|
|
 Previous Article | Table of Contents | Next Article 
Blood, Vol. 91 No. 2 (January 15), 1998:
pp. 392-398
High Efficiency Adenovirus-Mediated Gene Transfer to Human Dendritic
Cells
By
Allan B. Dietz and
Stanimir Vuk-Pavlovi
From the Stem Cell Laboratory, Mayo Cancer Center, Mayo Clinic and
Mayo Foundation, Rochester, MN.
 |
ABSTRACT |
The interest in the use of human dendritic cells in cancer
immunotherapy calls for efficient ex vivo methods of dendritic cell
education. To extend the range of methods available, we generated phenotypically characteristic dendritic cells from peripheral blood
monocytes incubated with granulocyte-macrophage colony-stimulating factor and interleukin-4 and infected them with an adenovirus containing a humanized version of green fluorescent protein as a marker
of gene expression. The levels of expressed protein were high, but they
were further increased in combination with cationic liposomes. In
comparison to transfection efficiency of the homologous expression
plasmid, adenovirus-mediated gene transfer was substantially more
efficient. With the aid of liposome-mediated infection, gene transfer
into CD83+ dendritic cells was highly effective,
resulting in more than 90% of the cells expressing the transgene.
| |
INTRODUCTION |
DENDRITIC CELLS process intracellular and
internalized antigens and present them to the naive T
cells.1 In vivo suppression of dendritic cell function
plays a role in pathogenesis of cancer2 and infectious
diseases, including acquired immunodeficiency syndrome.3,4 To overcome this suppression, ex vivo maturation of dendritic cell
precursors in the presence of antigen(s) has been explored as part of
an immunotherapeutic approach to malignancy.5-10 Generally, isolated dendritic cells have been incubated in vitro (pulsed) with
tumor-specific peptides5,7; preliminary reports on the
effectiveness of this technique have been
encouraging.5,7,11,12
Modification of gene expression in dendritic cells could provide
important advantages over peptide pulsing. For example, expression of a
transgene could extend the duration of antigen presentation. Also,
expression of transgenes for cytokines and chemokines, or for these
molecules fused with target peptides, could induce more potent immune
responses than peptides alone. Such fused cytokines and peptides have
been used as vaccines for B-cell lymphoma.10
Unlike somatic gene therapy that generally requires stable long-term
expression, education of dendritic cells for antigen presentation in
immunotherapy could benefit from a burst of transgene expression that
occurs in parallel with ex vivo dendritic cell processing. The hitherto
modes of gene delivery to dendritic cells include high-speed in vivo
delivery of naked DNA bound to microspheres,13 transfection
with plasmid constructs,14 and transduction with recombinant retroviruses.15,16 These techniques have shown delivery and expression of transgenes, but they have been more compatible with the requirements of somatic gene therapy. Therefore, for efficient DNA delivery to dendritic cells, as well as the favorable
kinetics of transgene expression in infected cells, we considered
recombinant adenoviruses. These viruses have been used successfully as
vectors for DNA transfer primarily into endothelial and epithelial
cells17,18 and recently into dendritic cells.19 Compared with other methods, adenovirus-mediated transfer stimulated the strongest antigen-specific cytotoxic T-lymphocyte
(CTL) response in mice and provided protective immunity to
a lethal antigen challenge.20,21
Recent advances in isolation and culture of dendritic cells from
circulating mononuclear cell precursors make it possible to prepare
dendritic cells in numbers useful for clinical trials.22-25 These cells are equivalent to mature blood-derived dendritic cells in
phenotype, morphology, and function.22 Availability of
these cells necessitates development of highly efficientmethods of gene transfer that would not compromise the later use of the cells in
humans. The lack of adenovirus integration into the genome and the high
level of transgene expression make this technique worth considering in
ex vivo antigen education of dendritic cells for therapy of malignant
and infectious diseases. In this communication, we report the use of a
recombinant adenovirus in combination with cationic liposomes for
highly effective gene transfer and expression in human dendritic cells.
| |
MATERIALS AND METHODS |
Cell culture media and recombinant cytokines.
In all experiments, we used the complete medium, RPMI-1640 supplemented
with 10% fetal bovine serum, 2 mmol/L L-glutamine, 100 U/mL
penicillin, 100 µg/mL streptomycin (all from GIBCO BRL, Gaithersburg,
MD). Recombinant granulocyte-macrophage colony-stimulating factor
(GM-CSF), interleukin-4 (IL-4), and tumor necrosis factor- (TNF-)
were obtained from R&D Systems (Minneapolis, MN). Cells were incubated
in a water-vapor saturated atmosphere containing 5% CO2 at
37°C.
Isolation of dendritic cells.
Dendritic cells were derived from monocytes isolated by immunomagnetic
adsorption26 and cultured essentially as described by
others.25 Buffy coat, derived from 1 U of whole blood drawn from a healthy volunteer, was supplied by the Components Laboratory, Department of Transfusion Medicine, Mayo Clinic, in compliance with
institutional guidelines. The cells, suspended in 50 mL of citrated
plasma, were mixed with an equal volume of phosphate-buffered saline
(PBS). The cell suspension (in aliquots of 8 mL) was layered onto 6 mL
of Lymphocyte Separation Medium (Organon Teknika, Durham, NC) into 12 15-mL conical tubes. The tubes were centrifuged at 400g for 30 minutes at room temperature (all centrifugation was performed at room
temperature unless noted). The cells layered between the phases were
aspirated and combined into 50 mL conical tubes, washed twice with 50 mL of PBS, and centrifuged at 250g for 10 minutes. The cells
were washed with 50 mL cold PBSFE (PBS with 0.5% bovine serum albumin
and 2 mmol/L EDTA), counted, and assayed for viability by trypan blue
exclusion.
Peripheral blood mononuclear cells (5 × 108 total
cells) were used as the source for immunomagnetic isolation of
CD14+ leukocytes according to the manufacturer's
instructions (Miltenyi, Auburn, CA). CD14+ cells (8 × 107) were resuspended at 5 × 106 cells/mL
in serum-free RPMI-1640 for 1 hour at 37°C. The nonadherent cells
were discarded and the attached cells were rinsed with PBS (Fig 1A). Complete medium was supplemented
with 1,000 IU/mL of GM-CSF and 1,000 IU/mL IL-4. Within 48 hours,
loosely attached grapelike clusters formed with some remaining adherent
cells (Fig 1B). Two days after plating with cytokines, the cells were
suspended at 2 × 106 cells/mL of complete medium with
GM-CSF and IL-4 and incubated for an additional 72 hours. Cells were
harvested by vigorous pipetting, counted, and used for flow cytometry
or gene transfer. At this stage, cells were more than 92% viable as
determined by trypan blue exclusion.
View larger version (148K):
[in this window]
[in a new window]
View larger version (146K):
[in this window]
[in a new window]
| Fig 1.
Morphology of isolated cells in culture. (A) Purified
adherent monocytes used for initiation of dendritic cells. (B) After 48 hours in culture with GM-CSF and IL-4, monocytes clustered into
aggregates typical of dendritic cell precursors.
|
|
Adenovirus-mediated gene transfer.
For gene transfer, we used the recombinant adenovirus Ad5RSVGFP alone,
Ad5RSVGFP in combination with cationic liposomes,27 the
expression plasmid pRSVGFP containing the adenovirus expression cassette, and pRSVGFP with cationic liposomes. Ad5RSVGFP contained a
humanized version of the green fluorescent protein (GFP; GFP-S65T; Clontech, Palo Alto, CA) under the control of the Rous Sarcoma virus
(RSV) promoter. Ad5RSVGFP and pRSVGFP were provided by the University
of Iowa Gene Transfer Vector Core (Iowa City, IA). The adenovirus
contained 1.1 × 109 particles/µL and was stored at
 70°C until needed.
For infection, AdRSVGFP was prepared by suspending 7.5 × 109 particles (5,000 particles per cell) in 50 µL total
serum free medium (Opti-MEM; GIBCO BRL). Meanwhile, 1.25 µg cationic
liposomes (Lipofectamine; GIBCO BRL) were suspended in 50 µL
serum-free medium. For experiments without liposomes, the adenovirus
was suspended in 100 µL serum-free medium. For plasmid transfections, we used the equivalent cell numbers, liposome amounts, and virus expression equivalents (5,000 plasmids per cell) as for Ad5RSVGFP infection. The vector and liposome solutions were mixed and incubated for 15 minutes.
For gene tranfer experiments, purified dendritic cells were centrifuged
at 250g for 10 minutes and the medium was carefully removed.
The cells (1.5 × 106 cells for each trial) were
resuspended in 100 µL of serum-free medium and combined with the
vector or vector/liposome. The suspension was gently mixed and
incubated for 105 minutes at 37°C. (The final adenovirus/cell ratio
was 5,000 virus particles per cell, resulting in a multiplicity of
infection factor of about 50.) After incubation, the cells were
centrifuged, the supernatant was removed, and the cells were
resuspended at 0.5 × 106 cells/mL in complete medium
containing 400 IU/mL GM-CSF and 200 IU/mL TNF-. The cells were
incubated at 37°C for 72 hours before analysis for GFP expression
by flow cytometry and fluorescence microscopy.
Flow cytometric characterization of dendritic cells.
The purity of dendritic cells was assessed by flow cytometry with the
aid of a FACScan cytometer (Becton Dickinson, Sunnyvale, CA) at the
Mayo Flow Cytometry Core Facility. All immunoreagents were obtained
commercially and were used with appropriate isotype controls. The
reagent for CD56 was obtained from Becton Dickinson (San Jose, CA); for
CD14, CD19, CD1a, CD54, CD3, CD4, CD11c, fluorescein isothiocyanate
(FITC) isotype control, phycoerythrin (PE) isotype control, and HLA-DR
from BioSource International (Camarillo, CA); for CD83 and CD64 from
Coulter (Hialeah, FL); and for CD86 from Ancell (Bayport, MN). For each
analysis, 100,000 cells in 100 µL PBS were incubated for 30 minutes
on ice with the manufacturer's recommended amount of PE-labeled or
FITC-labeled antibody. After incubation, the cells were washed with 3 mL of 0.1% sodium azide in PBS and centrifuged and the supernatant was
removed. The cells were fixed with 500 µL of 2% paraformaldehyde in
PBS and 10,000 events were analyzed for surface phenotype by flow
cytometry (Fig 2).
View larger version (42K):
[in this window]
[in a new window]
View larger version (33K):
[in this window]
[in a new window]
| Fig 2.
Flow cytometric analysis of human dendritic cells
cultured from CD14+ leukocytes. (A) Status of 12 markers
on dendritic cells after incubation with GM-CSF and IL-4; these cells
were subsequently used for gene transfer. Cells in the dot plot (upper
row left) were gated for homogeneity of size (forward scatter) and
shape (side scatter). (B) Status of HLA-DR, CD83, CD1a, CD54, CD11c, and CD86 after ex vivo maturation with TNF- (shaded) shown together with the status before maturation.
|
|
Quantitative and qualitative characterization of expressed GFP in
dendritic cells.
Expression of the GFP transgene was characterized by flow cytometry
with the instrument set for fluorescein detection. Three days after
infection, the majority of cells did not adhere to the substrate. These
cells were washed from the plates and combined with the adherent cells
collected after scraping the culture flask. The cells were centrifuged
and fixed for flow cytometry as described. Alternatively, dendritic
cells were visualized with an IM35 Zeiss fluorescent microscope
equipped with the fluorescein filter set and photographed with ISO 200 speed Royal Gold or PJM Multispeed film (Kodak, Rochester, NY).
All experiments were performed at least in duplicate with similar
results.
| |
RESULTS |
Dendritic cells were cultured from monocytes by a technique that
required minimum manipulation and resulted in high
yields.22-24 Positive isolation of monocytes by
magnetically labeled CD14-specific antibodies increased dendritic cell
purity without reducing cell yield. Starting with 5 × 108 peripheral blood mononuclear cells, we typically ended
up with more than 3 × 107 dendritic cell precursors.
Figure 2A shows the results of flow cytometric analysis of cells after
incubation with GM-CSF and IL-4. The cells contained markers typical of
dendritic cell precursors22-25,28; they were positive for
HLA-DR, CD54 (ICAM), CD11c, CD86, CD4, and CD1a and negative for CD56
(natural killer [NK] cells), CD64 (monocytes/macrophages), CD3 (T
cells), and CD19 (B cells). Significantly, the cells were negative for
CD83, a marker of mature dendritic cells.22,28 Some
dendritic cells in this system expressed CD14 upon development of the
ability to adhere,23 although this has not been
consistently documented.22 In addition to adherent cells,
the culture contained some multicell aggregates typical of
proliferating dendritic cells (Fig 1B).
Incubation with GM-CSF and TNF- transformed the immature dendritic
cells into veiled cells, strongly adherent cells with dendritic
appendages, and some remaining cell clusters. Flow cytometric analysis
indicated an increase in expression of CD83, HLA-DR, CD54, and CD86
(Fig 2B). Although the cells were still CD1a+ and
CD11c+, the level of expression of these markers was
reduced. CD83 has been characterized as a unique cell surface marker of
mature dendritic cells.22,28 An analysis of cell size
(forward scatter) and cell geometry (side scatter) showed a discrete
population of larger and less regular cells (Fig 2A). In this discrete
population, we found that more than 50% of the cells were
CD83+ (Fig 2B). Significantly, more than 90% of cells in
this population were also HLA-DR+ (Fig 2B). Thus, the large
cells in the gated region were a population of mature dendritic cells.
To determine the role of the recombinant adenovirus in GFP expression
in dendritic cells, we transfected the cells before maturation with
pRSVGFP, the plasmid contained in Ad5RSVGFP
(Fig 3B), and with pRSVGFP in the presence
of liposomes (Fig 3C). Also, we infected the cells with the
equivalent (in comparison to pRSVGFP) amount of Ad5RSVGFP in the
absence (Fig 3D) or in the presence of liposomes (Fig 3E).
View larger version (38K):
[in this window]
[in a new window]
| Fig 3.
GFP expression in human dendritic cells infected with
pRSVGFP and Ad5RSVGFP in the absence and the presence of cationic
liposomes. Cells in the gated region (Fig 2A) were analyzed for GFP
fluorescence. They were untreated (A), treated with pRSVGFP in the
absence (B) and in the presence of liposomes (C), or infected with
Ad5RSVGFP in the absence (D) and in the presence of liposomes (E).
|
|
Whereas cells treated with pRSVGFP alone (Fig 3B) were no more
fluorescent that untreated controls (Fig 3A), inclusion of liposomes in
the transfection medium resulted in GFP expression in some 5% of cells
(Fig 3C). Interestingly, the equivalent amount of Ad5RSVGFP alone
resulted in more successful gene transfer; about 20% of dendritic
cells expressed GFP (Fig 3D). Adenovirus combined with liposomes was
the most effective, resulting in approximately 90% of cells expressing
GFP (Fig 3E). In control cells (Fig 3A) and cells transfected with the
plasmid (with or without liposomes; Fig 3B and C), the mean
fluorescence intensity, a measure of the average level of GFP
expression per cell, was low and indistinguishable among the groups.
However, cells infected with the adenovirus (Fig 3D) were characterized
by the mean fluorescence intensity 2.5 times above control and the
cells treated with the adenovirus and liposomes (Fig 3E) were
characterized by the mean fluorescence intensity 8.5 times above
control. Clearly, both adenovirus infection and liposome treatment
increased the levels of transgene expression.
To determine whether cells expressing GFP were mature dendritic cells,
we analyzed CD83 and HLA-DR fluorescence, respectively, versus GFP
fluorescence. Of CD83+ cells, 90% expressed GFP
(Fig 4C). Similarly, of HLA-DR+
cells, some 90% expressed GFP (Fig 4D). Thus, the GFP+
cells harbored the two characteristic markers of dendritic cells.
View larger version (43K):
[in this window]
[in a new window]
| Fig 4.
Expression of CD83 (A and C) and HLA-DR (B and D) versus
green fluorescence in uninfected cells (A and B) and cells infected with Ad5RSVGFP in the presence of liposomes (C and D). The vertical and
horizontal lines distinguish CD83 and
HLA-DR cells (left of and below the line, respectively)
from CD83+ and HLA-DR+ cells.
|
|
GFP expression was observed in adherent, loosely adherent, and
nonadherent cells (Fig 5). The cells were
morphologically typical: the unattached cells contained small
pseudopodia and the attached cells displayed long bifurcated
appendages. Thus, infection and transgene expression had no apparent
effect on dendritic cell morphology and viability.
View larger version (138K):
[in this window]
[in a new window]
View larger version (108K):
[in this window]
[in a new window]
| Fig 5.
Expression of GFP in cultured dendritic cells. (A)
Transmitted light micrograph of dendritic cells 3 days after infection with AdRSVGFP in the presence of cationic liposomes. (B) Same field as
in (A) under fluorescence detection using a fluorescein filter set. GFP
was expressed in adherent, nonadherent, and loosely attached cells.
|
|
| |
DISCUSSION |
The purpose of this work was to establish a technique for high
efficiency gene transfer into human dendritic cells derived from
peripheral blood. We used expression of GFP as proof of infection and
effective expression of the transgene. To increase the efficiency of
transgene expression, we applied the recently described
liposome-enhanced adenovirus infection27 to human dendritic
cells. Adenoviruses alone can infect these cells, but they are rather
ineffective (ie, they need high levels of multiplicity of
infection19). By the use of liposomes, we dramatically
increased infection efficiency.
Although the adenovirus combined with liposomes was highly infective,
it was unclear whether the virus contributed to gene expression in
comparison to the plasmid transfection in the presence of liposomes. To
resolve the role of the adenovirus, we studied gene expression in cells
transfected with the plasmid containing the adenovirus expression
cassette. In cells transfected with the plasmid alone, there was
virtually no transgene expression. Liposomes did induce measurable
expression, but it was more than one order of magnitude below the level
obtained with the adenovirus/liposome combination. Clearly, the role of
the adenovirus was critical even when the transgene was transferred
across the cell membrane by the liposome-mediated
mechanism.27 The practical consequence of this finding is
that one can achieve superinfectivity with rather low amounts of
adenovirus. Thus, the high levels of gene expression achieved by the
use of adenovirus in combination with liposomes make this system well
suited for antigen expression and presentation.
GFP expression by cells infected by the adenovirus alone indicates that
dendritic cells harbor CD51 (integrin-v), the molecule critical for effective adenovirus infection, or its functional equivalent. In a murine dendritic cell line, GM-CSF increased the
expression of CD51.21 Expression of CD51 as part of the functional integrin v  3 and/or
v 5 raises questions about the role of
vitronectin binding integrins in the biology of dendritic cells and
particularly in their maturation. In mice, vitronectin receptor was
found in thymocytes, splenocytes, and bone marrow cells.29
In thymocytes, expression of vitronectin receptor depended on the stage
of development.29 Expression of vitronectin receptors in
dendritic cells may help colocalize these cells with developing T cells
in the course of immune system education. This hypothesis is in line
with the observation that blocking adhesion also blocks T-cell
proliferation. Work is underway in our laboratory to determine the role
of CD51 in human dendritic cells.
A salient feature of this work is that liposomes enhanced the
adenovirus-mediated gene expression without any apparent interference with dendritic cell maturation; high levels of CD83 and HLA-DR were
measured in infected cells. Because CD83 is highly expressed in
adenovirus/liposome-treated cells (this work) and coexpressed with
CD1a,b,c, class I, class II, CD80, and CD86, it is likely that the
adenovirus/liposome-treated cells contain all the factors required for
their function in immunity.
The ability of antigen-presenting dendritic cells to elicit a specific
immune response has stimulated interest in the use of dendritic cells
as adjuvants in immunotherapy of tumors and of autoimmune and
infectious diseases. Currently, clinical trials are conducted to
evaluate the usefulness of ex vivo educated dendritic cells in therapy
of B-cell lymphoma7 and prostate cancer,30 whereas laboratory observations promise that this treatment modality will soon be extended to other malignant diseases.31 Thus,
the high efficiency gene transfer and transgene expression by dendritic cells can extend the range of potential targets for dendritic cell
mediated immunotherapy. To investigate this hypothesis, we are
studying antigen-dependent stimulation of the T-cell response by
transgene expressing dendritic cells.
The technique that we have described is rapid and requires only a
single additional manipulation step to the standard dendritic cell
isolation protocol. It results in high levels of transgene expression
in CD83/HLA-DR+ dendritic cells. An additional advantage of
adenoviruses in ex vivo manipulation of cells for human use is their
inability to integrate into the genome and change the oncogenic
potential of manipulated cells. Ex vivo adenovirus-mediated gene
transfer fits well into future clinical protocols for isolation and ex
vivo education of dendritic cells.
| |
FOOTNOTES |
Submitted August 18, 1997;
accepted October 24, 1997.
Supported by a grant from Adelyn L. Luther, Singer Island, FL, and by
the Mayo Cancer Center. A.B.D. is a Glen and Florence Voyles Foundation
Scholar. The University of Iowa Gene Transfer Vector Core is supported
in part by a trust from the Carver Foundation.
Address reprint requests to Allan B. Dietz, PhD, Stem Cell Laboratory,
Mayo Cancer Center, Room 1311B Guggenheim, 200 First St SW, Rochester,
MN 55905.
The publication costs of this article were defrayed in part by page
charge payment. This article must therefore be hereby marked
"advertisement" in accordance with 18 U.S.C. section 1734 solely
to indicate this fact.
| |
ACKNOWLEDGMENT |
The authors thank the Mayo Flow Cytometry Core Facility for expert
assistance, Dr Beverly Davidson and Richard Anderson of the University
of Iowa Vector Core Facility for the supply of pRSVGFP and Ad5RSVGFP,
and Dr Franklyn G. Prendergast for his continuing interest and support.
| |
REFERENCES |
1.
Steinman RM:
The dendritic cell system and its role in immunogenicity.
Annu Rev Immunol
9:271,
1991[Medline]
[Order article via Infotrieve]
2.
Gabrilovich DI,
Chen HL,
Girgis KR,
Cunningham HT,
Meny GM,
Nadaf S,
Kavanaugh D,
Carbone DP:
Production of vascular endothelial growth factor by human tumors inhibits the functional maturation of dendritic cells.
Nat Med
2:1096,
1996[Medline]
[Order article via Infotrieve]
3.
Gabrilovich DI,
Patterson S,
Timofeev AV,
Harvey JJ,
Knight SC:
Mechanism of dendritic cell dysfunction in retrovirual infection of mice.
Clin Immunol Immunopathol
80:139,
1996[Medline]
[Order article via Infotrieve]
4.
Patterson S,
Gross J,
English N,
Stackpoole A,
Bedford P,
Knight SC:
CD4 expression on dendritic cells and their infection by human immunodeficiency virus.
J Gen Virol
76:1155,
1995[Abstract/Free Full Text]
5.
Inaba K,
Metlay JP,
Crowley MT,
Steinman RM:
Dendritic cells pulsed with protein antigens in vitro can prime antigen-specific, MHC-restricted T cells in situ.
J Exp Med
172:631,
1990[Abstract/Free Full Text]
6.
Gong J,
Chen D,
Kashiwaba M,
Kufe D:
Induction of antitumor activity by immunization with fusions of dendritic and carcinoma cells.
Nat Med
3:558,
1997[Medline]
[Order article via Infotrieve]
7.
Hsu FJ,
Benike C,
Fagnoni F,
Liles TM,
Czerwinski D,
Taidi B,
Engleman EG,
Levy R:
Vaccination of patients with B-cell lymphoma using autologous antigen-pulsed dendritic cells.
Nat Med
2:52,
1996[Medline]
[Order article via Infotrieve]
8.
Gabrilovich DI,
Nadaf S,
Corak J,
Berzofsky JA,
Carbone DP:
Dendritic cells in antitumor immune responses. II. Dendritic cells grown from bone marrow precursors, but not mature DC from tumor-bearing mice, are effective antigen carriers in the therapy of established tumors.
Cell Immunol
170:111,
1996[Medline]
[Order article via Infotrieve]
9.
Mayordomo JI,
Zorina T,
Storkus WJ,
Zitvogel L,
Celluzzi C,
Falo LD,
Melief CJ,
Ildstad ST,
Kast WM,
Deleo AB,
Lotze MT:
Bone marrow-derived dendritic cells pulsed with synthetic tumour peptides elicit protective and therapeutic antitumour immunity.
Nat Med
1:1297,
1995[Medline]
[Order article via Infotrieve]
10.
Tao M-H,
Levy R:
Idiotype/granulocyte-macrophage colony-stimulating factor fusion protein as a vaccine for B-cell lymphoma.
Nature
362:755,
1993[Medline]
[Order article via Infotrieve]
11.
Paglia P,
Chiodoni C,
Rodolfo M,
Colombo MP:
Murine dendritic cells loaded in vitro with soluble protein prime cytotoxic T lymphocytes against tumor antigen in vivo.
J Exp Med
183:317,
1996[Abstract/Free Full Text]
12.
Zitvogel L,
Mayordomo JI,
Tjandrawan T,
DeLeo AB,
Clarke MR,
Lotze MT,
Storkus WJ:
Therapy of murine tumors with tumor peptide-pulsed dendritic cells: Dependence on T cells, B7 costimulation, and T helper cell 1-associated cytokines.
J Exp Med
183:87,
1996[Abstract/Free Full Text]
13.
Condon C,
Watkins SC,
Celluzzi CM,
Thompson K,
Falo LDJ:
DNA-based immunization by in vivo transfection of dendritic cells.
Nat Med
2:1122,
1996[Medline]
[Order article via Infotrieve]
14.
Alijagic S,
Moller P,
Artuc M,
Jurgovsky K,
Czarnetzki BM,
Schadendorf D:
Dendritic cells generated from peripheral blood transfected with human tyrosinase induce specific T cell activation.
Eur J Immunol
25:3100,
1995[Medline]
[Order article via Infotrieve]
15.
Henderson RA,
Nimgaonkar MT,
Watkins SC,
Robbins PD,
Ball ED,
Finn OJ:
Human dendritic cells genetically engineered to express high levels of the human epithelial tumor antigen mucin (MUC-1).
Cancer Res
56:3763,
1996[Abstract/Free Full Text]
16.
Reeves ME,
Royal RE,
Lam JS,
Rosenberg SA,
Hwu P:
Retroviral transduction of human dendritic cells with a tumor-associated antigen gene.
Cancer Res
56:5672,
1996[Abstract/Free Full Text]
17.
Wickham TJ,
Mathias P,
Cheresh DA,
Nemerow GR:
Integrins v3 and v5 promote adenovirus internalization but not virus attachment.
Cell
73:309,
1993[Medline]
[Order article via Infotrieve]
18.
Greber UF,
Willetts M,
Webster P,
Helenius A:
Stepwise dismantling of adenovirus 2 during entry into cells.
Cell
75:477,
1993[Medline]
[Order article via Infotrieve]
19.
Arthur JF,
Butterfield LH,
Roth MD,
Bui LA,
Kiertscher SM,
Lau R,
Dubinett S,
Glaspy J,
McBride WH,
Economou JS:
A comparison of gene transfer methods in human dendritic cells.
Cancer Gene Ther
4:17,
1997[Medline]
[Order article via Infotrieve]
20.
Ribas A,
Butterfield LH,
McBride WH,
Jilani SM,
Bui LA,
Vollmer CM,
Lau R,
Dissette VB,
Hu B,
Chen AY,
Glaspy JA,
Economou JS:
Genetic immunization for the melanoma antigen MART-1/MELAN-A using recombinant adenovirus-transduced murine dendritic cells.
Cancer Res
57:2865,
1997[Abstract/Free Full Text]
21.
Brossart P,
Goldrath AW,
Butz EA,
Martin S,
Bevan MJ:
Virus-mediated delivery of antigenic epitopes into dendritic cells as a means to induce CTL.
J Immunol
158:3270,
1997[Abstract]
22.
Zhou L-J,
Tedder TF:
CD14+ blood monocytes can differentiate into functionally mature CD83+ dendritic cells.
Proc Natl Acad Sci USA
93:2588,
1996[Abstract/Free Full Text]
23.
Radmayr C,
Bock G,
Hobisch A,
Klocker H,
Bartsch G,
Thurnher M:
Dendritic antigen-presenting cells from the peripheral blood of renal-cell-carcinoma patients.
Int J Cancer
63:627,
1995[Medline]
[Order article via Infotrieve]
24.
Romani N,
Gruner S,
Brang D,
Kampgen E,
Lenz A,
Trockenbacher B,
Konwalinka G,
Fritsch PO,
Steinman RM,
Schuler G:
Proliferating dendritic cell progenitors in human blood.
J Exp Med
180:83,
1994[Abstract/Free Full Text]
25.
Thurnher M,
Papesh C,
Ramoner R,
Gastl G,
Bock G,
Radmayr C,
Klocker H,
Bartsch G:
In vitro generation of CD83+ human blood dendritic cells for active tumor immunotherapy.
Exp Hematol
25:232,
1997[Medline]
[Order article via Infotrieve]
26.
Miltenyi S,
Muller W,
Weichel W,
Radbruch A:
High gradient magnetic cell separation with MACS.
Cytometry
11:231,
1990[Medline]
[Order article via Infotrieve]
27.
Fasbender A,
Zabner J,
Chillon M,
Moninger TO,
Puga AP,
Davidson BL,
Welsh MJ:
Complexes of adenovirus with polycationic polymers and cationic lipids increase the efficiency of gene transfer in vitro and in vivo.
J Biol Chem
272:6479,
1997[Abstract/Free Full Text]
28.
Zhou L-J,
Tedder TF:
Human blood dendritic cells selectively express CD83, a member of the immunoglobulin superfamily.
J Immunol
154:3821,
1995[Abstract]
29.
Gerber DJ,
Pereira P,
Huang SY,
Pelletier C,
Tonegawa S:
Expression of v and 3 integrin chains on murine lymphocytes.
Proc Natl Acad Sci USA
93:14698,
1996[Abstract/Free Full Text]
30.
Salgaller ML,
Tjoa B,
Lodge PA,
Murphy G,
Ragde H,
Kenny B,
Boynton A:
Monitoring of prostate-specific membrane antigen (PSMA)-restricted responses in patients treated with dendritic cells pulsed with PSMA-derived peptides.
Proc Am Assoc Cancer Res
38:4138,
1997
31.
Choudhury A,
Gajewski JL,
Liang JC,
Popat U,
Claxton DF,
Kliche K-O,
Andreeff M,
Champlin RE:
Use of leukemic dendritic cells for the generation of antileukemic cellular cytotoxicity against Philadelphia chromosome-positive chronic myelogenous leukemia.
Blood
89:1133,
1997[Abstract/Free Full Text]
 CiteULike  Connotea  Del.icio.us  Digg  Reddit  Technorati What's this?
This article has been cited by other articles:
|
|
|
|
 
A. Krause, J. H. Joh, N. R. Hackett, P. W. Roelvink, J. T. Bruder, T. J. Wickham, I. Kovesdi, R. G. Crystal, and S. Worgall
Epitopes expressed in different adenovirus capsid proteins induce different levels of epitope-specific immunity.
J. Virol.,
June 1, 2006;
80(11):
5523 - 5530.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
S. Worgall, A. Busch, M. Rivara, D. Bonnyay, P. L. Leopold, R. Merritt, N. R. Hackett, P. W. Rovelink, J. T. Bruder, T. J. Wickham, et al.
Modification to the Capsid of the Adenovirus Vector That Enhances Dendritic Cell Infection and Transgene-Specific Cellular Immune Responses
J. Virol.,
March 1, 2004;
78(5):
2572 - 2580.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
K. Brown, W. Gao, S. Alber, A. Trichel, M. Murphey-Corb, S. C. Watkins, A. Gambotto, and S. M. Barratt-Boyes
Adenovirus-Transduced Dendritic Cells Injected into Skin or Lymph Node Prime Potent Simian Immunodeficiency Virus-Specific T Cell Immunity in Monkeys
J. Immunol.,
December 15, 2003;
171(12):
6875 - 6882.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
N. Belousova, N. Korokhov, V. Krendelshchikova, V. Simonenko, G. Mikheeva, P. L. Triozzi, W. A. Aldrich, P. T. Banerjee, S. D. Gillies, D. T. Curiel, et al.
Genetically Targeted Adenovirus Vector Directed to CD40-Expressing Cells
J. Virol.,
November 1, 2003;
77(21):
11367 - 11377.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
C. Bello-Fernandez, J. Stasakova, A. Renner, N. Carballido-Perrig, M. Koening, M. Waclavicek, O. Madjic, L. Oehler, O. Haas, J. M. Carballido, et al.
Retrovirus-mediated IL-7 expression in leukemic dendritic cells generated from primary acute myelogenous leukemias enhances their functional properties
Blood,
March 15, 2003;
101(6):
2184 - 2190.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
Y. Tanaka, S. F. Dowdy, D. C. Linehan, T. J. Eberlein, and P. S. Goedegebuure
Induction of Antigen-Specific CTL by Recombinant HIV Trans-Activating Fusion Protein-Pulsed Human Monocyte-Derived Dendritic Cells
J. Immunol.,
February 1, 2003;
170(3):
1291 - 1298.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
E. Andreakos, C. Smith, C. Monaco, F. M. Brennan, B. M. Foxwell, and M. Feldmann
Ikappa B kinase 2 but not NF-kappa B-inducing kinase is essential for effective DC antigen presentation in the allogeneic mixed lymphocyte reaction
Blood,
February 1, 2003;
101(3):
983 - 991.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
T. D. de Gruijl, S. A. Luykx-de Bakker, B. W. Tillman, A. J. M. van den Eertwegh, J. Buter, S. M. Lougheed, G. J. van der Bij, A. M. Safer, H. J. Haisma, D. T. Curiel, et al.
Prolonged Maturation and Enhanced Transduction of Dendritic Cells Migrated from Human Skin Explants After In Situ Delivery of CD40-Targeted Adenoviral Vectors
J. Immunol.,
November 1, 2002;
169(9):
5322 - 5331.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
M. D. Roth, Q. Cheng, A. Harui, S. K. Basak, K. Mitani, T. A. Low, and S. M. Kiertscher
Helper-Dependent Adenoviral Vectors Efficiently Express Transgenes in Human Dendritic Cells but Still Stimulate Antiviral Immune Responses
J. Immunol.,
October 15, 2002;
169(8):
4651 - 4656.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
Abstract
Introduction
Abstract