Adjuvant vaccination with melanoma antigen-pulsed dendritic cells in stage III melanoma patients.

Med Oncol. 2012 Feb 3;

Authors: Markowicz S, Nowecki ZI, Rutkowski P, Lipkowski AW, Biernacka M, Jakubowska-Mucka A, Switaj T, Misicka A, Skurzak H, Polowniak-Pracka H, Walewski J

Abstract
Dendritic cells may be successfully used to induce in vivo-specific anti-tumor responses when combined with the appropriate antigen in the appropriate context. The purpose of this study was to evaluate efficacy of peptide-loaded DC vaccine in high-risk stage III melanoma patients after lymph node dissection (LND). HLA-A2(+), -A1(+), or -A3(+) melanoma patients (N = 22), stage III, N1b-N3, received 5-16 (median: 11) DC vaccines loaded with MHC class-I-restricted melanoma peptides respective to the patient's haplotype, and with autologous tumor lysate, if available. Vaccinated patients were matched to unvaccinated stage III controls (22 of 869) by sex, number of metastatic lymph nodes, extracapsular involvement, LND type, Breslow stage, and ulceration. Vaccination elicited cutaneous delayed-type hypersensitivity (DTH) or/and IFN-γ-producing CD8(+) cell response to melanoma peptides in 15 of 22 patients. Three-year overall survival (OS) rate was 68.2% in the vaccinated group versus 25.7% in the control group, P value accounting for matching: 0.0290. In a Cox regression model, hazard ratio (HR) for death of vaccinated patients was 0.31 [95% confidence interval (CI): 0.10-0.94]. The corresponding values for 3-year disease-free survival rate were 40.9 versus 14.5%, P = 0.1083; HR of recurrence for vaccinated, 0.46 (95% CI: 0.18-1.22). There was no grade >1 toxicity. The DC/peptide vaccine was well tolerated and elicited immune responses to melanoma antigens. Vaccinated patients had significantly longer OS after LND than the matched controls, but a significant improvement in the primary endpoint DFS was not achieved.

PMID: 22302285 [PubMed - as supplied by publisher]

Review of Chinese clinical trials on CIK cell treatment for malignancies.

Clin Transl Oncol. 2012 Feb;14(2):102-8

Authors: Li XD, Xu B, Wu J, Ji M, Xu BH, Jiang JT, Wu CP

Abstract
China is the country where the most clinical trials on CIK cells have been performed. We aimed to provide definite evidence for using CIK cell treatment and extrapolate a common applicative standard for malignancies. We chose the VIP database of Chinese scientific and technological journals to search the literature. We entered the keywords "CIK" or "xi bao yin zi you dao de sha shang xi bao" (the equivalent Chinese phrase for CIK cells, by Chinese characters) and searched for in vivo human trials. In 24 collected trials, 936 patients were treated with CIK cells, 525 men and 246 women. The cultivation time of CIK cells ranged from 7 to 28 days. In five studies, CIK cells were co-cultured with dendritic cells. The total number of CIK cells used ranged from 6×10(6) to 1.5×10(10). The total number of DC-CIK cells used ranged from 1×10(9) to 1.3×10(10). In all studies, those immune parameters and tumour markers examined increased, but not all increased significantly. Of the reported 563 patients, 40 had a complete response, 126 had a partial response, 125 had a minimal response, 135 had stable disease and 58 had progressive disease. The remaining 76 patients did not reach an objective response. The total response rate was 51.7% (291/563). The toxicities were slight. CIK cell treatment is a promising and safe modality for treating malignancies. We proposed a standard for cultivating CIK cells.

PMID: 22301398 [PubMed - in process]

Transgene expression and local tissue distribution of naked and polymer-condensed plasmid DNA after intradermal administration in mice.

J Control Release. 2012 Jan 24;

Authors: Palumbo RN, Zhong X, Panus D, Han W, Ji W, Wang C

Abstract
DNA vaccination using cationic polymers as carriers has the potential to be a very powerful method of immunotherapy, but typical immune responses generated have been less than robust. To better understand the details of DNA vaccine delivery in vivo, we prepared polymer/DNA complexes using three structurally distinct cationic polymers and fluorescently labeled plasmid DNA and injected them intradermally into mice. We analyzed transgene expression (luciferase) and the local tissue distribution of the labeled plasmid at the injection site at various time points (from hours to days). Comparable numbers of luciferase expressing cells were observed in the skin of mice receiving naked plasmid or polyplexes one day after transfection. At day 4, however, the polyplexes appeared to result in more transfected skin cells than naked plasmid. Live animal imaging revealed that naked plasmid dispersed quickly in the skin of mice after injection and had a wider distribution than any of the three types of polyplexes. However, naked plasmid level dropped to below detection limit after 24h, whereas polyplexes persisted for up to 2weeks. The PEGylated polyplexes had a significantly wider distribution in the tissue than the nonPEGylated polyplexes. PEGylated polyplexes also distributed more broadly among dermal fibroblasts and allowed greater interaction with antigen-presenting cells (APCs) (dendritic cells and macrophages) starting at around 24h post-injection. By day 4, co-localization of polyplexes with APCs was observed at the injection site regardless of polymer structure, whereas small amounts of polyplexes were found in the draining lymph nodes. These in vivo findings demonstrate the superior stability of PEGylated polyplexes in physiological milieu and provide important insight on how cationic polymers could be optimized for DNA vaccine delivery.

PMID: 22300619 [PubMed - as supplied by publisher]

Decrease in calcium concentration triggers neuronal retinoic acid synthesis during homeostatic synaptic plasticity.

J Neurosci. 2011 Dec 7;31(49):17764-71

Authors: Wang HL, Zhang Z, Hintze M, Chen L

Abstract
Blockade of synaptic activity induces homeostatic plasticity, in part by stimulating synthesis of all-trans retinoic acid (RA), which in turn increases AMPA receptor synthesis. However, the synaptic signal that triggers RA synthesis remained unknown. Using multiple activity-blockade protocols that induce homeostatic synaptic plasticity, here we show that RA synthesis is activated whenever postsynaptic Ca(2+) entry is significantly decreased and that RA is required for upregulation of synaptic strength under these homeostatic plasticity conditions, suggesting that Ca(2+) plays an inhibitory role in RA synthesis. Consistent with this notion, we demonstrate that both transient Ca(2+) depletion by membrane-permeable Ca(2+) chelators and chronic blockage of L-type Ca(2+)-channels induces RA synthesis. Moreover, the source of dendritic Ca(2+) entry that regulates RA synthesis is not specific because mild depolarization with KCl is sufficient to reverse synaptic scaling induced by L-type Ca(2+)-channel blocker. By expression of a dihydropyridine-insensitive L-type Ca(2+) channel, we further show that RA acts cell autonomously to modulate synaptic transmission. Our findings suggest that, in synaptically active neurons, modest "basal" levels of postsynaptic Ca(2+) physiologically suppress RA synthesis, whereas in synaptically inactive neurons, decreases in the resting Ca(2+) levels induce homeostatic plasticity by stimulating synthesis of RA that then acts in a cell-autonomous manner to increase AMPA receptor function.

PMID: 22159093 [PubMed - indexed for MEDLINE]

Shared and distinct functions of the transcription factors IRF4 and IRF8 in myeloid cell development.

PLoS One. 2011;6(10):e25812

Authors: Yamamoto M, Kato T, Hotta C, Nishiyama A, Kurotaki D, Yoshinari M, Takami M, Ichino M, Nakazawa M, Matsuyama T, Kamijo R, Kitagawa S, Ozato K, Tamura T

Abstract
Interferon regulatory factor (IRF) 8 and IRF4 are structurally-related, hematopoietic cell-specific transcription factors that cooperatively regulate the differentiation of dendritic cells and B cells. Whilst in myeloid cells IRF8 is known to modulate growth and differentiation, the role of IRF4 is poorly understood. In this study, we show that IRF4 has activities similar to IRF8 in regulating myeloid cell development. The ectopic expression of IRF4 in myeloid progenitor cells in vitro inhibits cell growth, promotes macrophages, but hinders granulocytic cell differentiation. We also show that IRF4 binds to and activates transcription through the IRF-Ets composite sequence (IECS). Furthermore, we demonstrate that Irf8⁻/⁻Irf4⁻/⁻ mice exhibit a more severe chronic myeloid leukemia (CML)-like disease than Irf8⁻/⁻ mice, involving a disproportionate expansion of granulocytes at the expense of monocytes/macrophages. Irf4⁻/⁻ mice, however, display no obvious abnormality in myeloid cell development, presumably because IRF4 is expressed at a much lower level than IRF8 in granulocyte-macrophage progenitors. Our results also suggest that IRF8 and IRF4 have not only common but also specific activities in myeloid cells. Since the expression of both the IRF8 and IRF4 genes is downregulated in CML patients, these results may add to our understanding of CML pathogenesis.

PMID: 22003407 [PubMed - indexed for MEDLINE]

The soluble recombinant Neisseria meningitidis adhesin NadA(Δ351-405) stimulates human monocytes by binding to extracellular Hsp90.

PLoS One. 2011;6(9):e25089

Authors: Cecchini P, Tavano R, Polverino de Laureto P, Franzoso S, Mazzon C, Montanari P, Papini E

Abstract
The adhesin NadA favors cell adhesion/invasion by hypervirulent Neisseria meningitidis B (MenB). Its recombinant form NadA(Δ351-405,) devoid of the outer membrane domain, is an immunogenic candidate for an anti-MenB vaccine able to stimulate monocytes, macrophages and dendritic cells. In this study we investigated the molecular mechanism of NadA(Δ351-405) cellular effects in monocytes. We show that NadA(Δ351-405) (against which we obtained polyclonal antibodies in rabbits), binds to hsp90, but not to other extracellular homologous heat shock proteins grp94 and hsp70, in vitro and on the surface of monocytes, in a temperature dependent way. Pre-incubation of monocytes with the MenB soluble adhesin interfered with the binding of anti-hsp90 and anti-hsp70 antibodies to hsp90 and hsp70 at 37°C, a condition in which specific cell-binding occurs, but not at 0°C, a condition in which specific cell-binding is very diminished. Conversely, pre-incubation of monocytes with anti-hsp90 and anti-hsp70 antibodies did not affected NadA(Δ351-405) cell binding in any temperature condition, indicating that it associates to another receptor on their plasma membrane and then laterally diffuses to encounter hsp90. Consistently, polymixin B interfered with NadA(Δ351-405) /hsp90 association, abrogated the decrease of anti-hsp90 antibodies binding to the cell surface due to NadA(Δ351-405) and inhibited adhesin-induced cytokine/chemokine secretion without affecting monocyte-adhesin binding. Co-stimulation of monocytes with anti-hsp90 antibodies and NadA(Δ351-405) determined a stronger but polymixin B insensitive cell activation. This indicated that the formation of a recombinant NadA/hsp90/hsp70 complex, although essential for full monocyte stimulation, can be replaced by anti-hsp90 antibody/hsp90 binding. Finally, the activation of monocytes by NadA(Δ351-405) alone or in the presence of anti-hsp90 antibodies were both inhibited by neutralizing anti-TLR4 antibodies, but not by anti-TLR2 antibodies. We propose that hsp90-dependent recruitment into an hsp90/hsp70/TLR4 transducing signal complex is necessary for the immune-stimulating activity of NadA(Δ351-405) anti-MenB vaccine candidate.

PMID: 21949862 [PubMed - indexed for MEDLINE]

Different toll-like receptor stimuli have a profound impact on cytokines required to break tolerance and induce autoimmunity.

PLoS One. 2011;6(9):e23940

Authors: Lin AC, Dissanayake D, Dhanji S, Elford AR, Ohashi PS

Abstract
Although toll-like receptor (TLR) signals are critical for promoting antigen presenting cell maturation, it remains unclear how stimulation via different TLRs influence dendritic cell (DC) function and the subsequent adaptive response in vivo. Furthermore, the relationship between TLR-induced cytokine production by DCs and the consequences on the induction of a functional immune response is not clear. We have established a murine model to examine whether TLR3 or TLR4 mediated DC maturation has an impact on the cytokines required to break tolerance and induce T-cell-mediated autoimmunity. Our study demonstrates that IL-12 is not absolutely required for the induction of a CD8 T-cell-mediated tissue specific immune response, but rather the requirement for IL-12 is determined by the stimuli used to mature the DCs. Furthermore, we found that IFNα is a critical pathogenic component of the cytokine milieu that circumvents the requirement for IL-12 in the induction of autoimmunity. These studies illustrate how different TLR stimuli have an impact on DC function and the induction of immunity.

PMID: 21931625 [PubMed - indexed for MEDLINE]

Mycobacterium tuberculosis lineage influences innate immune response and virulence and is associated with distinct cell envelope lipid profiles.

PLoS One. 2011;6(9):e23870

Authors: Krishnan N, Malaga W, Constant P, Caws M, Tran TH, Salmons J, Nguyen TN, Nguyen DB, Daffé M, Young DB, Robertson BD, Guilhot C, Thwaites GE

Abstract
The six major genetic lineages of Mycobacterium tuberculosis are strongly associated with specific geographical regions, but their relevance to bacterial virulence and the clinical consequences of infection are unclear. Previously, we found that in Vietnam, East Asian/Beijing and Indo-Oceanic strains were significantly more likely to cause disseminated tuberculosis with meningitis than those from the Euro-American lineage. To investigate this observation we characterised 7 East Asian/Beijing, 5 Indo-Oceanic and 6 Euro-American Vietnamese strains in bone-marrow-derived macrophages, dendritic cells and mice. East Asian/Beijing and Indo-Oceanic strains induced significantly more TNF-α and IL-1β from macrophages than the Euro-American strains, and East Asian/Beijing strains were detectable earlier in the blood of infected mice and grew faster in the lungs. We hypothesised that these differences were induced by lineage-specific variation in cell envelope lipids. Whole lipid extracts from East Asian/Beijing and Indo-Oceanic strains induced higher concentrations of TNF-α from macrophages than Euro-American lipids. The lipid extracts were fractionated and compared by thin layer chromatography to reveal a distinct pattern of lineage-associated profiles. A phthiotriol dimycocerosate was exclusively produced by East Asian/Beijing strains, but not the phenolic glycolipid previously associated with the hyper-virulent phenotype of some isolates of this lineage. All Indo-Oceanic strains produced a unique unidentified lipid, shown to be a phenolphthiocerol dimycocerosate dependent upon an intact pks15/1 for its production. This was described by Goren as the 'attenuation indictor lipid' more than 40 years ago, due to its association with less virulent strains from southern India. Mutation of pks15/1 in a representative Indo-Oceanic strain prevented phenolphthiocerol dimycocerosate synthesis, but did not alter macrophage cytokine induction. Our findings suggest that the early interactions between M. tuberculosis and host are determined by the lineage of the infecting strain; but we were unable to show these differences are driven by lineage-specific cell-surface expressed lipids.

PMID: 21931620 [PubMed - indexed for MEDLINE]

Dysregulation in retinal para-inflammation and age-related retinal degeneration in CCL2 or CCR2 deficient mice.

PLoS One. 2011;6(8):e22818

Authors: Chen M, Forrester JV, Xu H

Abstract
We have shown previously that a para-inflammatory response exists at the retinal/choroidal interface in the aging eye; and this response plays an important role in maintaining retinal homeostasis under chronic stress conditions. We hypothesized that dysregulation of the para-inflammatory response may result in an overt pro-inflammatory response inducing retinal degeneration. In this study, we examined this hypothesis in mice deficient in chemokine CCL2 or its cognate receptor CCR2. CCL2- or CCR2-deficient mice developed retinal degenerative changes with age, characterized as retinal pigment epithelial (RPE) cell and photoreceptor cell death. Retinal cell death was associated with significantly more subretinal microglial accumulation and increased complement activation. In addition, monocytes from CCL2- or CCR2-deficient mice had reduced capacity for phagocytosis and chemotaxis, expressed less IL-10 but more iNOS, IL-12 and TNF-α when compared to monocytes from WT mice. Complement activation at the site of RPE cell death resulted in C3b/C3d but not C5b-9 deposition, indicating only partial activation of the complement pathway. Our results suggest that altered monocyte functions may convert the protective para-inflammatory response into an overtly harmful inflammation at the retina/choroidal interface in CCL2- or CCR2-deficient mice, leading to RPE and photoreceptor degeneration. These data support a concept whereby a protective para-inflammatory response relies upon a normally functioning innate immune system. If the innate immune system is deficient chronic stress may tip the balance towards an overt inflammatory response causing cell/tissue damage.

PMID: 21850237 [PubMed - indexed for MEDLINE]

Antigen presenting cells in a non-mammalian model system, the chicken.

Immunobiology. 2011 Nov;216(11):1177-83

Authors: Wu Z, Kaiser P

Abstract
The chicken has a different repertoire of tissues, cells and genes of the immune response compared to mammals, yet generally survives infection with viral, bacterial, protozoal and fungal pathogens, and also worms and ectoparasites, just like mammals. Poultry are also probably the most heavily vaccinated group of farmed animals. Antigen presentation to the adaptive immune response therefore obviously normally occurs efficiently in birds. Although comparatively much is known about macrophages and B cells in the chicken, there is as yet little work on the other, professional, antigen-presenting cells, the dendritic cells (DC). Birds also have at least two other sets of phagocytic cells, heterophils and thrombocytes, which may also have the ability to present antigen. Here we review the current state of knowledge about antigen presenting cells in the chicken, concentrating mainly on recent advances in our knowledge of DC.

PMID: 21719145 [PubMed - indexed for MEDLINE]

Clinical and immunomodulatory effects of celecoxib plus interferon-alpha in metastatic renal cell carcinoma patients with COX-2 tumor immunostaining.

J Clin Immunol. 2011 Aug;31(4):690-8

Authors: Schwandt A, Garcia JA, Elson P, Wyckhouse J, Finke JH, Ireland J, Triozzi P, Zhou M, Dreicer R, Rini BI

Abstract
INTRODUCTION: Cycloxygenase-2 (COX-2) is an enzyme involved in prostaglandin E2 (PGE(2)) synthesis associated with higher renal cell carcinoma stage. COX-2 inhibition enhances interferon (IFN-α) anti-tumor immune effects in pre-clinical models. A phase II trial of celecoxib and IFN-α in a targeted population of metastatic renal cell carcinoma patients with maximal COX-2 expression was conducted.
METHODS: Cytokine-naive metastatic renal cell carcinoma patients with tumors expressing ≥10% maximal COX-2 staining by immunohistochemistry received IFN-α 5 million units daily and celecoxib 400 mg orally twice daily in an open-label, single-arm phase II trial.
RESULTS: There were 3 partial responses among 17 patients (objective response rate 18%; 95% confidence interval, 4-43%). Time to progression was 5.6 months. Increased tumor staining 3+ for COX-2 was associated with increased baseline peripheral blood PGE(2) levels, and these patients demonstrated less PGE(2) decrease with therapy. Patients with more 3+ COX-2 staining had significantly more CD3(+) (p = 0.004) and CD4(+) (p = 0.002) IFN-γ T cells at baseline and a significantly greater decrease in these cells with therapy.
DISCUSSION: Celecoxib plus IFN-α in renal cell carcinoma (RCC) patients with maximally staining COX-2 tumors does not significantly enhance overall response rates over IFN monotherapy.
CONCLUSION: COX-2-expressing RCC demonstrates inherent immunosuppression. COX-2 inhibition with IFN results in minimal immunomodulation and no augmented clinical activity in RCC.

PMID: 21487892 [PubMed - indexed for MEDLINE]

Identification of a novel peptide derived from the M-phase phosphoprotein 11 (MPP11) leukemic antigen recognized by human CD8+ cytotoxic T lymphocytes.

Hematol Oncol Stem Cell Ther. 2010;3(1):24-33

Authors: Al Qudaihi G, Lehe C, Dickinson A, Eltayeb K, Rasheed W, Chaudhri N, Aljurf M, Dermime S

Abstract
BACKGROUND AND OBJECTIVES: There is an urgent need for the development of leukemia-targeted immunotherapeutic approaches using defined leukemia-associated antigens that are preferentially expressed by most leukemia subtypes and absent or minimally expressed in vital tissues. M-phase phosphoprotein 11 protein (MPP11) is extensively overexpressed in leukemic cells and therefore is considered an attractive target for leukemia T cell therapy. We sought to identify potential CD8+ cytotoxic T lymphocytes that specifically recognised peptides derived from the MPP11 antigen.
METHODS: A computer-based epitope prediction program SYFPEITHI, was used to predict peptides from the MPP11 protein that bind to the most common HLA- A*0201 molecule. Peptide binding capacity to the HLA-A*0201 molecule was measured using the T2 TAP-deficient, HLA-A*0201-positive cell line. Dendritic cells were pulsed with peptides and then used to generate CD8+ cytotoxic T lymphocytes (CTL). The CML leukemic cell line K562-A2.1 naturally expressing the MPP11 antigen and engineered to express the HLA-A*0201 molecule was used as the target cell.
RESULTS: We have identified a potential HLA-A*0201 binding epitope (STLCQVEPV) named MPP-4 derived from the MPP11 protein which was used to generate a CTL line. Interestingly, this CTL line specifically recognized peptide-loaded target cells in both ELISPOT and cytotoxic assays. Importantly, this CTL line exerted a cytotoxic effect towards the CML leukemic cell line K562-A2.1.
CONCLUSION: This is the first study to describe a novel epitope derived from the MPP11 antigen that has been recognized by human CD8+ CTL.

PMID: 20231810 [PubMed - indexed for MEDLINE]