Jacqueline Y. Channon

Differential infectivity and division of Toxoplasma gondii in human peripheral blood leukocytes.

ABSTRACT When tachyzoites were incubated with human peripheral blood leukocytes in vitro, more monocytes and dendritic cells than neutrophils or lymphocytes were infected. Although tachyzoites were able to divide in each of these cell types, monocytes and dendritic cells were more permissive to rapid tachyzoite division than neutrophils or lymphocytes.

A Randomized Trial of Ex vivo CD40L Activation of a Dendritic Cell Vaccine in Colorectal Cancer Patients: Tumor-Specific Immune Responses Are Associated with Improved Survival

Purpose: To determine whether an autologous dendritic cell (DC) vaccine could induce antitumor immune responses in patients after resection of colorectal cancer metastases and whether these responses could be enhanced by activating DCs with CD40L. Experimental Design: Twenty-six patients who had undergone resection of colorectal metastases were treated with intranodal injections of an autologous tumor lysate– and control protein [keyhole limpet hemocyanin (KLH)]–pulsed DC vaccine. Patients were randomized to receive DCs that had been either activated or not activated with CD40L. All patients were followed for a minimum of 5.5 years. Results: Immunization induced an autologous tumor-specific T-cell proliferative or IFNγ enzyme-linked immunospot response in 15 of 24 assessable patients (63%) and a tumor-specific DTH response in 61%. Patients with evidence of a vaccine-induced, tumor-specific T-cell proliferative or IFNγ response 1 week after vaccination had a markedly better recurrence-free survival (RFS) at 5 years (63% versus 18%, P = 0.037) than nonresponders. In contrast, no association was observed between induction of KLH-specific immune responses and RFS. CD40L maturation induced CD86 and CD83 expression on DCs but had no effect on immune responses or RFS. Conclusion: Adjuvant treatment of patients after resection of colorectal metastases with an autologous tumor lysate–pulsed, DC vaccine–induced, tumor-specific immune responses in a high proportion of patients. There was an association between induction of tumor-specific immune responses and RFS. Activation of this DC vaccine with CD40L did not lead to increased immune responses. Clin Cancer Res; 16(22); 5548–56. ©2010 AACR.

Toxoplasma gondii induces granulocyte colony-stimulating factor and granulocyte-macrophage colony-stimulating factor secretion by human fibroblasts: Implications for neutrophil apoptosis

ABSTRACT Human neutrophils are rescued from apoptosis following incubation with once-washed, fibroblast-derived Toxoplasma gondii tachyzoites. Both infected and uninfected neutrophils are rescued, implicating a soluble mediator. In this study we investigated the origin and identity of this soluble mediator. Neutrophils were incubated either with purified tachyzoites or with conditioned medium derived from T. gondii-infected human fibroblasts. Conditioned medium was found to be a potent stimulus that delayed neutrophil apoptosis up to 72 h, whereas purified and extensively washed tachyzoites had no effect. Delayed apoptosis correlated with up-regulation of the neutrophil antiapoptotic protein, Mcl-1, and the neutrophil interleukin 3 receptor α subunit (IL-3Rα), suggesting a role for granulocyte-macrophage colony-stimulating factor (GM-CSF). GM-CSF and granulocyte colony-stimulating factor (G-CSF) were measurable in conditioned medium by enzyme-linked immunosorbent assay. Neutralizing antibodies to GM-CSF and G-CSF were additive in abrogating delayed neutrophil apoptosis induced by conditioned medium. Inhibitors of Src family tyrosine kinases, Gi proteins, phosphatidylinositol 3-kinase, p44erk1 and p42erk2 mitogen-activated protein kinases, and Jak2 kinases partially attenuated the effect of conditioned medium, consistent with a role for G-CSF and/or GM-CSF. Hence, delayed neutrophil apoptosis is mediated by GM-CSF and G-CSF secreted by T. gondii-infected human fibroblasts. This enhanced neutrophil survival may contribute to the robust proinflammatory response elicited in the T. gondii-infected host.

The natural killer-activating receptor, NKG2D, on CD3+CD8+ T cells plays a critical role in identifying and killing autologous myeloma cells

The NKG2D receptor, one of the natural killer (NK) cell–activating receptors, is expressed on the surface of CD3+CD8+ T cells, γδ+ T cells, NK cells, NKT cells, and a few CD4+ T cells. We show, for the first time, a critical role for the NKG2D receptor on CD3+CD8+ T cells isolated from myeloma patients, in identifying and killing autologous myeloma cells isolated from the same patients’ marrow. We also show that blocking NKG2D using anti‐NKG2D reverses the cytotoxicity while blocking HLA‐I using antibodies does not have the same effect, showing that the autologous cytotoxicity is NKG2D dependent and major histocompatibility complex (MHC)‐I independent. We further confirmed the NKG2D specificity by small interfering RNA (siRNA) down regulation of NKG2D receptor.

Corr4A and VRT325 do not reduce the inflammatory response to P. aeruginosa in human cystic fibrosis airway epithelial cells.

Background: P. aeruginosa chronically colonizes the lung in CF patients and elicits a proinflammatory response. Excessive secretion of IL-6 and IL-8 by CF airway cells in response to P. aeruginosa infection in the CF airway is though to contribute to lung injury. Accordingly, the goal of this study was to test the hypothesis that Corr4a and VRT325, investigational compounds that increase ΔF508-CFTR mediated Cl- secretion in human CF airway cells, reduce the pro-inflammatory response to P. aeruginosa. Methods: IL-6 and IL-8 secretion by polarized CF human airway epithelial cells (CFBE41o-) were measured by multiplex analysis, and ΔF508-CFTR Cl- secretion was measured in Ussing chambers. Airway cells were exposed to P. aeruginosa (PAO1 or PA14) and Corr4a or VRT325. Results: Corr4a and VRT325 increased ΔF508-CFTR Cl- secretion but did not reduce either constitutive IL-6 or IL-8 secretion, or IL-6 and IL-8 secretion stimulated by P. aeruginosa (PA14 or PAO1). Conclusions: Corr4a and VRT325 do not reduce the inflammatory response to P. aeruginosa in human cystic fibrosis airway epithelial cells.

Alteration of CD39 + Foxp3 + CD4 T cell and cytokine levels in EAE/MS following anti-CD52 treatment

While examining the therapeutic value of anti-CD52 antibody against EAE/MS, we identified a unique subset of CD39+ Tregs in repopulating GALT tissues, a major lymphoid reservoir, which was accompanied by amelioration of disease. Furthermore, anti-CD52 treatment leads to increased expression of BDNF, IL-10, and SMAD3 in the brains of EAE mice. This condition is associated with suppression of IL-17, a critical inflammatory factor in EAE/MS progression. Additionally, we found elevated levels of CD4+CD39+ Tregs in PBMCs of RRMS patients treated with humanized anti-CD52 mAb. Thus, anti-CD52 can affect multiple immune mediated pathways involved in the pathogenesis of EAE/MS.

Dietary B Vitamin Intake Is Associated with Lower Urinary Monomethyl Arsenic and Oxidative Stress Marker 15-F2t-Isoprostane among New Hampshire Adults

Background: Arsenic exposure has been associated with an increased risk of cardiovascular disease (CVD). Growing evidence suggests that B vitamins facilitate arsenic metabolism and may protect against arsenic toxicity. However, to our knowledge, few studies have evaluated this in US populations.Objective: Our objective was to examine whether higher B vitamin intake is associated with enhanced arsenic metabolism and lower concentrations of preclinical markers of CVD among New Hampshire adults.Methods: We used weighted quantile sum (WQS) regression to evaluate the collective impact of 6 dietary B vitamins (thiamin, riboflavin, folate, niacin, and vitamins B-6 and B-12) on 1) the proportion of arsenic metabolites in urine and 2) 6 CVD-related markers [including urinary 15-F2t-isoprostane (15-F2t-IsoP)] among 418 participants (26-75 y of age) from the New Hampshire Health Study. Contributions of arsenic metabolites to B vitamin-CVD marker associations were also explored in structural equation models.Results: In WQS models, the weighted sum of B vitamin intakes from food sources was inversely associated with the proportion of monomethyl arsenic species in urine (uMMA) (β: -1.03; 95% CI: -1.91, -0.15; P = 0.02). Thiamin and vitamins B-6 and B-12 contributed the most to this association, whereas riboflavin had a negligible effect. Higher overall B vitamin intake was also inversely associated with 15-F2t-IsoP (β: -0.21; 95% CI: -0.32, -0.11; P < 0.01), with equal contributions from the 6 B vitamins, which was partially explained by differences in the proportion of uMMA (indirect effect β: -0.01; 95% CI: -0.04, -0.00).Conclusions: Among New Hampshire adults, higher intakes of certain B vitamins (particularly thiamin and vitamins B-6 and B-12 from food sources) may reduce the proportion of uMMA, an intermediate of arsenic metabolism that has been associated with an increased risk of CVD. Higher overall B vitamin intake may also reduce urinary 15-F2t-IsoP, a marker of oxidative stress and potential risk factor for CVD, in part by reducing the proportion of uMMA.

Abstract 2400: Treatment of colorectal cancer patients after resection of metastases with a tumor lysate pulsed dendritic cell vaccine: association between immune response and recurrence free survival

Introduction. Phase III trials have shown that a vaccine consisting of irradiated autologous tumor cells plus bacille Calmette-Guerin (BCG) can induce anti-tumor immune responses in patients after resection of colorectal cancer. We have demonstrated in preclinical studies that an intranodally injected tumor lysate pulsed dendritic cell (DC) vaccine induces a more potent immune response than tumor cells plus BCG. Preclinical studies also indicate that DC maturation via signaling through CD40 increases vaccine potency. Patients and Methods. Twenty six patients who had undergone resection of colorectal metastases were treated with an intranodal injection of a vaccine consisting of autologous tumor lysate pulsed dendritic cells. Keyhole limpet hemocyanin (KLH) was added to the lysate as an adjuvant and a control protein. Patients were randomized to receive DCs that had been either activated or not activated with rhCD40L. Immune responses were evaluated with a dye dilution proliferation assay, an IFNγ ELISPOT assay and delayed type hypersensitivity (DTH) testing. All patients were followed for a minimum of 5.5 years after vaccination. Results. The vaccine was administered to all patients with minimal toxicity. Immunization induced an autologous tumor specific proliferative T cell immune response in 8 of 24 assessable patients (33%), a tumor specific IFNγ secretory response in 10 of 24 patients (42%) and a DTH response to autologous tumor cells in 14 of 23 patients (61%). KLH specific responses were induced in 54%, 83% and 60% of patients by proliferation, IFNγ ELISPOT and DTH assays, respectively. Use of this whole cell antigen source induced peptide (CEA, Her-2 neu, Muc-1) specific T cell immune responses in half of the assayed patients. Relapse free survival (RFS) was 58%, 41% and 37% at 1, 2 and 5 years. Patients with evidence of a vaccine induced anti-tumor proliferative T cell immune response one week after vaccination had a markedly better RFS at 5 years (67% vs 31%, p=0.057) than non-responders. There was a trend towards better RFS at 5 years (67% vs 23%, p= 0.09) in patients who developed a vaccine induced tumor specific IFNγ T cell response. No association was observed between induction of KLH specific immune responses and RFS. DCs grown in the presence of rhCD40L were observed to have a significantly greater expression of the costimulatory molecule CD86 and the maturation marker CD83. However, rhCD40L DC activation did not significantly affect the percentage of patients with positive immune responses. Conclusions. Adjuvant treatment of patients after resection of colorectal metastases with a tumor lysate pulsed dendritic cell vaccine induced tumor specific immune responses in a high proportion of patients. There was an association between induction of tumor specific immune responses and recurrence free survival. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 2400.