRISPR/Cas drop-out screens on a panel of seven B-ALL cell lines. Our results demonstrate that while there was a significant overlap in gene essentiality between ALL cell lines and other cancer types survival of ALL cell lines was dependent on several unique metabolic pathways, including an exquisite sensitivity to GPX4 depletion and ferroptosis induction. Detailed molecular analysis of B-ALL cells suggest that they are primed to undergo ferroptosis as they exhibit high steady-state oxidative stress potential, a low buffering capacity, and a disabled GPX4-independent secondary lipid peroxidation detoxification pathway. Finally, we validated the sensitivity of BALL to ferroptosis induction using patient-derived B-ALL samples.

Paediatric inflammatory multisystem syndrome in Canada: population-based surveillance and role of SARS-CoV-2 linkage.

BACKGROUND: Paediatric inflammatory multisystem syndrome (PIMS) is a rare condition temporally associated with SARS-CoV-2 infection. Using national surveillance data, we compare presenting features and outcomes among children hospitalized with PIMS by SARS-CoV-2 linkage, and identify risk factors for intensive care (ICU). METHODS: Cases were reported to the Canadian Paediatric Surveillance Program by a network of >2800 pediatricians between March 2020 and May 2021. Patients with positive versus negative SARS-CoV-2 linkages were compared, with positive linkage defined as any positive molecular or serologic test or close contact with confirmed COVID-19. ICU risk factors were identified with multivariable modified Poisson regression. RESULTS: We identified 406 children hospitalized with PIMS, including 49.8% with positive SARS-CoV-2 linkages, 26.1% with negative linkages, and 24.1% with unknown linkages. The median age was 5.4 years (IQR 2.5-9.8), 60% were male, and 83% had no comorbidities. Compared to cases with negative linkages, children with positive linkages experienced more cardiac involvement (58.8% vs. 37.4%; p < 0.001), gastrointestinal symptoms (88.6% vs. 63.2%; p < 0.001), and shock (60.9% vs. 16.0%; p < 0.001). Children aged ≥6 years and those with positive linkages were more likely to require ICU. CONCLUSIONS: Although rare, 30% of PIMS hospitalizations required ICU or respiratory/hemodynamic support, particularly those with positive SARS-CoV-2 linkages. IMPACT: We describe 406 children hospitalized with paediatric inflammatory multisystem syndrome (PIMS) using nationwide surveillance data, the largest study of PIMS in Canada to date. Our surveillance case definition of PIMS did not require a history of SARS-CoV-2 exposure, and we therefore describe associations of SARS-CoV-2 linkages on clinical features and outcomes of children with PIMS. Children with positive SARS-CoV-2 linkages were older, had more gastrointestinal and cardiac involvement, and hyperinflammatory laboratory picture. Although PIMS is rare, one-third required admission to intensive care, with the greatest risk amongst those aged ≥6 years and those with a SARS-CoV-2 linkage.

Granulocyte Transfusions in Patients with Chronic Granulomatous Disease Undergoing Hematopoietic Cell Transplantation or Gene Therapy.

Granulocyte transfusions are sometimes used as adjunctive therapy for the treatment of infection in patients with chronic granulomatous disease (CGD). However, granulocyte transfusions can be associated with a high rate of alloimmunization, and their role in CGD patients undergoing hematopoietic cell transplantation (HCT) or gene therapy (GT) is unknown. We identified 27 patients with CGD who received granulocyte transfusions pre- (within 6 months) and/or post-HCT or GT in a retrospective survey. Twelve patients received granulocyte transfusions as a bridge to cellular therapy. Six (50%) of these patients had a complete or partial response. However, six of 10 (60%) patients for whom testing was performed developed anti-HLA antibodies, and three of the patients also had severe immune-mediated cytopenia within the first 100 days post-HCT or GT. Fifteen patients received granulocyte transfusions post-HCT only. HLA antibodies were not checked for any of these 15 patients, but there were no cases of early immune-mediated cytopenia. Out of 25 patients who underwent HCT, there were 5 (20%) cases of primary graft failure. Three of the patients with primary graft failure had received granulocyte transfusions pre-HCT and were subsequently found to have anti-HLA antibodies. In this small cohort of patients with CGD, granulocyte transfusions pre-HCT or GT were associated with high rates of alloimmunization, primary graft failure, and early severe immune-mediated cytopenia post-HCT or GT. Granulocyte transfusions post-HCT do not appear to confer an increased risk of graft failure.

CD5 levels define functionally heterogeneous populations of naïve human CD4+ T cells.

Studies in murine models show that subthreshold TCR interactions with self-peptide are required for thymic development and peripheral survival of naïve T cells. Recently, differences in the strength of tonic TCR interactions with self-peptide, as read-out by cell surface levels of CD5, were associated with distinct effector potentials among sorted populations of T cells in mice. However, whether CD5 can also be used to parse functional heterogeneity among human T cells is less clear. Our study demonstrates that CD5 levels correlate with TCR signal strength in human naïve CD4+ T cells. Further, we describe a relationship between CD5 levels on naïve human CD4+ T cells and binding affinity to foreign peptide, in addition to a predominance of CD5hi T cells in the memory compartment. Differences in gene expression and biases in cytokine production potential between CD5lo and CD5hi naïve human CD4+ T cells are consistent with observations in mice. Together, these data validate the use of CD5 surface levels as a marker of heterogeneity among human naïve CD4+ T cells with important implications for the identification of functionally biased T- cell populations that can be exploited to improve the efficacy of adoptive cell therapies.

IL-27 shapes the immune properties of human astrocytes and their impact on encountered human T lymphocytes.

BACKGROUND: Interleukin-27 (IL-27) can trigger both pro- and anti-inflammatory responses. This cytokine is elevated in the central nervous system (CNS) of multiple sclerosis (MS) patients, but how it influences neuroinflammatory processes remains unclear. As astrocytes express the receptor for IL-27, we sought to determine how these glial cells respond to this cytokine and whether such exposure alters their interactions with infiltrating activated T lymphocytes. To determine whether inflammation shapes the impact of IL-27, we compared the effects of this cytokine in non-inflamed and inflamed conditions induced by an IL-1ß exposure. MAIN BODY: Transcriptomic analysis of IL-27-exposed human astrocytes showed an upregulation of multiple immune genes. Human astrocytes increased the secretion of chemokines (CXCL9, CXCL10, and CXCL11) and the surface expression of proteins (PD-L1, HLA-E, and ICAM-1) following IL-27 exposure. To assess whether exposure of astrocytes to IL-27 influences the profile of activated T lymphocytes infiltrating the CNS, we used an astrocyte/T lymphocyte co-culture model. Activated human CD4+ or CD8+ T lymphocytes were co-cultured with astrocytes that have been either untreated or pre-exposed to IL­27 or IL-1ß. After 24 h, we analyzed T lymphocytes by flow cytometry for transcription factors and immune molecules. The contact with IL-27-exposed astrocytes increased the percentages of T-bet, Eomes, CD95, IL-18Rα, ICAM-1, and PD-L1 expressing CD4+ and CD8+ T lymphocytes and reduced the proportion of CXCR3-positive CD8+ T lymphocytes. Human CD8+ T lymphocytes co-cultured with human IL-27-treated astrocytes exhibited higher motility than when in contact with untreated astrocytes. These results suggested a preponderance of kinapse-like over synapse-like interactions between CD8+ T lymphocytes and IL-27-treated astrocytes. Finally, CD8+ T lymphocytes from MS patients showed higher motility in contact with IL-27-exposed astrocytes compared to healthy donors' cells. CONCLUSION: Our results establish that IL-27 alters the immune functions of human astrocytes and shapes the profile and motility of encountered T lymphocytes, especially CD8+ T lymphocytes from MS patients.

Excellent outcomes following hematopoietic cell transplantation for Wiskott-Aldrich syndrome: a PIDTC report.

Wiskott-Aldrich syndrome (WAS) is an X-linked disease caused by mutations in the WAS gene, leading to thrombocytopenia, eczema, recurrent infections, autoimmune disease, and malignancy. Hematopoietic cell transplantation (HCT) is the primary curative approach, with the goal of correcting the underlying immunodeficiency and thrombocytopenia. HCT outcomes have improved over time, particularly for patients with HLA-matched sibling and unrelated donors. We report the outcomes of 129 patients with WAS who underwent HCT at 29 Primary Immune Deficiency Treatment Consortium centers from 2005 through 2015. Median age at HCT was 1.2 years. Most patients (65%) received myeloablative busulfan-based conditioning. With a median follow-up of 4.5 years, the 5-year overall survival (OS) was 91%. Superior 5-year OS was observed in patients <5 vs ≥5 years of age at the time of HCT (94% vs 66%; overall P = .0008). OS was excellent regardless of donor type, even in cord blood recipients (90%). Conditioning intensity did not affect OS, but was associated with donor T-cell and myeloid engraftment after HCT. Specifically, patients who received fludarabine/melphalan-based reduced-intensity regimens were more likely to have donor myeloid chimerism <50% early after HCT. In addition, higher platelet counts were observed among recipients who achieved full (>95%) vs low-level (5%-49%) donor myeloid engraftment. In summary, HCT outcomes for WAS have improved since 2005, compared with prior reports. HCT at a younger age continues to be associated with superior outcomes supporting the recommendation for early HCT. High-level donor myeloid engraftment is important for platelet reconstitution after either myeloablative or busulfan-containing reduced intensity conditioning. (This trial was registered at www.clinicaltrials.gov as #NCT02064933.).

Rituximab-induced hypogammaglobulinemia and infection risk in pediatric patients.

BACKGROUND: Rituximab is a B-cell depleting agent used in B-cell malignancies and autoimmune diseases. A subset of adult patients may develop prolonged and symptomatic hypogammaglobulinemia following rituximab treatment. However, this phenomenon has not been well delineated in the pediatric population. OBJECTIVES: This study sought to determine the prevalence, risk factors, and clinical significance of hypogammaglobulinemia following rituximab therapy in children. METHODS: This was a multicenter, retrospective cohort study that extracted clinical and immunological data from pediatric patients who received rituximab. RESULTS: The cohort comprised 207 patients (median age, 12.0 years). Compared to baseline values, there was a significant increase in hypogammaglobulinemia post-rituximab therapy, with an increase in prevalence of hypo-IgG (28.7%-42.6%; P = .009), hypo-IgA (11.1%-20.4%; P = .02), and hypo-IgM (20.0%-62.0%; P < .0001). Additionally, low IgG levels at any time post-rituximab therapy were associated with a higher risk of serious infections (34.4% vs 18.9%; odds ratio, 2.3; 95% CI, 1.1-4.8; P = .03). Persistent IgG hypogammaglobulinemia was observed in 27 of 101 evaluable patients (26.7%). Significant risk factors for persistent IgG hypogammaglobulinemia included low IgG and IgA levels pre-rituximab therapy. Nine patients (4.3%) within the study were subsequently diagnosed with a primary immunodeficiency, 7 of which received rituximab for autoimmune cytopenias. CONCLUSIONS: Hypogammaglobulinemia post-rituximab treatment is frequently diagnosed within the pediatric population. Low IgG levels are associated with a significant increase in serious infections, and underlying primary immunodeficiencies are relatively common in children receiving rituximab, thus highlighting the importance of immunologic monitoring both before and after rituximab therapy.

HIV Infection and Persistence in Pulmonary Mucosal Double Negative T Cells In Vivo.

The lungs are relatively unexplored anatomical human immunodeficiency virus (HIV) reservoirs in the antiretroviral therapy (ART) era. Double negative (DN) T cells are a subset of T cells that lack expression of CD4 and CD8 (CD4- CD8-) and may have both regulatory and effector functions during HIV infection. Notably, circulating DN T cells were previously described as cellular HIV reservoirs. Here, we undertook a thorough analysis of pulmonary versus blood DN T cells of people living with HIV (PLWH) under ART. Bronchoalveolar lavage (BAL) fluid and matched peripheral blood were collected from 35 PLWH on ART and 16 uninfected volunteers without respiratory symptoms. Both PLWH and HIV-negative (HIV-) adults displayed higher frequencies of DN T cells in BAL versus blood, and these cells mostly exhibited an effector memory phenotype. In PLWH, pulmonary mucosal DN T cells expressed higher levels of HLA-DR and several cellular markers associated with HIV persistence (CCR6, CXCR3, and PD-1) than blood. We also observed that DN T cells were less senescent (CD28- CD57+) and expressed less immunosuppressive ectonucleotidase (CD73/CD39), granzyme B, and perforin in the BAL fluid than in the blood of PLWH. Importantly, fluorescence-activated cell sorter (FACS)-sorted DN T cells from the BAL fluid of PLWH under suppressive ART harbored HIV DNA. Using the humanized bone marrow-liver-thymus (hu-BLT) mouse model of HIV infection, we observed higher infection frequencies of lung DN T cells than those of the blood and spleen in both early and late HIV infection. Overall, our findings show that HIV is seeded in pulmonary mucosal DN T cells early following infection and persists in these potential cellular HIV reservoirs even during long-term ART.IMPORTANCE Reservoirs of HIV during ART are the primary reasons why HIV/AIDS remains an incurable disease. Indeed, HIV remains latent and unreachable by antiretrovirals in cellular and anatomical sanctuaries, preventing its eradication. The lungs have received very little attention compared to other anatomical reservoirs despite being immunological effector sites exhibiting characteristics ideal for HIV persistence. Furthermore, PLWH suffer from a high burden of pulmonary non-opportunistic infections, suggesting impaired pulmonary immunity despite ART. Meanwhile, various immune cell populations have been proposed to be cellular reservoirs in blood, including CD4- CD8- DN T cells, a subset that may originate from CD4 downregulation by HIV proteins. The present study aims to describe DN T cells in human and humanized mice lungs in relation to intrapulmonary HIV burden. The characterization of DN T cells as cellular HIV reservoirs and the lungs as an anatomical HIV reservoir will contribute to the development of targeted HIV eradication strategies.

The Tumor-Immune Response Is Not Compromised by Mesenchymal Stromal Cells in Humanized Mice.

Therapeutic uses of mesenchymal stromal cells (MSCs) have emerged over the past decade. Yet, their effect on tumor growth remains highly debated, particularly in an immune competent environment. In this study, we wanted to investigate the impact of human umbilical cord-derived MSCs (hUC-MSCs) on tumor growth in humanized mice generated by the human adoptive transfer of PBMCs or the cotransplantation of hematopoietic stem cells and human thymic tissue (human BLT [Hu-BLT]). Our results showed that the growth and immune rejection of engineered human fibroblastic tumors was not altered by the injection of hUC-MSCs in immune-deficient or humanized mice, respectively. This was observed whether tumor cells were injected s.c. or i.v. and independently of the injection route of the hUC-MSCs. Moreover, only in Hu-BLT mice did hUC-MSCs have some effects on the tumor-immune infiltrate, yet without altering tumor growth. These results demonstrate that hUC-MSCs do not promote fibroblastic tumor growth and neither do they prevent tumor infiltration and rejection by immune cells in humanized mice.