Activated PI3Kinase Delta Syndrome-A Multifaceted Disease.

Autosomal dominant gain-of-function mutations in the PIK3CD gene encoding the catalytic subunit p110δ of phosphoinositide 3-kinase-δ (PI3K-δ) or autosomal dominant loss-of-function mutations in the PIK3R1 gene encoding the p85α, p55α and p50α regulatory subunits cause Activated PI3-kinase-δ syndrome (APDS; referred as type 1 APDS and type 2 APDS, respectively). Consequences of these mutations are PI3K-δ hyperactivity. Clinical presentation described for both types of APDS patients is very variable, ranging from mild or asymptomatic features to profound combined immunodeficiency. Massive lymphoproliferation, bronchiectasis, increased susceptibility to bacterial and viral infections and, at a lesser extent, auto-immune manifestations and occurrence of cancer, especially B cell lymphoma, have been described for both types of APDS patients. Here, we review clinical presentation and treatment options as well as fundamental immunological and biological features associated to PI3K-δ increased signaling.

Two Monogenetic Disorders, Activated PI3-Kinase-δ Syndrome 2 and Smith-Magenis Syndrome, in One Patient: Case Report and a Literature Review of Neurodevelopmental Impact in Primary Immunodeficiencies Associated With Disturbed PI3K Signaling.

Activated PI3-kinase-δ syndrome 2 (APDS2) is caused by autosomal dominant mutations in the PIK3R1 gene encoding the p85α, p55α, and p50α regulatory subunits. Most diagnosed APDS2 patients carry mutations affecting either the splice donor or splice acceptor sites of exon 11 of the PIK3R1 gene responsible for an alternative splice product and a shortened protein. The clinical presentation of APDS2 patients is highly variable, ranging from mild to profound combined immunodeficiency features as massive lymphoproliferation, increased susceptibility to bacterial and viral infections, bronchiectasis, autoimmune manifestations, and occurrence of cancer. Non-immunological features such as growth retardation and neurodevelopmental delay have been reported for APDS2 patients. Here, we describe a patient suffering from an APDS2 associated with a Smith-Magenis syndrome (SMS), a complex genetic disorder affecting, among others, neurological manifestations and review the literature describing neurodevelopmental impacts in APDS2 and other PIDs/monogenetic disorders associated with dysregulated PI3K signaling.

Known and potential molecules associated with altered B cell development leading to predominantly antibody deficiencies.

Predominantly antibody deficiencies (PADs) encompass a heterogeneous group of disorders characterized by low immunoglobulin serum levels in the presence or absence of peripheral B cells. Clinical presentation of affected patients may include recurrent respiratory and gastrointestinal infections, invasive infections, autoimmune manifestations, allergic reactions, lymphoproliferation, and increased susceptibility to malignant transformation. In the last decades, several genetic alterations affecting B-cell development/maturation have been identified as causative of several forms of PADs, adding important information on the genetic background of PADs, which in turn should lead to a better understanding of these disorders and precise clinical management of affected patients. This review aimed to present a comprehensive overview of the known and potentially involved molecules in the etiology of PADs to elucidate the pathogenesis of these disorders and eventually offer a better prognosis for affected patients.

Genomic Spectrum and Phenotypic Heterogeneity of Human IL-21 Receptor Deficiency.

Biallelic inactivating mutations in IL21R causes a combined immunodeficiency that is often complicated by cryptosporidium infections. While eight IL-21R-deficient patients have been reported previously, the natural course, immune characteristics of disease, and response to hematopoietic stem cell transplantation (HSCT) remain to be comprehensively examined. In our study, we have collected clinical histories of 13 patients with IL-21R deficiency from eight families across seven centers worldwide, including five novel patients identified by exome or NGS panel sequencing. Eight unique mutations in IL21R were identified in these patients, including two novel mutations. Median age at disease onset was 2.5 years (0.5-7 years). The main clinical manifestations were recurrent bacterial (84.6%), fungal (46.2%), and viral (38.5%) infections; cryptosporidiosis-associated cholangitis (46.2%); and asthma (23.1%). Inflammatory skin diseases (15.3%) and recurrent anaphylaxis (7.9%) constitute novel phenotypes of this combined immunodeficiency. Most patients exhibited hypogammaglobulinemia and reduced proportions of memory B cells, circulating T follicular helper cells, MAIT cells and terminally differentiated NK cells. However, IgE levels were elevated in 50% of IL-21R-deficient patients. Overall survival following HSCT (6 patients, mean follow-up 1.8 year) was 33.3%, with pre-existing organ damage constituting a negative prognostic factor. Mortality of non-transplanted patients (n = 7) was 57.1%. Our detailed analysis of the largest cohort of IL-21R-deficient patients to date provides in-depth clinical, immunological and immunophenotypic features of these patients, thereby establishing critical non-redundant functions of IL-21/IL-21R signaling in lymphocyte differentiation, humoral immunity and host defense against infection, and mechanisms of disease pathogenesis due to IL-21R deficiency. Outcome following HSCT depends on prior chronic infections and organ damage, which should thus be considered as early as possible following molecular diagnosis.

Class Switch Recombination Defects: impact on B cell maturation and antibody responses.

To assess how B cell phenotype analysis correlates with antigen responses in patients with class switch recombination defects (CSRD) we quantified memory B cells by flow-cytometry and immunized CSRD patients with the neoantigen bacteriophage phiX174 (phage). CSRD patients showed uniformly absent or markedly reduced switched memory B cells (IgM-IgD-CD27+). CD40L patients had reduced CD27+ memory B cells (both non-switched and switched). In NEMO patients, results varied depending on the IKKγ gene variant. Three of four AID patients had normal percentages of CD27+ memory B cells while CD27+IgM-IgD- switched memory B cells were markedly reduced in all AID patients. Antibody response to phage was remarkably decreased with lack of memory amplification and class-switching in immunized CD40L, UNG deficient, and NEMO patients. Distinct B-cell phenotype pattern correlated with abnormal antibody responses to a T-cell dependent neoantigen, representing a powerful tool to identify CSRD patients.

Clinical, Immunological, and Functional Characterization of Six Patients with Very High IgM Levels.

Very high IgM levels represent the hallmark of hyper IgM (HIGM) syndromes, a group of primary immunodeficiencies (PIDs) characterized by susceptibility to infections and malignancies. Other PIDs not fulfilling the diagnostic criteria for HIGM syndromes can also be characterized by high IgM levels and susceptibility to malignancies. The aim of this study is to characterize clinical phenotype, immune impairment, and pathogenic mechanism in six patients with very high IgM levels in whom classical HIGM syndromes were ruled out. The immunological analysis included extended B-cell immunophenotyping, evaluation of class switch recombination and somatic hypermutation, and next generation sequencing (NGS). Recurrent or severe infections and chronic lung changes at the diagnosis were reported in five out of six and two out of six patients, respectively. Five out of six patients showed signs of lymphoproliferation and four patients developed malignancies. Four patients showed impaired B-cell homeostasis. Class switch recombination was functional in vivo in all patients. NGS revealed, in one case, a pathogenic mutation in PIK3R1. In a second case, the ITPKB gene, implicated in B- and T-cell development, survival, and activity was identified as a potential candidate gene. Independent of the genetic basis, very high IgM levels represent a risk factor for the development of recurrent infections leading to chronic lung changes, lymphoproliferation, and high risk of malignancies.

Increased activation of PI3 kinase-δ predisposes to B-cell lymphoma.

Activated phosphatidylinositol 3-kinase-δ (PI3K-δ) syndrome (APDS) is a rare primary combined immunodeficiency caused by either dominant gain-of-function mutations in the PIK3CD gene encoding the catalytic subunit p110δ of PI3K-δ (referred to as type 1 APDS) or dominant loss-of-function mutations in the PIK3R1 gene encoding the p85α, p55α, and p50α regulatory subunits (type 2 APDS). In types 1 and 2 APDS, the PI3K-δ hyperactivity resulting from the gene mutations leads to similar clinical presentations, characterized by increased susceptibility to bacterial and viral infections and (to a lesser extent) autoimmune manifestations. A hallmark of this disease is lymphoproliferation, which may even be life threatening and require repeated surgical treatment. A major complication of APDS is malignancy (especially B-cell lymphomas), which greatly worsens the prognosis. Here, we review the different neoplastic conditions observed in patients with APDS and discuss the uncontrolled PI3K-δ activity in B and T cells that leads to malignant transformation.

Hematopoietic stem cell transplantation for progressive combined immunodeficiency and lymphoproliferation in patients with activated phosphatidylinositol-3-OH kinase δ syndrome type 1.

BACKGROUND: Activated phosphatidylinositol-3-OH kinase δ syndrome type 1 (APDS1) is a recently described primary immunodeficiency syndrome characterized by recurrent respiratory tract infections, lymphoid hyperplasia, and Herpesviridae infections caused by germline gain-of-function mutations of PIK3CD. Hematopoietic stem cell transplantation (HSCT) can be considered to ameliorate progressive immunodeficiency and associated malignancy, but appropriate indications, methods, and outcomes of HSCT for APDS1 remain undefined. OBJECTIVE: Our objective was to analyze the clinical manifestations, laboratory findings, prognosis, and treatment of APDS1 and explore appropriate indications and methods of HSCT. METHODS: We reviewed retrospectively the medical records of cohorts undergoing HSCT at collaborating facilities. RESULTS: Thirty-year overall survival was 86.1%, but event-free survival was 39.6%. Life-threatening events, such as severe infections or lymphoproliferation, were frequent in childhood and adolescence and were common indications for HSCT. Nine patients underwent HSCT with fludarabine-based reduced-intensity conditioning. Seven patients survived after frequent adverse complications and engraftment failure. Most symptoms improved after HSCT. CONCLUSION: Patients with APDS1 showed variable clinical manifestations. Life-threatening progressive combined immunodeficiency and massive lymphoproliferation were common indications for HSCT. Fludarabine-based reduced-intensity conditioning-HSCT ameliorated clinical symptoms, but transplantation-related complications were frequent, including graft failure.

Loss of ARHGEF1 causes a human primary antibody deficiency.

ARHGEF1 is a RhoA-specific guanine nucleotide exchange factor expressed in hematopoietic cells. We used whole-exome sequencing to identify compound heterozygous mutations in ARHGEF1, resulting in the loss of ARHGEF1 protein expression in 2 primary antibody-deficient siblings presenting with recurrent severe respiratory tract infections and bronchiectasis. Both ARHGEF1-deficient patients showed an abnormal B cell immunophenotype, with a deficiency in marginal zone and memory B cells and an increased frequency of transitional B cells. Furthermore, the patients' blood contained immature myeloid cells. Analysis of a mediastinal lymph node from one patient highlighted the small size of the germinal centers and an abnormally high plasma cell content. On the molecular level, T and B lymphocytes from both patients displayed low RhoA activity and low steady-state actin polymerization (even after stimulation of lysophospholipid receptors). As a consequence of disturbed regulation of the RhoA downstream target Rho-associated kinase I/II (ROCK), the patients' lymphocytes failed to efficiently restrain AKT phosphorylation. Enforced ARHGEF1 expression or drug-induced activation of RhoA in the patients' cells corrected the impaired actin polymerization and AKT regulation. Our results indicate that ARHGEF1 activity in human lymphocytes is involved in controlling actin cytoskeleton dynamics, restraining PI3K/AKT signaling, and confining B lymphocytes and myelocytes within their dedicated functional environment.

Clinical spectrum and features of activated phosphoinositide 3-kinase δ syndrome: A large patient cohort study.

BACKGROUND: Activated phosphoinositide 3-kinase δ syndrome (APDS) is a recently described combined immunodeficiency resulting from gain-of-function mutations in PIK3CD, the gene encoding the catalytic subunit of phosphoinositide 3-kinase δ (PI3Kδ). OBJECTIVE: We sought to review the clinical, immunologic, histopathologic, and radiologic features of APDS in a large genetically defined international cohort. METHODS: We applied a clinical questionnaire and performed review of medical notes, radiology, histopathology, and laboratory investigations of 53 patients with APDS. RESULTS: Recurrent sinopulmonary infections (98%) and nonneoplastic lymphoproliferation (75%) were common, often from childhood. Other significant complications included herpesvirus infections (49%), autoinflammatory disease (34%), and lymphoma (13%). Unexpectedly, neurodevelopmental delay occurred in 19% of the cohort, suggesting a role for PI3Kδ in the central nervous system; consistent with this, PI3Kδ is broadly expressed in the developing murine central nervous system. Thoracic imaging revealed high rates of mosaic attenuation (90%) and bronchiectasis (60%). Increased IgM levels (78%), IgG deficiency (43%), and CD4 lymphopenia (84%) were significant immunologic features. No immunologic marker reliably predicted clinical severity, which ranged from asymptomatic to death in early childhood. The majority of patients received immunoglobulin replacement and antibiotic prophylaxis, and 5 patients underwent hematopoietic stem cell transplantation. Five patients died from complications of APDS. CONCLUSION: APDS is a combined immunodeficiency with multiple clinical manifestations, many with incomplete penetrance and others with variable expressivity. The severity of complications in some patients supports consideration of hematopoietic stem cell transplantation for severe childhood disease. Clinical trials of selective PI3Kδ inhibitors offer new prospects for APDS treatment.

Clinical and immunologic phenotype associated with activated phosphoinositide 3-kinase δ syndrome 2: A cohort study.

BACKGROUND: Activated phosphoinositide 3-kinase δ syndrome (APDS) 2 (p110δ-activating mutations causing senescent T cells, lymphadenopathy, and immunodeficiency [PASLI]-R1), a recently described primary immunodeficiency, results from autosomal dominant mutations in PIK3R1, the gene encoding the regulatory subunit (p85α, p55α, and p50α) of class IA phosphoinositide 3-kinases. OBJECTIVES: We sought to review the clinical, immunologic, and histopathologic phenotypes of APDS2 in a genetically defined international patient cohort. METHODS: The medical and biological records of 36 patients with genetically diagnosed APDS2 were collected and reviewed. RESULTS: Mutations within splice acceptor and donor sites of exon 11 of the PIK3R1 gene lead to APDS2. Recurrent upper respiratory tract infections (100%), pneumonitis (71%), and chronic lymphoproliferation (89%, including adenopathy [75%], splenomegaly [43%], and upper respiratory tract lymphoid hyperplasia [48%]) were the most common features. Growth retardation was frequently noticed (45%). Other complications were mild neurodevelopmental delay (31%); malignant diseases (28%), most of them being B-cell lymphomas; autoimmunity (17%); bronchiectasis (18%); and chronic diarrhea (24%). Decreased serum IgA and IgG levels (87%), increased IgM levels (58%), B-cell lymphopenia (88%) associated with an increased frequency of transitional B cells (93%), and decreased numbers of naive CD4 and naive CD8 cells but increased numbers of CD8 effector/memory T cells were predominant immunologic features. The majority of patients (89%) received immunoglobulin replacement; 3 patients were treated with rituximab, and 6 were treated with rapamycin initiated after diagnosis of APDS2. Five patients died from APDS2-related complications. CONCLUSION: APDS2 is a combined immunodeficiency with a variable clinical phenotype. Complications are frequent, such as severe bacterial and viral infections, lymphoproliferation, and lymphoma similar to APDS1/PASLI-CD. Immunoglobulin replacement therapy, rapamycin, and, likely in the near future, selective phosphoinositide 3-kinase δ inhibitors are possible treatment options.

A homozygous PMS2 founder mutation with an attenuated constitutional mismatch repair deficiency phenotype.

BACKGROUND: Inherited mutations in DNA mismatch repair genes predispose to different cancer syndromes depending on whether they are mono-allelic or bi-allelic. This supports a causal relationship between expression level in the germline and phenotype variation. As a model to study this relationship, our study aimed to define the pathogenic characteristics of a recurrent homozygous coding variant in PMS2 displaying an attenuated phenotype identified by clinical genetic testing in seven Inuit families from Northern Quebec. METHODS: Pathogenic characteristics of the PMS2 mutation NM_000535.5:c.2002A>G were studied using genotype-phenotype correlation, single-molecule expression detection and single genome microsatellite instability analysis. RESULTS: This PMS2 mutation generates a de novo splice site that competes with the authentic site. In homozygotes, expression of the full-length protein is reduced to a level barely detectable by conventional diagnostics. Median age at primary cancer diagnosis is 22 years among 13 NM_000535.5:c.2002A>G homozygotes, versus 8 years in individuals carrying bi-allelic truncating mutations. Residual expression of full-length PMS2 transcript was detected in normal tissues from homozygotes with cancers in their 20s. CONCLUSIONS: Our genotype-phenotype study of c.2002A>G illustrates that an extremely low level of PMS2 expression likely delays cancer onset, a feature that could be exploited in cancer preventive intervention.

The CARD11-BCL10-MALT1 (CBM) signalosome complex: Stepping into the limelight of human primary immunodeficiency.

Next-generation DNA sequencing has accelerated the genetic characterization of many human primary immunodeficiency diseases (PIDs). These discoveries can be lifesaving for the affected patients and also provide a unique opportunity to study the effect of specific genes on human immune function. In the past 18 months, a number of independent groups have begun to define novel PIDs caused by defects in the caspase recruitment domain family, member 11 (CARD11)-B-cell chronic lymphocytic leukemia/lymphoma 10 (BCL10)-mucosa-associated lymphoid tissue lymphoma translocation gene 1 (MALT1 [CBM]) signalosome complex. The CBM complex forms an essential molecular link between the triggering of cell-surface antigen receptors and nuclear factor κB activation. Germline mutations affecting the CBM complex are now recognized as the cause of novel combined immunodeficiency phenotypes, which all share abnormal nuclear factor κB activation and dysregulated B-cell development as defining features. For this "Current perspectives" article, we have engaged experts in both basic biology and clinical immunology to capture the worldwide experience in recognizing and managing patients with PIDs caused by CBM complex mutations.

A human immunodeficiency caused by mutations in the PIK3R1 gene.

Recently, patient mutations that activate PI3K signaling have been linked to a primary antibody deficiency. Here, we used whole-exome sequencing and characterized the molecular defects in 4 patients from 3 unrelated families diagnosed with hypogammaglobulinemia and recurrent infections. We identified 2 different heterozygous splice site mutations that affect the same splice site in PIK3R1, which encodes the p85α subunit of PI3K. The resulting deletion of exon 10 produced a shortened p85α protein that lacks part of the PI3K p110-binding domain. The hypothetical loss of p85α-mediated inhibition of p110 activity was supported by elevated phosphorylation of the known downstream signaling kinase AKT in patient T cell blasts. Analysis of patient blood revealed that naive T and memory B cell counts were low, and T cell blasts displayed enhanced activation-induced cell death, which was corrected by addition of the PI3Kδ inhibitor IC87114. Furthermore, B lymphocytes proliferated weakly in response to activation via the B cell receptor and TLR9, indicating a B cell defect. The phenotype exhibited by patients carrying the PIK3R1 splice site mutation is similar to that of patients carrying gain-of-function mutations in PIK3CD. Our results suggest that PI3K activity is tightly regulated in T and B lymphocytes and that various defects in the PI3K-triggered pathway can cause primary immunodeficiencies.

Defective anti-polysaccharide response and splenic marginal zone disorganization in ALPS patients.

Autoimmune lymphoproliferative syndrome (ALPS) caused by impaired FAS-mediated apoptosis of lymphocytes is characterized by lymphoproliferation, autoimmunity, but also an increased risk of invasive bacterial infection, notably following splenectomy. We surveyed a cohort of 100 ALPS patients (including 33 splenectomized) and found that 12 (10 splenectomized) had experienced 23 invasive bacterial infections mainly caused by Streptococcus pneumoniae. This vulnerability was associated with evidence of defective B-cell function characterized by low serum immunoglobulin (Ig) M, low IgM antibody production in response to S pneumoniae following nonconjugated immunization, and low blood memory B-cells counts (including marginal zone [MZ] B-cell counts). This immunodeficiency strongly correlated with intensity of lymphoproliferation. Spleen sections from 9 ALPS patients revealed double-negative T-cell (DN-T) infiltration of the MZ, which was depleted of B cells. MZ in ALPS patients contained an abnormally thick layer of MAdCAM-1((+)) stromal cells and an excess of DN-Ts. DN-Ts were shown to express MAdCAM-1 ligand, the α4ß7 integrin. These observations suggest that accumulating DN-Ts are trapped within stromal cell meshwork and interfere with correct localization of MZ B cells. Similar observations were made in spleens of fas-deficient mice. Our data revealed an unexpected mechanism by which ALPS results in anti-polysaccharide IgM antibody production-specific defect. Splenectomy should be avoided.

Analysis of somatic hypermutations in the IgM switch region in human B cells.

BACKGROUND: The molecular mechanism of class-switch recombination (CSR) in human subjects has not been fully elucidated. The CSR-induced mutations occurring in the switch region of the IgM gene (Smu-SHMs) in in vitro CSR-activated and in vivo switched B cells have been analyzed in mice but not in human subjects. OBJECTIVE: We sought to better characterize the molecular mechanism of CSR in human subjects. METHODS: Smu-SHMs were analyzed in vitro and in vivo by using healthy control subjects and patients with molecularly defined CSR defects. RESULTS: We found that Smu-SHMs can be induced in vitro by means of CSR activation in human subjects. We also found large amounts of Smu-SHMs in in vivo class-switched memory B cells, smaller (although significant) amounts in unswitched memory B cells, and very low amounts in naive B cells. In class-switched memory B cells a high frequency of Smu-SHMs was found throughout the Smu. In unswitched memory B cells, the Smu-SHM frequency was significantly decreased in the 5' part of the Smu. The difference between switched and unswitched B cells suggests that the extension of somatic hypermutation (SHM) to the 5' upstream region of the Smu might be associated with the effective induction of CSR. The analysis of the pattern of mutations within and outside the WRCY/RGYW (W, A/T; R, A/G; and Y, C/T) motifs, as well as the Smu-SHMs, in CD27(+) B cells from CD40 ligand (CD40L)-, activation-induced cytidine deaminase (AID)-, and uracil-DNA glycosylase (UNG)-deficient patients revealed the dependence of Smu-SHM on CD40L, AID, UNG, and the mismatch repair system in human subjects. CONCLUSION: CD40L-, AID-, UNG-, and mismatch repair system-dependent Smu-SHMs and extension to the 5' region of Smu are necessary to accomplish effective CSR in human subjects.

First report of the Hyper-IgM syndrome Registry of the Latin American Society for Immunodeficiencies: novel mutations, unique infections, and outcomes.

Hyper-IgM (HIGM) syndrome is a heterogeneous group of disorders characterized by normal or elevated serum IgM levels associated with absent or decreased IgG, IgA and IgE. Here we summarize data from the HIGM syndrome Registry of the Latin American Society for Immunodeficiencies (LASID). Of the 58 patients from 51 families reported to the registry with the clinical phenotype of HIGM syndrome, molecular defects were identified in 37 patients thus far. We retrospectively analyzed the clinical, immunological and molecular data from these 37 patients. CD40 ligand (CD40L) deficiency was found in 35 patients from 25 families and activation-induced cytidine deaminase (AID) deficiency in 2 unrelated patients. Five previously unreported mutations were identified in the CD40L gene (CD40LG). Respiratory tract infections, mainly pneumonia, were the most frequent clinical manifestation. Previously undescribed fungal and opportunistic infections were observed in CD40L-deficient patients but not in the two patients with AID deficiency. These include the first cases of pneumonia caused by Mycoplasma pneumoniae, Serratia marcescens or Aspergillus sp. and diarrhea caused by Microsporidium sp. or Isospora belli. Except for four CD40L-deficient patients who died from complications of presumptive central nervous system infections or sepsis, all patients reported in this study are alive. Four CD40L-deficient patients underwent successful bone marrow transplantation. This report characterizes the clinical and genetic spectrum of HIGM syndrome in Latin America and expands the understanding of the genotype and phenotype of this syndrome in tropical areas.

Naive and memory human B cells have distinct requirements for STAT3 activation to differentiate into antibody-secreting plasma cells.

Long-lived antibody memory is mediated by the combined effects of long-lived plasma cells (PCs) and memory B cells generated in response to T cell-dependent antigens (Ags). IL-10 and IL-21 can activate multiple signaling pathways, including STAT1, STAT3, and STAT5; ERK; PI3K/Akt, and potently promote human B cell differentiation. We previously showed that loss-of-function mutations in STAT3, but not STAT1, abrogate IL-10- and IL-21-mediated differentiation of human naive B cells into plasmablasts. We report here that, in contrast to naive B cells, STAT3-deficient memory B cells responded to these STAT3-activating cytokines, differentiating into plasmablasts and secreting high levels of IgM, IgG, and IgA, as well as Ag-specific IgG. This was associated with the induction of the molecular machinery necessary for PC formation. Mutations in IL21R, however, abolished IL-21-induced responses of both naive and memory human B cells and compromised memory B cell formation in vivo. These findings reveal a key role for IL-21R/STAT3 signaling in regulating human B cell function. Furthermore, our results indicate that the threshold of STAT3 activation required for differentiation is lower in memory compared with naive B cells, thereby identifying an intrinsic difference in the mechanism underlying differentiation of naive versus memory B cells.