Expression Of Co-signaling Molecules Research Articles

Blinatumomab in Combination with Immune Checkpoint Inhibitors (ICIs) of PD-1 and CTLA-4 in Adult Patients with Relapsed/Refractory (R/R) CD19 Positive B-Cell Acute Lymphoblastic Leukemia (ALL): Results of a Phase I Study

Background: Blinatumomab (blina) improves outcomes in R/R CD19+ ALL compared to chemotherapy. However, the overall response rate to blina was 44%, and median overall survival (OS) is just 7.7 mos (Kantarjian. NEJM. 2017). Preclinical studies show increased PD-L1 expression on leukemic blasts potentially contributes to relapse after blina. The addition of PD-1 +/- CTLA-4 blockade to blina leads to increased in vitro T cell proliferation and enhanced cytotoxicity (Feucht. Oncotarget 2016). Thus, blockade of co-inhibitory pathways may enhance blina efficacy in vivo. We describe results of a multi-center phase I study combining blina with immune checkpoint inhibitors (ICIs) targeting PD-1 (nivolumab) +/- CTLA-4 (ipilimumab). Methods: In a dose-escalation study, we evaluated the safety, tolerability, and preliminary efficacy of blina with nivolumab (nivo) +/- ipilimumab (ipi) using a 3+3 design. Pts ³16 years-old with R/R CD19+ B-ALL or mixed phenotype acute leukemia (MPAL) were eligible. Pts ³60 years could be untreated. Table 1 presents the dose escalation schema. Expansion cohorts of 6 pts were planned at the maximum tolerated dose (MTD) for blina + nivo AND blina + nivo + ipi. Pts received up to 5 cycles of blina and 1 year of ICIs. Pts removed from the study during the blina lead-in (days 1-10) were replaced. Dose-limiting toxicities (DLTs) were defined as grade 3+ non-hematologic toxicities requiring the permanent discontinuation of treatment in the first 42 days. Primary endpoints were toxicities and MTD. Secondary endpoints included complete remission (CR), MRD-negativity at a sensitivity of 0.01%, duration of response and OS from treatment initiation. Results: Twenty-seven pts were enrolled to 3 dose levels (14 DLA1, 3 DLB1, and 10 DLB-1) from September 2017-December 2022. Six pts were replaced (4 prior to ICI dosing and 2 requiring subsequent therapy in the DLT window). The median age of enrolled pts was 55 (range 24-84), 14 were male (52%), and median baseline BM blasts were 60% (range 0.2-98%). Baseline characteristics are presented in Table 2. A single DLT (G4 infusion reaction and G3 hypotension) occurred among 6 pts at DLA1. Based on this safety and efficacy data, blina + nivo was expanded to include 6 additional pts without additional DLTs. There were 2 DLTs at DLB1 (G5 pneumonitis and G2 GVHD), which led to de-escalation to DLB-1, where there was 1 DLT (G3 delirium). Grade 2+ immune-related adverse events attributable to ICIs included pneumonitis (G2+G5), rash (G2+G3), transaminitis (G2+G3), colitis (G3), and hypothyroidism (G2). Grade 3+ adverse events attributable to blina included neutropenia (5-G4+3-G3), dysphasia (2-G3), weakness (G3), seizure (G4), and transaminitis (1-G4+4-G3). Among 22 pts evaluable for response, the CR rate was 68% (100% MRD-negative). Among 13 pts with >50% BM blasts at baseline, 62% achieved CR. Pts with B-myeloid MPAL (0/2) and a history of extramedullary disease (1/4) were less likely to respond. Nine responders subsequently relapsed including 3 with isolated extramedullary disease and 2 with CD19-negative disease. As shown in Figure 1, relapse-free survival (RFS) at 1 year was 27% (95% CI 10-46), while OS at 1 year was 63% (95% CI 38-79). Twelve pts underwent allogeneic blood or marrow transplant (alloBMT) following study treatment. At 1 year for alloBMT pts, RFS was 51% (95% CI 19-76) and OS was 61% (95% CI 26-83). Ongoing analyses of changes in T cell subpopulations, co-signaling molecule expression, and single cell RNA seq results will be presented. Conclusions: Combination therapy with blina and ICIs in R/R ALL is safe, feasible, and associated with a high MRD-negative response rate. Long-term survival was promising in comparison to prior results with blina monotherapy, especially following consolidation with alloBMT. A randomized trial of blina +/- nivolumab is needed to confirm the benefit of this combination.

A Phase IB/II Study of Blinatumomab in Patients with B-Cell Acute Lymphoblastic Leukemia (ALL) and B-Cell Non-Hodgkin Lymphoma (NHL) As Post-Allogeneic Blood or Marrow Transplant (alloBMT) Remission Maintenance

Background: AlloBMT can be curative as consolidation for high risk B ALL and NHL. However, transplant-related toxicity and disease relapse limit survival. Post-transplantation cyclophosphamide (PTCy) as graft-versus-host disease (GVHD) prophylaxis limits GVHD and facilitates alternative allograft sources. Following PTCy, cellular immune reconstitution is favorable for strategies to augment anti-tumor immunity. Blinatumomab (blina) is effective in the treatment of CD19+ ALL and NHL. Blina leads to T cell activation that may enhance post-transplant tumor-specific T cell responses, leading to a more potent graft-versus-tumor effect. A study of 21 B ALL pts demonstrated the feasibility of post-transplant blina but did not improve survival when pts universally remained on immunosuppression for at least 1 cycle of treatment (Gaballa. Blood. 2022). We present results of a phase Ib/II trial to assess the tolerability and preliminary efficacy of blina as post-alloBMT remission maintenance in B-cell ALL and NHL in pts off immunosuppression. Methods: Pts ≥1 month-old with high risk CD19+ B ALL or ≥18 years-old with NHL who underwent alloBMT with PTCy were eligible. Pts had to be 60-180 days post-transplant with count recovery and in remission. Pts had to be off immunosuppression for ≥4 weeks prior to treatment, and without a history of grade ≥3 acute GVHD or severe chronic GVHD. Pts could receive 2 cycles of blina if they had evidence of disease (including MRD) at their pre- and/or post-transplant evaluations but otherwise received 1 cycle. Blina was given as a continuous infusion at 9 mcg/day on C1D1-7 and 28 mcg/day on C1D8-28 and C2D1-28. Results: The study closed to accrual on January 31, 2023 following enrollment of 42 pts (26 male/16 female) with a median age of 54 (Range 30-73). Among enrolled pts, 19 had Ph-negative B ALL and 23 had B-cell NHL. All pts underwent reduced-intensity conditioning alloBMT using a regimen of fludarabine, cyclophosphamide, and total body irradiation. Median prior lines of therapy were 2 (range 1-6). Additional transplant details are presented in Table 1. All B ALL pts were in an MRD-negative CR by flow cytometry at a sensitivity of 0.01% at alloBMT. All pts received a single cycle of therapy. Pts started blina a median of 137 days post-transplant (range, 90-182). Four pts failed to complete a full cycle due to G4 transaminitis (1), G4 neutropenia (1), relapse (1), and patient preference in the setting of tremors (1). Grade 3 or 4 adverse events felt to be at least possibly related to the study drug included G3 neutropenia (7%), G4 neutropenia (19%), anemia (5%), G3 ALT (5%), G4 AST (2%), G3 AST (2%), and neurotoxicity (7%). Two pts (5%) developed chronic GVHD following blina requiring the resumption of immunosuppression. At a median follow-up of 40 months post-alloBMT, the 3-year relapse-free survival is 73% (95% CI 54-85%) due to a 24% (95% CI 12-39%) incidence of relapse and a 4% (95% CI 0-17%) incidence of non-relapse mortality, as shown in Figure 2. All 3 (100%) relapsed B ALL pts and 2/6 (33%) relapsed B NHL pts presented with CNS involvement. Four pts with CNS relapse (80%) had CNS involvement prior to alloBMT. Among 2 pts who died without relapse, 1 was unrelated to alloBMT, while the other died of therapy-related myeloid neoplasm. Data on biomarkers including changes in T cell subpopulations in both BM and PB, and co-signaling molecule expression will be presented. Conclusions: Post-alloBMT maintenance therapy with blina is feasible with minimal toxicity in pts off immunosuppression. Initial survival outcomes are promising with the majority of relapses involving the CNS. A randomized trial of maintenance with blina after alloBMT with PTCy is needed to confirm efficacy in this high-risk population.

A Phase IB Study of Blinatumomab (blina) in Patients with B Cell Acute Lymphoblastic Leukemia (ALL) and B-Cell Non-Hodgkin Lymphoma (NHL) As Post-Allogeneic Blood or Marrow Transplant (allo-BMT) Remission Maintenance

Background: AlloBMT can be curative as consolidation for high risk B ALL and NHL. However, long term survival is limited by transplant-related toxicity and particularly by disease relapse. Post-transplantation cyclophosphamide (PTCy) as graft-versus-host disease (GVHD) prophylaxis limits GVHD and facilitates the use of alternative allograft sources. Moreover, following PTCy, cellular immune reconstitution is favorable for the integration of strategies to augment anti-tumor immunity. Blina, a CD19/CD3 bispecific T cell engager antibody construct, is effective in the treatment of CD19+ ALL and NHL. Blina leads to T cell activation that may enhance established posttransplant tumor-specific T cell responses, leading to a more potent graft-versus-tumor effect. Thus, we have undertaken a phase Ib trial to assess the tolerability and preliminary efficacy of blina as post-alloBMT remission maintenance in B-cell ALL and NHL. Methods: Patients ³18 years-old with high risk CD19+ B ALL or NHL who underwent alloBMT using PTCy were eligible including those with prior blina exposure. Patients had to be 60-180 days from transplant with documented count recovery and no evidence of disease progression. Patients had to be off all post-transplant immunosuppression including steroids for the treatment of GVHD for ≥4 weeks prior to treatment initiation, and without a history of grade ≥3 acute GVHD or severe chronic GVHD. Patients could receive two cycles of blina if they had evidence of disease (including MRD) at their pre- and/or post-transplant evaluations but otherwise received only one cycle. Blina was given as a continuous infusion at 9 mcg/day on C1D1-7 and 28 mcg/day on C1D8-28 and C2D1-28. Results: As of July 23, 2019, 12 adults (10 males/2 females) have enrolled including 4 patients with B ALL and 8 patients with NHL. Among the B ALL patients, two with known TP53 mutations were transplanted in CR1, while two with relapsed disease were transplanted in CR3. Among the NHL patients, five had large cell transformation (3 from follicular and 2 from CLL); one had relapsed primary CNS lymphoma (PCNSL); one had relapsed mantle cell lymphoma (MCL); and one had diffuse large B cell lymphoma (DLBCL). The median age was 53 (range, 30-73). All patients underwent alloBMT using a conditioning regimen of fludarabine, cyclophosphamide, and total body irradiation (TBI). Eight patients received allografts from haploidentical donors, three from matched-unrelated donors, and one from a matched-related donor. Five patients received peripheral blood allografts, and seven bone marrow. Two patients were enrolled after a second alloBMT, and 3/4 ALL patients previously received blina. Baseline characteristics are presented in Figure 1. Patients started blina a median of 144 days post-transplant (range, 90-180). One patient stopped treatment on day 5 due to a grade 2 tremor, and one patient required dose reduction on day 25 due to grade 4 neutropenia. Toxicities were otherwise mild and are presented in Figure 2. There were no exacerbations of GVHD. At a median follow-up of 13.7 months after BMT (range 3.8-23 months), ten patients remain in remission, while one patient suffered a third CNS relapse of ALL at 20.6 months after his 2nd transplant and another had relapse of his transformed lymphoma. Data on biomarkers including changes in T cell subpopulations in both BM and PB, and co-signaling molecule expression will be presented. Conclusions: Post-alloBMT maintenance therapy with blina is feasible with minimal toxicity. 83% of the very high risk patients treated on study remain in CR at a median of 13.7 months post-transplant. Based on promising safety and efficacy data from the phase IB, the plan is to proceed to the phase II portion of the study. Disclosures Webster: Amgen: Consultancy; Genentech: Research Funding; Pfizer: Consultancy. Wagner-Johnston:Jannsen: Membership on an entity's Board of Directors or advisory committees; Bayer: Membership on an entity's Board of Directors or advisory committees; Gilead: Membership on an entity's Board of Directors or advisory committees; ADC Therapeutics: Membership on an entity's Board of Directors or advisory committees. Luznik:Merck: Research Funding, Speakers Bureau; Genentech: Research Funding; AbbVie: Consultancy; WindMiL Therapeutics: Patents & Royalties: Patent holder. Gojo:Abbvie: Consultancy, Honoraria; Novartis: Consultancy, Honoraria; Merck: Research Funding; Jazz: Consultancy, Honoraria; Amgen Inc: Consultancy, Honoraria, Research Funding; Juno: Research Funding; Amphivena: Research Funding. OffLabel Disclosure: Blinatumomab is not labeled for use as post-transplant maintenance therapy or for use in the non-Hodgkin lymphoma.

Blinatumomab in Combination with Immune Checkpoint Inhibitors of PD-1 and CTLA-4 in Adult Patients with Relapsed/Refractory (R/R) CD19 Positive B-Cell Acute Lymphoblastic Leukemia (ALL): Preliminary Results of a Phase I Study

Background: Blinatumomab, a CD19/CD3 bispecific T cell engager antibody construct, leads to improved outcomes in patients with R/R CD19+ ALL compared to standard chemotherapy. However, most adults fail to achieve complete remission (CR) with blinatumomab, and the median duration of remission is only 7.3 months. Preclinical studies have shown significantly increased PD-L1 expression on leukemic blasts in patients who are refractory to or relapse after response to blinatumomab. Additionally, expression of the exhaustion markers PD-1 and TIM-3 on bone marrow (BM) CD3+ T cells is significantly higher among ALL patients than controls. The addition of PD-1 blockade +/- CTLA-4 blockade to blinatumomab and ALL blasts in vitro leads to increased T cell proliferation and enhanced blinatumomab-mediated cytotoxicity (Feucht et al, Oncotarget 2016). Thus, blockade of co-inhibitory pathways represents a viable strategy to enhance blinatumomab efficacy. We describe early results of a multi-center phase I study combining blinatumomab with monoclonal antibodies targeting PD-1 (nivolumab) +/- CTLA-4 (ipilimumab) in R/R CD19+ ALL.Methods: This phase I dose-escalation study evaluates the safety and tolerability (MTD) of blinatumomab in combination with nivolumab +/- ipilimumab using a 3+3 design. Patients ≥16 years-old with R/R CD19+ Pre-B ALL or MPAL are eligible including those with prior blinatumomab and/or prior allogeneic transplant (allo-SCT). Patients ≥60 years may be untreated and those 16-21 must be R/R to ³2 lines of therapy. The trial started at dose level (DL) A1 (Fig. 1). Upon determining the MTD for the combination of blinatumomab and nivolumab, dose escalation will add ipilimumab (DLB1). Patients may receive up to 5 cycles of blinatumomab and 1 year of nivolumab/ipilimumab. Patients achieving CR may proceed to allo-SCT. Patients removed from the study during the blinatumomab lead-in (days 1-10) will be replaced.Results: As of July 31, 2018, 8 adults (4 males/4 females) had enrolled at DLA1. The median age was 55 (range, 25-75) and baseline BM blast percentage was 73% (range, 8-98%). Baseline characteristics are presented in Figure 2. Seven patients received cytoreduction before treatment (6 steroids only and 1 steroids + Cytoxan). Two patients previously treated with blinatumomab were withdrawn from the study during the blinatumomab lead-in (1 for G3 pericardial effusion 2/2 disease progression and 1 for G3 hyperbilirubinemia). Among the 5 patients who received nivolumab to date, drug-related non-hematologic AEs of grade ≥3 included elevated AST (20%), ALT (20%), amylase (20%), and lipase (G4, 20%); hypophosphatemia (20%); rash (20%); infusion-related reaction (G4, 20%); and hypotension (20%). The elevated AST, ALT, amylase and lipase occurred prior to nivolumab dosing and resolved. One patient was removed from the study for a G4 infusion-related reaction following the 2nd dose of nivolumab that was considered a DLT. One patient in CR developed G4 neutropenia in cycles 2 + 3 but recovered spontaneously. Among the 5 evaluable patients, 4 (80%) achieved CR without MRD (2 after 1 cycle and 2 after 2 cycles of blinatumomab) with 3 ongoing remissions (median f/u 5 months) and 1 extramedullary relapse at day 125. Data on biomarkers including changes in T cell subpopulations in both BM and PB, and co-signaling molecule expression will be presented.Conclusions: Combination therapy with blinatumomab and nivolumab in R/R ALL with is feasible with acceptable toxicity. The MRD-negative CR rate was (80%) despite heavily pre-treated patients with significant baseline disease burden. The last patient treated at DLA1 is undergoing treatment before dose escalation to include ipilimumab. DisclosuresDeAngelo:Shire: Honoraria; ARIAD: Consultancy, Research Funding; Takeda: Honoraria; BMS: Consultancy; Amgen: Consultancy; Blueprint Medicines: Honoraria, Research Funding; Pfizer Inc: Consultancy, Honoraria; Glycomimetics: Research Funding; Novartis Pharmaceuticals Corporation: Consultancy, Honoraria; Incyte: Consultancy, Honoraria. Luznik:WIndMIL Therapeutics: Equity Ownership, Patents & Royalties. Gojo:Amgen: Membership on an entity's Board of Directors or advisory committees; Novartis: Membership on an entity's Board of Directors or advisory committees; Amgen: Research Funding; Merck inc: Research Funding; Jazz Pharmaceuticals: Membership on an entity's Board of Directors or advisory committees.

Case-control study of PD-1, PD-L1, and B7-H3 expression in human immunodeficiency virus infected (HIV+) and uninfected patients with head and neck cancers.

e23194 Background: HIV+patients are typically excluded from immunotherapy trials but there are limited data on co-signaling molecule expression in these patients, including in head and neck squamous cell carcinoma (HNSCC). Methods: A case control study was conducted to evaluate tumor tissue sections for PD-L1, PD-1, and B7-H3 expression in HIV+HNSCC patients (n = 12) and HNSCC controls (n = 12). Cases and controls were matched for age at diagnosis, race, gender, TNM stage, and primary site. Smoking status, HIV RNA viral load (VL), and CD4 count were also recorded. Immunostained tumor sections were analyzed for percent of tumor cells expressing PD-L1 (PD-L1%) and B7-H3(B7-H3%) (Abcam), and percent of tumor infiltrating lymphocytes (TIL) expressing PD-1 (TIL-PD-1%) and PD-L1 (TIL-PD-L1%) (Abcam). Statistical analysis used the non-parametric Mann-Whitney test, chi-square test and Spearman’s rank correlation, Rs. Results: The 12 HIV+ HNSCC cases were predominantly male (67%), black race (67%) and cigarette smokers (100%), with a median age of 50.4 years, median viral load (VL) of 52399 copies/mL, median CD4 count (CD4) of 236cells/uL, predominantly locally advanced (75% Stage III/IVa/IVb), and oropharyngeal primary site (42%). Defining positive expression as > 5%, 42% of HIV+ HNSCC patients tumors were positive for PD-L1, 100% for B7-H3, and 92% had TILs that expressed PD-1 and PD-L1. HIV+ patients had significantly higher B7-H3% (Median 60% vs. 20%, p = 0.03) and TIL-PD-L1% (Median 15% vs. 10%, p = 0.045) compared to controls. There were no differences in PD-L1% or TIL-PD-1%. In HIV+ cases, increased VL correlated with increased TIL-PD-1% (Rs = 0.73, p = 0.011) and increased CD4 count correlated with increased tumor PD-L1% (Rs = 0.62, p = 0.04). There were no other significant correlations between CD4 count or VL and co-signaling molecule expression. Conclusions: HIV+ HNSCC patients had significantly higher tumor B7-H3 and TIL PD-L1 expression, with similar tumor PD-L1 and TIL PD-1 expression, compared to HIV negative HNSCC control patients. These findings support inclusion of HIV+ HNSCC patients in immunotherapy trials with checkpoint inhibitors.

Abstract 5001: High dose IL-2 (HDIL-2) results in increased PD-1 expression on CD45RA-CCR7-CD8+ and CD45RA-CCR7-CD4+ T cells in melanoma patients

Abstract Introduction: HDIL-2 is associated with long-term remissions in Stage IV melanoma in less than 10% of patients. We hypothesized that HDIL-2 alters the expression of PD-1 on T cell subsets in melanoma patients. Methods: Samples were collected after written consent under an IRB approved protocol. Peripheral blood was collected at baseline and serially post-treatment at 24, 48, 72, and 96 hours from 6 patients with advanced stage melanoma undergoing HDIL-2. All patients were treatment naïve with regards to anti-PD-1 and anti-CTLA4 therapies. PBMCs were isolated and underwent intracellular cytokine and extracellular receptor staining. Co-signaling molecule expression was analyzed via flow cytometry. Statistical analysis was performed using paired t tests via Prism 6.0e software. Results: HDIL-2 was associated with a preferential loss of effector memory (TEM; CD45RA-CCR7-) CD4+ and CD8+ T cells and a relative enrichment of naïve (CD45RA+CCR7+) CD4+ and CD8+ T cells in the blood during the first 3 days of treatment. On day 4, frequencies of CD4+ and CD8+ TEM had returned to baseline. Importantly, PD-1 expression was significantly increased on day 4 post-treatment on CD8+ TEM relative to baseline (22.1% +/- 13.2% on day 4 compared to 13.4% +/- 7.4% at baseline; p = 0.04). This finding was also true for CD4+ TEM (11.4% +/- 6.1% on day 4 compared to 6.1% +/- 3.7% at baseline, p = 0.04). PD-1 expression was not statistically different within the other CD4+ and CD8+ memory T cell subsets including: naïve, central memory (CD45RA-CCR7+), or TEMRA (CD45RA+CCR7-) lymphocytes. Conclusion: These results demonstrate that HDIL-2 is associated with an increase in expression of PD-1 on TEM cells in patients with advanced melanoma suggesting a possible mechanism contributing to the limited efficacy of HDIL-2 in most patients. It is known that TEM cells function by migrating to peripheral tissues, and that CD8+ TEM cells are found specifically in metastatic melanoma lesions. Importantly, our findings illustrate a potential synergistic effect between HDIL-2 and PD-1 blockade. Upregulation of PD-1 and the chronologic pattern in which its expression changes under the effects of HDIL-2 provide a mechanistic rationale for testing the combination of HDIL-2 and anti-PD-1 therapy in patients with advanced melanoma. Citation Format: Maggie L. Diller, Susan Maio, Cabell E. Eysmans, David Lawson, Keith A. Delman, Ragini R. Kudchadkar, Mandy L. Ford. High dose IL-2 (HDIL-2) results in increased PD-1 expression on CD45RA-CCR7-CD8+ and CD45RA-CCR7-CD4+ T cells in melanoma patients. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 5001. doi:10.1158/1538-7445.AM2015-5001

Analysis of CTLA4 expression on T regulatory cells in patients with advanced stage melanoma in the setting of high dose IL-2 (HDIL-2).

9043 Background: HDIL-2 is associated with long-term remissions in stage IV melanoma, but in less than 10% of patients. Combination therapy with HDIL-2 and agents immunomodulating CTLA-4 have been undertaken. We investigated the effect of HDIL-2 on CTLA4 expression on T cell subsets in patients with melanoma. Methods: Peripheral blood was collected at baseline and serially post-treatment at 24, 48, 72, and 96 hours from 6 patients with advanced stage melanoma undergoing HDIL-2. All patients were treatment naive with regard to anti-CTLA4 therapy. PBMCs were isolated and restimulated ex vivo for 4 hours using PMA/ionomycin. Fluorophore conjugated anti-CTLA4 antibody was added at time of restimulation and incubated with cells for 4 hours. Intracellular and extracellular cytokine staining was then performed and co-signaling molecule expression was quantified via flow cytometry and measured on a continuous scale. Statistical analysis was performed using paired t tests via Prism 6.0e software. Results: HDIL-2 r...

Infusion of Stably Immature Monocyte-Derived Dendritic Cells Plus CTLA4Ig Modulates Alloimmune Reactivity in Rhesus Macaques

Immature dendritic cells (DC) can promote long-term transplant survival in rodents. We assessed the impact of stably immature, donor-derived DC on alloimmune reactivity in rhesus macaques. CD14 monocytes isolated from leukapheresis products of Macacca mulatta were cultured in granulocyte-macrophage colony stimulating factor plus interleukin (IL)-4+/-vitamin (vit) D3, and IL-10. Major histocompatibility complex class II and cosignaling molecule expression was determined on CD11c cells by flow cytometry. T-cell allostimulatory capacity of the DC, including DC exposed to proinflammatory cytokines, was determined in mixed leukocyte reaction. To test their influence in vivo, purified DC were infused intravenously into allogeneic recipients, either alone or followed by CTLA4Ig, 24 hr later. Proliferative responses of recipient CFSE-labeled T cells to donor or third party DC, cytokine production by stimulated T cells, and circulating alloantibody levels were determined by flow cytometry, up to 100 days postinfusion. VitD3/IL-10-conditioned, monocyte-derived DC were resistant to maturation and failed to induce allogeneic T cell proliferation in vitro. After their infusion, an increase in anti-donor and anti-third party T-cell reactivity was observed, that subsequently subsided to fall significantly below pretreatment levels (by day 56) only in animals also given CTLA4Ig. No increase in circulating immunoglobulin (Ig) M or IgG anti-donor alloantibody titers compared with pretreatment values was detected. With DC+CTLA4Ig infusion, alloreactive IL-10-producing T cells were prevalent in the circulation after day 28. Maturation-resistant rhesus DC infusion is well-tolerated. DC+CTLA4Ig infusion modulates allogeneic T-cell responses and results in hyporesponsiveness to donor and third party alloantigens.

Ethanol affects the generation, cosignaling molecule expression, and function of plasmacytoid and myeloid dendritic cell subsets in vitro and in vivo

The influence of ethanol (EtOH) on multiple dendritic cell (DC) subsets, in the steady state or following their mobilization in vivo, has not been characterized. Herein, generation of mouse bone marrow-derived DC (BMDC) in response to fms-like tyrosine kinase 3 ligand was inhibited by physiologically relevant concentrations of EtOH with selective suppression of plasmacytoid (p)DC. EtOH reduced surface expression of costimulatory molecules (CD40, CD80, CD86) but not that of coinhibitory CD274 (B7-H1) on resting or CpG-stimulated DC subsets. Interleukin (IL)-12p70 production by activated DC was impaired. Consistent with these findings, EtOH-exposed BMDC exhibited a reduced capacity to induce naïve, allogeneic T cell proliferation and impaired ability to prime T cells in vivo. DC subsets freshly isolated from EtOH-fed mice were also examined. Liver DC, inherently immature and resistant to maturation, exhibited little change in their low surface cosignaling molecule expression, whereas splenic DC showed reduced expression of surface costimulatory molecules in response to CpG stimulation in vivo. These splenic DC elicited reduced naïve, allogeneic T cell proliferation in vitro, and the stimulatory capacity of resting but not CpG-activated liver DC was reduced by chronic EtOH administration. T cells from animals primed with EtOH-exposed DC produced elevated levels of IL-10 following ex vivo challenge with donor alloantigen. Thus, EtOH impairs cytokine-driven differentiation and function of myeloid DC and pDC in vitro. Hepatic DC from chronic EtOH-fed mice are less affected than splenic DC, which exhibit impaired functional maturation following CpG stimulation. These results indicate a potential mechanism by which alcohol consumption is associated with immunosuppression.

Keratinocytes in atopic dermatitis: inflammatory signals.

Atopic dermatitis (AD) is a chronic inflammatory skin disorder that usually predates the development of allergic airway disease. In most cases, this is thought to be an allergen-driven disease with prominent roles played by antigen presenting cells and effector Th2 cells. But keratinocytes, by virtue of their location, provide an important window to the environment and are also thought to contribute to the development of AD. In this review, we discuss several biologic attributes of keratinocytes that are relevant for AD: 1) intrinsic defects in barrier function, 2) production of inflammatory mediators that promote or maintain allergic inflammation, 3) keratinocyte apoptosis, 4) effects of staphylococcal toxins on keratinocytes, and 5) potential consequences of the expression of cosignaling molecules (eg, B7 family members) and receptors important for innate immune responses (eg, Toll receptors). Clearly, these findings have highlighted a more active role played by the epithelium than was previously recognized.