Low B-cell Counts Research Articles

Intrinsic functional defects in B cells of patients with NFKB2 mutations.

Mutations in the human nuclear factor-κB2 gene (NFKB2) are associated with common variable immunodeficiency (CVID) or combined immunodeficiency diseases (CID), characterized by B-cell lymphopenia, hypogammaglobulinemia, and T cell dysfunction. This study investigated whether B cells with NFKB2 mutations exhibit intrinsic impairments in activation, class-switch recombination, and differentiation. We analyzed five patients from four unrelated families with CVID, each carrying a heterozygous NFKB2 mutation: P1 (C.2595_2614del, p.A867Gfs*12), P2 (C.2597G>A, p.S866N), P3 (C.2540dupT, p.R848Efs*38), and P4 and P5 (C.2570_2571insCAGCACA, p.A860Qfs*28). The patients with frameshift mutations (P1, P3, P4, and P5) exhibited truncated proteins detectable in their peripheral blood mononuclear cells, while P2 had a missense mutation. All identified mutations disrupted the processing of p100 into the active p52 form, resulting in NF-κB2 loss-of-function and IκBδ gain-of-function. Clinically, P1, P2, and P3 exhibited B-cell lymphopenia, and all five patients presented with hypogammaglobulinemia. Notably, P2 exhibited a markedly low B-cell count, associated with increased proportions of memory B and IgD-CD27- double negative B cells. In vitro experiments with naïve B cells from P1 and P4 demonstrated decreased survival, impaired activation, and reduced differentiation into CD27+IgD- cells and plasmablasts, while class switch recombination was unaffected. These findings reveal novel B cell-intrinsic functional defects in patients with NFKB2 mutations.

Variants in IGLL1 cause a broad phenotype from agammaglobulinemia to transient hypogammaglobulinemia

BackgroundAgammaglobulinemia due to variants in IGLL1 has traditionally been considered an exceedingly rare form of severe B-cell deficiency, with only eight documented cases in the literature. Surprisingly, the first agammaglobulinemic patient identified by newborn screening (NBS) through quantification of kappa-deleting recombination excision circles harbored variants in IGLL1. ObjectiveTo provide a comprehensive overview of the clinical and immunological findings of patients with B-cell deficiency attributed to variants in IGLL1. MethodsNBS programs reporting using kappa-deleting recombination excision circle assays, the European Society for Immunodeficiencies Registry, and authors of published reports featuring patients with B-cell deficiency linked to IGLL1 variants were contacted. Only patients with (likely) pathogenic variants, reduced CD19+ counts and no alternative diagnosis were included. ResultsThe study included 13 patients identified through NBS, two clinically diagnosed patients, and two asymptomatic siblings. All had severely reduced CD19+ B-cells (< 0.1×109/L) on first evaluation, yet subsequent follow-ups indicated residual immunoglobulin production. Specific antibody responses to vaccine antigens varied, with a predominant reduction observed during infancy. Clinical outcomes were favorable with immunoglobulin G substitution. Two patients successfully discontinued substitution without developing susceptibility to infections and maintaining immunoglobulin levels. The pooled incidence of homozygous or compound heterozygous pathogenic IGLL1 variants identified by NBS in Austria, Czechia, and Switzerland was 1.3:100´000, almost double of X-linked agammaglobulinemia. ConclusionB-cell deficiency resulting from IGLL1 variants appears to be more prevalent than initially believed. Despite markedly low B-cell counts, the clinical course in some patients may be milder than reported in the literature so far.

B-cell immune deficiency in twin sisters expands the phenotype of MOPDI.

Microcephalic osteodysplastic primordial dwarfism type I (MOPDI) is a very rare and severe autosomal recessive disorder characterized by marked intrauterine growth retardation, skeletal dysplasia, microcephaly and brain malformations. MOPDI is caused by biallelic mutations in RNU4ATAC, a non-coding gene involved in U12-type splicing of 1% of the introns in the genome, which are recognized by their specific splicing consensus sequences. Here, we describe a unique observation of immunodeficiency in twin sisters with mild MOPDI, who harbor a novel n.108_126del mutation, encompassing part of the U4atac snRNA 3' stem-loop and Sm protein binding site, and the previously reported n.111G>A mutation. Interestingly, both twin sisters show mild B-cell anomalies, including low naive B-cell counts and increased memory B-cell and plasmablasts counts, suggesting partial and transitory blockage of B-cell maturation and/or excessive activation of naive B-cells. Hence, the localization of a mutation in stem II of U4atac snRNA, as observed in another RNU4ATAC-opathy with immunodeficiency, that is, Roifman syndrome (RFMN), is not required for the occurrence of an immune deficiency. Finally, we emphasize the importance of considering immunodeficiency in MOPDI management to reduce the risk of serious infectious episodes.

Correlation Analysis of Peripheral Blood B Cell Count with Clinical Features and Prognosis in Patients Newly Diagnosed with Diffuse Large B-Cell Lymphoma

To explore the correlation between peripheral blood B cell count and clinical features and prognosis of patients with newly diagnosed diffuse large B-cell lymphoma (DLBCL). The relationship of peripheral blood B cell count with clinical features, laboratory indexes and prognosis in 67 patients with newly diagnosed DLBCL was retrospectively analyzed. Patients were divided into low B-cell count group (B cell＜0.1×109/L, n=34) and high B-cell count group (B cell≥0.1×109/L, n=33) according to the median B cell count values. Compared with the high B cell count group, the low B cell count group had a higher proportion of patients with Lugano stage III-IV, elevated LDH, elevated β2-MG and IPI score 3-5 and increased CRP (P =0.033, 0.000, 0.023, 0.001, 0.033). The peripheral CD3+ and CD4+ cell counts of patients in the low B cell count group were significantly lower than those in the high B cell count group (P =0.010, 0.017). After initial treatment, overall response rate (ORR) and complete remission (CR) rate in high B cell count group were significantly higher than those in low B cell count group (P =0.032, 0.013). The median follow-up time of patients was 23(2-77) months, progression-free survival (PFS) and overall survival (OS) of patients in the high B cell count group were significantly better than those in the low B cell count group (P =0.001, 0.002). Univariate analysis showed that pretreatment low B cell count in the peripheral blood was associated with shortened PFS and OS (HR=4.108, P =0.002; HR=8.218, P =0.006). Multivariate analysis showed that low B cell count was an independent prognostic factor for shortened PFS (HR=3.116, P =0.037). Decreased peripheral blood B cell count in newly diagnosed DLBCL patients is associated with high-risk clinical features and may affect the efficacy of immunochemotherapy, which is associated with poor clinical prognosis.

Low circulating B cells in immunocompromised individuals are linked to poorer antibody responses to vaccines and a predisposition to viral infections.

B cells play an important role in protection against viral infections, not only through the production of antibodies but also through their ability to act as antigen-presenting cells and produce cytokines. To assess whether there is a link between low circulating B-cell counts and a predisposition to viral infections in immunocompromised individuals, we performed a retrospective cohort analysis at 2 National Health Service Clinical Immunology sites in England. Eligible patients were adults who were either diagnosed with or under investigation for an immunodeficiency and had recorded circulating B-cell counts. Information on viral infections was collected by using the departmental, hospital, and laboratory electronic information systems. Ageneralized linear model was used to analyze the relationship between B-cell counts and relevant indices of viral infection while controlling for patient age, diagnosis group, and T-cell and natural killer cell counts. A total of 376 eligible patients were identified, 134 of whom had B-cell counts that were below the laboratory-defined refence range (<0.11 ×109/L). Patients with low numbers of circulating B cells had lower pretreatment immunoglobulin levels and poorer antibody responses to vaccines (Streptococcus pneumonia, Clostridium tetani, and Haemophilus influenzae type B). An increased number of chronic or recurrent (P= .001), severe or unusual (P= .001), and PCR-confirmed viral infections (P= .04) were recorded in these patients versus in those with normal numbers of circulating B cells. Overall, there was a statistically significant association between low circulating B-cell counts and the incidence of clinically important viral infections in this patient cohort, even when controlling for relevant covariates. Clinicians caring for patients with immunodeficiency should be vigilant for these types of infections, particularly in patients with low peripheral B-cell counts. Aprospective study will be required to confirm these findings.

Low Peripheral B-Cell Counts in Patients With Systemic Rheumatic Diseases Due to Treatment With Belimumab and/or Rituximab Are Associated With Low Antibody Responses to Primary COVID-19 Vaccination.

Background: Immunosuppressive agents inhibit COVID-19 vaccine antibody (Ab) responses in patients with systemic rheumatic diseases. Rituximab may fully block Ab responses when B cells become undetected. The effect of detected but low number of B cells due to treatment with a B-cell agent (belimumab and/or rituximab) has not been established. Purpose: We sought to examine whether there is an association between a low number of B cells due to treatment with belimumab and/or rituximab and impaired primary COVID-19 vaccination spike Ab responses in patients with systemic rheumatic diseases. Methods: We retrospectively examined Ab responses to COVID-19 vaccinations, especially in relation to B-cell counts after treatment with belimumab and/or rituximab, in 58 patients with systemic rheumatic diseases: 22 on and 36 not on B-cell agents. We used Kruskal-Wallis and Mann-Whitney U tests for comparison of Ab values between the groups and Fisher exact test for relative risk calculations. Results: Median (interquartile range) postvaccination Ab responses were lower in patients on versus those not on B-cell agents: 3.91 (0.77-20.00) versus 20.00 (14.32-20.00), respectively. Among patients on belimumab and/or rituximab, Ab responses of less than 25% of the assay's upper limit were exclusively observed in those with B-cell counts lower than 40/µL. Patients with B-cell counts lower than 40/µL exhibit a relative risk of 6.092 (95% CI: 2.75-14.24) for Ab responses of less than 25% of the upper limit compared with patients not on B-cell agents. This relative risk remained significant, even after excluding patients with undetected B cells. Conclusion: This retrospective study found an association between low B-cell counts (less than 40/µL) and decreased Ab responses to primary COVID-19 vaccination in patients with systemic rheumatic diseases treated with belimumab and/or rituximab. Despite the small number of patients studied, these findings add to the accumulating evidence on the importance of B-cell count in predicting spike Ab responses to COVID-19 vaccination.

Humoral Response to SARS-CoV-2 Antigen in Patients Treated with Monoclonal Anti-CD20 Antibodies: It Is Not All about B Cell Recovery.

Anti-CD20 therapies decrease the humoral response to SARS-CoV-2 immunization. We aimed to determine the extent of the humoral response to SARS-CoV-2 antigens in correlation with peripheral B-cell dynamics among patients with central nervous system inflammatory disorders treated with anti-CD20 medications. We retrospectively included patients receiving anti-CD20 therapy after antigen contact who were divided into responders (&gt;7 binding antibody units (BAU)/mL) and non-responders (&lt;7 BAU/mL). In participants with first antigen contact prior to therapy, we investigated the recall response elicited once under treatment. We included 80 patients (responders n = 34, non-responders n = 37, recall cohort n = 9). The B-cell counts among responders were significantly higher compared to non-responders (mean 1012 cells/µL ± SD 105 vs. mean 17 cells/µL ± SD 47; p &lt; 0.001). Despite very low B-cell counts (mean 9 cells/µL ± SD 20), humoral response was preserved among the recall cohort (mean 1653 BAU/mL ± SD 2250.1) and did not differ significantly from responders (mean 735 BAU/mL ± SD 1529.9; p = 0.14). Our data suggest that peripheral B cells are required to generate antibodies to neo-antigens but not for a recall response during anti-CD20 therapy. Evaluation of B-cell counts and pre-existing SARS-CoV-2 antibodies might serve as biomarkers for estimating the immune competence to mount a humoral response to SARS-CoV-2 antigens.

Iberdomide (IBER) in Combination with Dexamethasone (DEX) in Relapsed/Refractory Multiple Myeloma (RRMM): Results from the Anti-B-Cell Maturation Antigen (BCMA)-Exposed Cohort of the CC-220-MM-001 Trial

Introduction: IBER, a novel, oral cereblon E3 ligase modulator (CELMoD®), has greater tumoricidal and immune-stimulatory effects compared with immunomodulatory drugs (IMiDs®), and has shown marked synergy with DEX and other antimyeloma therapies in preclinical models. IBER is being investigated in the phase 1/2 CC-220-MM-001 (NCT02773030) and phase 3 EXCALIBER-RRMM (NCT04975997) studies with different treatment combinations in patients (pts) with RRMM. In phase 1 of CC-220-MM-001, IBER + DEX showed notable efficacy (overall response rate [ORR] 31.9%) and a manageable safety profile. Given the emergence of novel anti-BCMA therapies in RRMM, assessment of pts with prior anti-BCMA exposure is an important consideration. Here we report results from the dose-expansion phase of IBER + DEX in heavily pretreated, triple-class-exposed (including ≥ 1 IMiD agent, ≥ 1 proteasome inhibitor [PI], and ≥ 1 anti-CD38 monoclonal antibody [mAb]) pts with RRMM who had also received prior anti-BCMA therapy. Methods: Eligible pts had RRMM, had received ≥ 3 prior lines of therapy (including lenalidomide or pomalidomide, a PI, an anti-CD38 mAb, and an anti-BCMA therapy) and had documented progressive disease (PD) on or within 60 days of their last antimyeloma therapy (documented PD if chimeric antigen receptor [CAR] T cell therapy was the last therapy). IBER 1.6 mg was given orally on days 1-21 of each 28-day cycle, plus weekly DEX (40 mg; 20 mg if > 75 years of age). The primary objectives were to determine preliminary efficacy (ORR) and safety. Results: As of April 15, 2022, 38 pts had received IBER + DEX in the anti-BCMA-exposed cohort. Median age was 65 (range 50-78) years and median time since initial diagnosis was 7.8 (0.6-24.8) years. High-risk cytogenetics were present in 31.6% of pts (52.6% pts were not evaluable), and 23.7% of pts had extramedullary plasmacytomas. Median number of prior regimens was 7 (4-15). All pts were triple-class exposed. Prior anti-BCMA therapies included CAR T cell therapy (36.8%), antibody-drug conjugates (34.2%), and T-cell engagers (TCEs; 23.7%) (Table); 78.9% of pts were refractory to the last antimyeloma regimen and 84.2% were triple-class refractory. Median follow-up was 8.1 (range 1.5-24.2) months, with a median number of 3.5 (1-19) cycles received and 21.1% of pts continuing treatment. Discontinuation was due mainly to PD, reported in 68.4% of pts. ORR (≥ partial response) was 36.8%, with 2 (5.3%) complete responses, 5 (13.2%) very good partial responses, and 7 (18.4%) partial responses. Clinical benefit rate (≥ minimal response) was 39.5%. Responses were observed regardless of prior anti-BCMA therapy (Figure; updated data will be presented at the congress). Median duration of response was 7.5 (95% confidence interval [CI] 3.2-not reached) months, median progression-free survival was 2.4 (95% CI 2.1-4.2) months, and median time to response was 1.4 (range 0.9-5.4) months. Grade (Gr) 3/4 treatment-emergent adverse events (TEAEs) occurred in 30 (78.9%) pts and were mostly hematologic; most frequent (≥ 20% pts) were neutropenia (50.0%, with 5.3% cases of febrile neutropenia), anemia (28.9%), leukopenia (23.7%), and thrombocytopenia (21.1%). Gr 3/4 infections occurred in 23.7% of pts and included pneumonia (21.1%); the occurrence of other Gr 3/4 non-hematologic TEAEs was low and included hypokalemia, hypertension, and mood alterations (all 5.3%). Two (5.3%) deaths were reported (due to sepsis and PD) which were not considered related to study treatment. IBER dose interruptions and reductions occurred in 24 (63.2%) and 7 (18.4%) pts, respectively. No pts discontinued IBER due to TEAEs. Immunophenotyping showed comparable immunodeficiency (low absolute B-cell counts, T-cell counts, and CD4:CD8 ratio) between anti-BCMA-exposed pts and triple-class-exposed pts without prior anti-BCMA exposure; this was more pronounced following TCE-based therapies. Importantly, IBER + DEX remained immune-stimulatory in this population, increasing T- and NK-cell proliferation and T-cell activation. Conclusions: IBER + DEX showed encouraging efficacy and safety in pts with triple-class-exposed RRMM and prior anti-BCMA therapy. The results are comparable to those from Cohort D of the same study, which is assessing IBER + DEX in pts with triple-class refractory RRMM. These findings support further development of IBER in RRMM, including in anti-BCMA-exposed pts. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

Blinatumomab for treating pediatric B-lineage acute lymphoblastic leukemia: A retrospective real-world study.

Blinatumomab was shown to be safe and effective for consolidation therapy in B-cell acute lymphoblastic leukemia (B-ALL). This study aimed to investigate the effectiveness and safety of blinatumomab in pediatric B-ALL patients in a real-world setting. This was a retrospective, observational study that included patients who initiated blinatumomab treatment between October 1, 2020 and June 20, 2022. Patients with B-ALL diagnosis, age below 18 years, and at least one blinatumomab treatment cycle were included. Treatment-related toxicities were assessed. Totally 23 pediatric patients were included in this study, with a median age of 6 years (range, 2 to 11 years). Blinatumomab therapy was applied for MRD-positive (disease ≥0.01%, n = 3) or chemotherapy-ineligible (n = 20) B-ALL cases. The median follow-up time was 9 months, and all evaluable patients achieved complete molecular remission with undetectable MRD. Four relapsed B-ALL cases proceeded to hematopoietic stem cell transplantation (HSCT) without further bridging therapy, while the others underwent maintenance chemotherapy after blinatumomab treatment. Grade ≥3 febrile neutropenia, white blood cell decrease and seizure were observed in 57%, 48% and 4.3% of patients, respectively. One case discontinued therapy due to neurologic toxicities. Elevated cytokine levels were observed in 4 patients. In all 23 patients, increased T-cell and low B-cell counts (<10/μl) were detected during blinatumomab therapy. These encouraging results suggest blinatumomab in pediatric B-ALL patients with MRD+ or chemotherapy-related toxicities is effective and safe in the short run, although long-term follow-up is still needed.

Two Doses of BNT162b2 mRNA Vaccine in Patients after Hematopoietic Stem Cell Transplantation: Humoral Response and Serological Conversion Predictors.

Simple SummaryVaccination against SARS-CoV-2 is currently the best tool in the fight against the COVID-19 pandemic. However, there are concerns about its efficacy and safety after hematopoietic stem cell transplantation. The aim of the study was to study the efficacy and safety of two doses of BNT162b2 mRNA vaccine in adult patients after autologous or allogeneic transplantation. We examined the presence of anti-SARS-CoV-2 antibodies before and after vaccination. We also searched for the potential predictors of serological conversion after vaccination, including the analysis of the impact of various lymphocyte subpopulations at the time of vaccination on post-vaccine antibody concentration and seroconversion. In addition, patients were followed-up for adverse events after vaccination, and the data on breakthrough SARS-CoV-2 infection were collected. The results of our study broaden the knowledge of the efficacy and safety of BNT162b2 mRNA vaccine in patients after HCT, providing new data on serological conversion predictors.Vaccination against SARS-CoV-2 is currently the best tool in the fight against the COVID-19 pandemic. However, there are limited data on its efficacy and safety after hematopoietic stem cell transplantation (HCT). We present the results of a prospective analysis of the humoral response to two doses of BNT162b2 mRNA vaccine in 93 adult patients, including 29 after autologous HCT (autoHCT) and 64 after allogeneic HCT (alloHCT). Positive anti-SARS-CoV-2 antibodies were detected before vaccination in 25% of patients despite a negative medical history of COVID-19. Seroconversion after vaccination was achieved in 89% of patients after alloHCT and in 96% after autoHCT, without grade 3/4 adverse events. Post-vaccination anti-SARS-CoV-2 antibody level correlated with the time from transplant and absolute B-cell count at the vaccination. In univariate analysis restricted to the alloHCT group, short time since transplantation, low B-cell count, low intensity conditioning, GvHD, and immunosuppressive treatment at the vaccination were associated with lack of seroconversion. In the multivariate model, the only negative predictor of seroconversion remained treatment with calcineurin inhibitor (CNI). In conclusion, the BNT162b2 mRNA vaccine is highly immunogenic in patients after HCT, but treatment with CNI at the time of vaccination has a strong negative impact on the humoral response

A Third Anti-Sars-Cov2 mRNA Dose Does Not Overcome the Pejorative Impact of Anti-CD20 Therapy and/or Low Immunoglobulin Level in CLL/Lymphoma Patients

Introduction: Patients with lymphoma and chronic lymphocytic leukemia (CLL) share immune-deficiencies due to the biological features of these diseases per se and to their treatments. They are at risk to develop severe and/or prolonged forms of Covid-19. The efficacy of vaccination against COVID in lymphoma/CLL patients also raises specific concerns: Antibody response to mRNA COVID-19 vaccine was shown to be negatively affected by CLL stage and its treatments, especially in case of previous administration of anti-CD20 antibodies (Herishanu et al). Therefore, the addition of a third dose of vaccine in immunosuppressed patients is currently recommended in France.Methods: To analyse the determinants of the antibody response after anti-SARS-Cov2 mRNA vaccines (Pfizer or Moderna), we conducted a retrospective monocentric study among adults with a past or current lymphoma/CLL who underwent serology 2 weeks or more after 2 or 3 previous injections. The decision of a third dose was at the discretion of each physician. Data were extracted from the patients' medical charts on their demographics, lymphoma history and detailed treatments, vaccinations and biological parameters (immunoglobulin (Ig) G dosage, lymphocytic, B-cell and T-cell counts) and serology (Elecsys ® Anti-SARS-CoV-2 S). Statistical tests were two-tailed and p-values < 0.05 were considered significant. Multivariable analysis was conducted using independent variables having univariate significance below 0.1.This study was conducted in accordance with the Declaration of Helsinki, and approved by the ethics committee of our institution.Results: Ninety-one patients with non-Hodgkin's lymphomas (NHL) (n=48), CLL (32) or Hodgkin's lymphoma (11) were enrolled. With a median interval since last dose of 47 days (range 15-125), 48 patients (53%) had a negative serology (Table). Gender and age were not associated with antibody response. The proportion of seronegativity was 57% among patients with B-cell NHL, and 41% among those with CLL. Among the biological variables reflecting immune deficiency, lymphocytopenia (<1.5G/L), low B-cell counts (<0.2G/L) and IgG levels (<6g/L) were significantly associated with a higher risk of seronegativity with odds-ratios (OR, [95% confidence interval]) of 5.1 [1.8-15.3], p<7x10 -4 , 15.0 [3.0-147.9], p<7x10 -5, and 15.3 [3.7-93.0], p<9x10 ⁻6 respectively).Patients under watch and wait attitude and those who did not have a lymphoma/CLL treatment since at least 12 months before vaccination had a much lower risk of negative serology (OR: 0.08 [<0.01 ; 0.4], p< 2.10 -4). Among lymphoma/CLL therapies, chemotherapy overall or targeted therapy with BTK inhibitors or Venetoclax were not significantly associated with a lower risk of negative serology. In contrast, the administration of anti-CD20 therapy during the year before first dose of vaccine was associated with a higher risk of negative serology (OR: 4.5 [1.7; 12.1], p<8.10 -4). Of note, this treatment was also significantly associated with both lymphocytopenia and low B-cell counts. There was no association between the number of vaccine doses and the risk of negative serology, 59% of the patients who received 3 doses remained seronegative.A multivariable analysis associating IgG level and treatment history showed that negative serology is significantly associated with a low level of IgG (OR: 25.74 [5.58; 201.14], p < 10 -3) and anti-CD20 treatment (OR: 28.70 [4.14, 382.60], p = 3x10 -3).Conclusion: Overall, previous anti-CD20 therapy and low IgG levels are the main independent factors associated with a lack of serological response after anti-SARS-Cov2 mRNA vaccine. Administration of a third vaccine does not overcome their pejorative impact. This should contribute to the elaboration of guidelines for the management of lymphoma/CLL patients during the Covid-19 era. In particular, in all non-critical clinical situation, SARS-CoV-2 vaccination should be proposed before the onset of lymphoma/CLL therapy. Meanwhile, individuals with CLL/lymphoma should receive the Covid-19 vaccine, be informed that they are unlikely to be protected and continue social distancing and adhere to other proven mitigation strategies. Systematic vaccination of their proxies and hospital workers should also benefit directly to patients. [Display omitted] DisclosuresRigaudeau: Takeda: Membership on an entity's Board of Directors or advisory committees.

B-Cell Regeneration Profile and Minimal Residual Disease Status in Bone Marrow of Treated Multiple Myeloma Patients

Simple SummaryB-cell regeneration during therapy has been associated with the outcome of multiple myeloma (MM) patients. However, the effects of therapy and hemodilution in bone marrow (BM) B-cell recovery have not been systematically evaluated. Here, we show that hemodilution is present in a significant fraction of MM BM samples, leading to lower total B-cell, B-cell precursor (BCP), and normal plasma cell (nPC) counts. Among MM BM samples, decreased percentages (vs. healthy donors) of BCP, transitional/naïve B-cell (TBC/NBC) and nPC populations were observed at diagnosis. BM BCP, but not TBC/NBC, increased after induction therapy. At day+100 post-autolo-gous stem cell transplantation, a greater increase in BCP with recovered TBC/NBC numbers but persistently low memory B-cell and nPC counts were found. At the end of therapy, complete response (CR) BM samples showed higher CD19− nPC counts vs. non-CR specimens with no clear association between BM B-cell regeneration profiles and patient outcomes.B-cell regeneration during therapy has been considered as a strong prognostic factor in multiple myeloma (MM). However, the effects of therapy and hemodilution in bone marrow (BM) B-cell recovery have not been systematically evaluated during follow-up. MM (n = 177) and adult (≥50y) healthy donor (HD; n = 14) BM samples were studied by next-generation flow (NGF) to simultaneously assess measurable residual disease (MRD) and residual normal B-cell populations. BM hemodilution was detected in 41 out of 177 (23%) patient samples, leading to lower total B-cell, B-cell precursor (BCP) and normal plasma cell (nPC) counts. Among MM BM, decreased percentages (vs. HD) of BCP, transitional/naïve B-cell (TBC/NBC) and nPC populations were observed at diagnosis. BM BCP increased after induction therapy, whereas TBC/NBC counts remained abnormally low. At day+100 postautologous stem cell transplantation, a greater increase in BCP with recovered TBC/NBC cell numbers but persistently low memory B-cell and nPC counts were found. At the end of therapy, complete response (CR) BM samples showed higher CD19− nPC counts vs. non-CR specimens. MRD positivity was associated with higher BCP and nPC percentages. Hemodilution showed a negative impact on BM B-cell distribution. Different BM B-cell regeneration profiles are present in MM at diagnosis and after therapy with no significant association with patient outcome.

High anti-JCPyV serum titers coincide with high CSF cell counts in RRMS patients

Background: Progressive multifocal leukoencephalopathy (PML) can in rare cases occur in natalizumab-treated patients with high serum anti-JCPyV antibodies, hypothetically due to excessive blockade of immune cell migration. Objective: Immune cell recruitment to the central nervous system (CNS) was assessed in relapsing-remitting multiple sclerosis (RRMS) patients stratified by low versus high anti-JCPyV antibody titers as indicator for PML risk. Methods: Cerebrospinal fluid (CSF) cell counts of 145 RRMS patients were quantified by flow cytometry. Generalized linear models were employed to assess influence of age, sex, disease duration, Expanded Disability Status Scale (EDSS), clinical/radiological activity, current steroid or natalizumab treatment, as well as anti-JCPyV serology on CSF cell subset counts. Results: While clinical/radiological activity was associated with increased CD4, natural killer (NK), B and plasma cell counts, natalizumab therapy reduced all subpopulations except monocytes. With and without natalizumab therapy, patients with high anti-JCPyV serum titers presented with increased CSF T-cell counts compared to patients with low anti-JCPyV serum titers. In contrast, PML patients assessed before (n = 2) or at diagnosis (n = 5) presented with comparably low CD8 and B-cell counts, which increased after plasma exchange (n = 4). Conclusion: High anti-JCPyV indices, which could be indicative of increased viral activity, are associated with elevated immune cell recruitment to the CNS. Its excessive impairment in conjunction with viral activity could predispose for PML development.

Frequent altered distribution of peripheral B-lymphocyte subsets in pediatric and adolescent patients with classical Hodgkin lymphoma

Peripheral lymphopenia is a well-known negative prognostic marker in classical Hodgkin lymphoma (cHL). We characterized the peripheral B-cell compartment in a prospective cohort of 83 pediatric cHL patients. We observed significantly low total B-cell counts (<100 cells/µl) in 31 of 83 patients (37%). More specifically, there was a smaller peripheral IgDhighCD27– naïve B-cell pool among B-cell lymphopenic patients than for non-B-cell lymphopenic patients (p < 0.01). The B-cell count was lower in patients without in situ Epstein Barr Virus (EBV) expression than among those with in situ EBV expression (p = 0.03). Peripheral B-cell lymphopenia was associated with the presence of poor prognostic features, such as advanced lymphoma stage (p < 0.01) and the presence of B symptoms (p = 0.04). Of interest, B-cell lymphopenia resolved in all six studied patients in long-term remission. Our findings support that cHL tumor-associated factors interfere with the distribution of peripheral B-cell subsets.

Increased Plasma Cells and Decreased B-cells in Tumor Infiltrating Lymphocytes are Associated with Worse Survival in Lung Adenocarcinomas.

Clinical significance of tumor-infiltrating plasma cells and B-cells in lung adenocarcinoma is not well known. CD3, CD20 and MUM1 immunostains were performed on representative tumor blocks selected from 120 consecutive lung adenocarcinoma cases treated by surgical resection at Mayo Clinic Rochester. CD3+ T-cells, CD20+ B-cells, and MUM1+ plasma cells were enumerated separately in the intraepithelial (IE) compartment and the stroma (ST) by digital image analyses using whole sections. Measured tumor-infiltrating plasma cells and B-cells were correlated with patient's overall survival (OS) using Cox proportional hazards analysis. Median age of patients was 69 years (range, 46-91 years) and 52 were male. Median numbers (interquartile range) of CD20+ B-cells per 1mm2 of tumor area (IE plus ST) and IE compartment within tumor area were 590 (224-1276) and 101 (38-109), respectively; the corresponding numbers of MUM1+ plasma cells were 298 (180-605), and 67 (22-145), respectively. The proportion of MUM1+ plasma cell among all TILs (MUM1+ cells/[CD3+ cells +CD20+ cells+MUM1+ cells] × 100) ranged 1%-59% (median13%) in the tumor area and showed a significant association with OS by univariate Cox analysis (negative correlation with hazard ratio (HR)=12.50 [95% confidence interval (CI), 1.75-89.27]). There was a significant association between IE CD20+ B-cells and the patient's OS in univariate analysis (positive correlation with HR=0.81 [95% CI, 0.68-0.96]). Both parameters remained significant by multivariate analysis. High plasma cell % among TILs in the tumor area and low IE B-cell count were associated with worse prognosis in lung adenocarcinoma patients.

Serum soluble Fas ligand levels and peripheral blood lymphocyte subsets in patients with drug-induced maculopapular rashes, dress, and viral exanthemas

BackgroundFatty acid synthetase (Fas)/Fas ligand (FasL)-dependent apoptotic pathways have been reported as being involved in the pathogenesis of drug-induced maculopapular rashes. ObjectiveWe investigated serum soluble FasL (sFasL) levels and peripheral blood lymphocyte subtypes to discriminate maculopapular drug eruptions (MPDE) from viral exanthema (VE). Patients/methodsChildren with confirmed MPDE (group I), VE (group II), and drug rashes with eosinophilia and systemic symptoms (DRESS) or drug-induced hypersensitivity syndrome (DIHS) (group III) were included. Serum sFasL levels and peripheral blood lymphocyte subtypes were analyzed in groups I–III on admission, and repeated twice (only once for group IV – controls). ResultsThere were no significant serum soluble FasL level differences among the groups for all the samples. In the initial samples, CD19+ cell numbers in group II were significantly higher than in group IV, and the CD4+/CD8+ ratio was higher than groups I and IV. In the second samples, CD4+ and CD19+ cell numbers were significantly higher in group II than group I. In the final samples, CD4+ cell numbers in group II were significantly higher than group I and group III. CD19+ cells numbers in group III were significantly lower than the other groups for all samples. ConclusionSerum sFasL levels were not found to be useful in discriminating viral exanthemas from other drug rashes. The significant differences between MPDE, VE, and DRESS were high CD4+ and CD19+ cell-count numbers in VE but low B-cell numbers in DRESS. This might be important for discriminating VE from DRESS, and the low B-cell count in early symptoms might be a useful predictor of DRESS development.

Effect of low-dose rituximab treatment on T- and B-cell lymphocyte imbalance in refractory myasthenia gravis

We aimed to explore the effects of low-dose rituximab (RTX) on circulating T- and B-cell lymphocytes and the improvement of clinical symptoms in refractory myasthenia gravis (MG) patients. Fifteen patients with refractory MG were treated with a low dose of 600 mg RTX and were evaluated by serial-clinical scales, flow cytometry of peripheral blood T and B cells, and antibody titer before and after six months of RTX treatment. The quantitative MG score (QMGS), manual muscle testing (MMT), MG-related activities of daily living (MG-ADL) and MG-specific quality-of-life (QOL) were significantly improved and the average steroid-dosage reduction was 40% (p = .001) in refractory MG patients at six months after RTX infusion. Compared to eighteen non-refractory MG patients and eighteen Healthy controls, our study showed that the frequencies of circulating regulatory B cells (Bregs) and regulatory T cells (Tregs) were significantly lower and the expression of B-cell activating factor receptors (BAFF-Rs) was greater in refractory MG patients without RTX treatment. Importantly, 600-mg RTX was sufficient to deplete B cells and maintain low B-cell counts for up to six months after infusion. Additionally, a low dose of RTX further increased the frequencies of Tregs. Hence, there is an immune imbalance in circulating T- and B-cell lymphocytes in refractory MG patients compared to non-refractory MG patients. We conclude that remarkable T- and B-cell lymphocyte imbalance exists in refractory MG. Low-dose RTX can improve myasthenic symptom and deplete B cells and increase Tregs%.

A High Prevalence of Gastrointestinal Manifestations in Common Variable Immunodeficiency.

OBJECTIVES:Common variable immunodeficiency (CVID) is associated with a spectrum of autoimmune complications. We studied the prevalence of gastrointestinal (GI) manifestations and infections in patients with CVID.METHODS:Complete clinical data of 132 Finnish patients with CVID (106 probable and 26 possible CVID) followed up between 2007 and 2016 were collected to a structured database. Data on endoscopies, histology, and laboratory studies were retrieved from patient files.RESULTS:Most common referral indications were diarrhea and/or weight loss (47%–67%). Patients with probable CVID had higher fecal calprotectin and α1-antitrypsin and lower blood vitamin B12 than patients with possible CVID. Gastroscopy and colonoscopy were done to 71 (67%) and 63 (59%) patients with probable CVID, respectively. Endoscopies showed that 15% of them had chronic active gastritis and 17% atrophic gastritis and 3% had gastric adenocarcinoma. A celiac sprue-like condition was found in 7 patients (10%), of whom 3 responded to a gluten-free diet. Colonoscopies demonstrated unspecific colitis (14%), ulcerative colitis (8%), microscopic colitis (10%), and Crohn's disease (2%). Colonic polyps were noted in 30% of patients, and 3% had lower GI malignancies. Thirty-five patients with CVID had bacterial or parasitic gastroenteritis; chronic norovirus was detected in 4 patients with probable CVID. Patients with GI inflammation had higher levels of fecal calprotectin and blood CD8+ T lymphocytes but lower counts of CD19+CD27+ memory B cells and/or CD19+ B cells. Immunophenotype with low B-cell counts was associated with higher fecal calprotectin levels.DISCUSSION:Patients with CVID had a high prevalence of GI manifestations and infections of the GI tract. GI inflammation was associated with a distinct immunophenotype and elevated fecal calprotectin.

P2.04-06 Increased Plasma Cell % and Decreased B-Cells in Tumor Immune Infiltrates Are Associated with Worse Prognosis in Lung Adenocarcinomas

Clinical significance of tumor-infiltrating plasma cells and B-cells in lung adenocarcinoma is not well known. CD3, CD20 and MUM1 immunostains were performed on representative tumor blocks selected from 120 consecutive lung adenocarcinoma cases resected. CD3-positive T-cells, CD20-positive B-cells, and MUM1-positive plasma cells were separately enumerated in the intraepithelial (IE) compartment and the stroma (ST) by digital image analyses. Distribution of measured tumor-infiltrating cells was systematically evaluated and their associations with patient’s overall survival (OS) modeled using Cox proportional hazards analysis. Median age of patients was 69 years (range, 46-91 years) and 52 patients were male. Eighty-two, 17, and 21 patients were tumor stage I, II, and III/IV, respectively. Ninety patients had surgery only; 30 had surgery with adjuvant chemotherapy and/or radiation therapy. Median numbers (interquartile range) of CD20-positive B-cells per 1mm2 in the tumor area (IE plus ST) and in IE compartment were 590 (224-1276) and 101 (38-109), respectively; the corresponding numbers of MUM1-positive plasma cells were 298 (180-605), and 67 (22-145), respectively. The percent of MUM1-positive plasma cells among all tumor immune infiltrate (i.e. MUM1-positive cells/[CD3-positive cells + CD20-positive cells + MUM1- positive cells] x 100) ranged from 0 to 60% (median 10%) in the tumor area and showed a significant association with OS by univariate Cox analysis (continuous variable; negative correlation with HR=12.50 [95% confidence interval [CI], 1.75-89.27]). There was a significant association between IE CD20-positive B-cells and the patient’s OS in univariate analysis (continuous variable; positive correlation with hazard ratio [HR]=0.81 (95% CI, 0.68-0.96). Both parameters remained significant by multivariate analysis. Cut-off points (low vs high) showing significant associations with patient’s OS were found by log-rank test (Table 1). High plasma cell % among immune infiltrate in the tumor area and low IE B-cell count were associated with worse prognosis in lung adenocarcinoma patients.

Responsiveness to low-dose rituximab in refractory generalized myasthenia gravis

We aim to investigate the effect of a low dose of rituximab (RTX) in improving the clinical symptoms of refractory generalized myasthenia gravis (MG). Eight patients with refractory generalized MG were treated with a low dose of 600mg RTX. Patients were evaluated by serial clinical scales, flow cytometry of peripheral blood B, T and NK cells, immunoglobulin, complement levels and antibody titer. The quantitative MG score (QMGS), manual muscle testing (MMT), MG-related activities of daily living (MG-ADL) and MG-specific quality-of-life (QOL) were recorded at baseline as well as 1, 3, and 6months after RTX infusion. The initial improvement was recorded at 1month after treatment. QMGS, MMT and MG-ADL were significantly improved and the average steroid dosage reduction was 43% (p=0.018) at 6months. 600mg RTX was sufficient to deplete B cells and maintain low B-cell counts until 6months after infusion. Treatment with RTX did not result in a significant change in the percentage of CD4+, CD8+ T-cells while an average increase in the percentage of NK cells. Our study found successful B cell depletion was parallel to symptoms remission and change in serum C3 and C4 levels. Serum AChR antibody levels were independent of clinical response and not influenced by RTX. Therefore, low dose of 600mg RTX may be sufficient in depleting B cells, maintaining low B-cell counts and improving the clinical symptoms of MG in 6months.