| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
|
|
|
|||||||
|
|||||||||
Divisions of
*
Surgical Oncology,
Hematology/Oncology, and
Experimental Radiation Oncology, University of California Los Angeles Medical Center, Los Angeles, CA 90095, and
§
Department of Medicine, University of Connecticut Health Center, Farmington, CT 06032
 |
Abstract |
|---|
 Top Abstract IntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
. Finally,
AdVMART1-transduced DC were able to generate MART-127–35
peptide-specific, class I-restricted CTL in PBL cultures from normal
donors. This study supports the use of tumor Ag-engineered DC in
genetic immunotherapy. |
Introduction |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
Dendritic cells (DC) are the most potent APCs identified (13, 14, 15, 16) and are capable of activating naive T cells. A potentially powerful strategy in cancer gene therapy involves the genetic engineering of DC with defined tumor Ag genes and their use as vaccines. We have recently described a murine model of genetic immunotherapy in which DC were transduced with an adenovirus vector expressing the human MART-1 gene (17). Vaccination of mice with murine DC transduced with MART-1 protected mice against a challenge of murine fibrosarcoma cells expressing the MART-1 gene.
Herein, we report the use of AdVMART1 to transduce human DC and generate MART-1-specific CTL in vitro from naive donor blood. DC transduced with AdVMART1 process and present the MART-127–35 immunodominant peptide. CTL generated from AdVMART1-transduced DC can recognize and lyse HLA-A2+/MART-1+ melanoma after only 7 days in culture.
|
Materials and Methods |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
All melanoma lines used (M201, M202, M207, M237, M238) were generated at UCLA, Los Angeles, CA, from surgical specimens, and two (M202 and M207) have been described previously (18). All melanoma lines were screened for HLA-A2 expression by flow cytometry and were subtyped by PCR and direct sequencing by the UCLA Tissue Typing Laboratory (P. Terasaki, director). MART-1 mRNA expression was assessed by RT-PCR, Northern blot analysis (6), or both. The HLA-A2.1+, Ag processing-deficient T2 cells were provided by Peter Cresswell (Yale University School of Medicine, New Haven, CT). Saos-2 osteosarcoma cells were provided by Arnold Berk (UCLA); K562 erythroleukemia cells, the W6/32 and BB7.2 hybridomas, and HepG2 hepatoma cells were obtained from the American Type Culture Collection (ATCC; Manassas, VA). HepG2 and Saos-2 cells were tested for HLA-A2 by flow cytometry.
Adenoviruses
The empty adenovirus vector, AdVRR5, and those containing luciferase (AdVLuc) and ß-galactosidase (AdVLacZ) reporter genes have been described previously and served as controls (18). Briefly, the AdVMART1 (17) contains the 400-bp MART-1 cDNA, originally cloned by RT-PCR and driven by the CMV promoter/enhancer in a pAC-CMVpLpA AdV type 5 backbone. The virus was prepared by recombination of this plasmid with pJM17, which contains the 35-kb AdV genome, deleted in the E1 region, in 293 cells which provide the E1 genes in trans. Recombinant viruses were released into the medium, purified by limiting dilution, and amplified on 293 cells. All viruses used have been purified on CsCl gradients as described (18).
Antibodies
The anti-MART-1 Ab used in immunohistochemistry is derived from clone A103, (Novacastra, Vector Laboratories, Burlingame, CA). Cells to be stained were either grown on coverslips or as cytospin preparations, fixed in 4% paraformaldehyde, and stained with the anti-MART-1 primary Ab. This was followed by an appropriate secondary Ab and peroxidase staining with the Vectastain Elite kit and reagent diaminobenzidine (Vector Laboratories), according to manufacturer’s instructions. Counterstaining was performed with either Giemsa (Mediatech) or hematoxylin (Vector Laboratories), as the described in the figure legend. As a control to ensure primary Ab-specific binding, MART-1+ melanoma cells were stained with every reagent except the primary Ab, and no color reaction was ever observed. The pan-anti-human class I Ab W6/32 and anti-HLA-A2 Ab BB7.2 were purified from hybridoma supernatant by Macrosep 30K column concentration (Filtron, Northborough, MA). The hybridoma cells were obtained from ATCC. Secondary goat anti-mouse FITC and preconjugated CD4 and CD8 Abs were obtained from Caltag (South San Francisco, CA). Anti-ß2-microglobulin used in some class I inhibition cytotoxicity assays was obtained from PharMingen (San Diego, CA).
RT-PCR
RT-PCR for MART-1 mRNA expression was performed as described previously (17). Human ß-actin PCR was used for semiquantitation in PCR reactions. The primers were 5'-GGCATCGTGATGGACTCCG and 3'-GTCGGAAGGTGGACAGCGA. Primers were synthesized by Life Technologies (Gaithersburg, MD).
AdV transduction
Transduction of cell lines was performed in the following manner. Cells were removed from flasks (using trypsin for adherent cells), washed, and resuspended in 1 ml of RPMI 1640 (Life Technologies) with penicillin-streptomycin-Fungizone (PSF; Gemini Bioproducts, Calabasas, CA) and 2% FBS (Gemini). Virus was added at a multiplicity of infection (moi) of 10, and the cells were incubated at 37°C for 2 h. Excess virus was washed off, and the cells were resuspended in growth medium (RPMI 1640-PSF-10% FBS) and replated for 24–48 h until use.
DC were transduced after harvest from the 7-day culture with 2% human AB serum (Omega Scientific, Tarzana, CA)-RPMI-PSF at an moi of 1000 unless stated otherwise. After transduction, DC were rinsed and either left in a tube or plated in 5% autologous serum-containing medium (without addition of exogenous cytokines) for 24–48 h until use.
DC generation
DC were prepared as described (19) with some modifications. Peripheral blood was drawn from healthy volunteers by venous puncture, and lymphocytes were purified by Ficoll (Pharmacia, Piscataway, NJ) gradient separation. Mononuclear cells (3–4 x 107) were plated in T-25 flasks (Costar, Cambridge, MA) in RPMI 1640-PSF-10% human AB serum (except for DC used in CTL generation, which were in 5% autologous serum instead of 10% AB) for 2 h at 37°C in a humidified CO2 incubator. The nonadherent cells were removed by gentle rinsing with PBS, and the loosely adherent cells were cultured in medium with 800 U/ml granulocyte/macrophage colony-stimulating factor (Immunex, Seattle, WA) and 500 U/ml IL-4 (R&D systems, Minneapolis, MN) for 7 days. The nonadherent and loosely adherent cells were harvested by vigorous washing. These cells were generally 50% DC as assessed by morphology and phenotyping (19). To compare levels of class I molecules on the DC surface with and without AdV transduction, DC were transduced as described, and 48 h later they were harvested, washed, and stained for either total class I (W6/32 Ab) or HLA-A2 expression (BB7.2 Ab). Controls included untransduced DC. All DC populations were also stained with the secondary Ab to control for nonspecific binding. The DC were gated based on their large size and granular cytoplasm by forward and side scatter. Flow cytometry was performed on a FACScan (Becton Dickinson, Mountain View, CA.
TIL line generation
The CD8+ MART-127–35-specific TIL line, CS TIL 8.0, was established from a metastatic melanoma nodule of an HLA-A2.1 patient (B.M., unpublished observations). CS TIL 8.0 was expanded in 50 U/ml rIL-2 and restimulated approximately every 10 days with an HLA-A2.1/MART-1+ allogeneic melanoma line and an HLA-A2.1-allogeneic EBV-transformed lymphoblastoid cell line. CS TIL 8.0 is oligoclonal and exhibits MART-127–35 specificity.
Generation of peptide-specific CTL
Peptide-specific CTL were generated using a published protocol by Plebanski (20). Briefly, PBMC from HLA-A2.1 or A2.4 donors were pulsed with MART-1 peptide (AAGIGILTV, prepared by the peptide synthesis facility at UCLA, Dr. Joseph Reeve, director) at 50 µg/ml peptide, at 3 x 107 cells/ml serum-free RPMI, at 37°C for 90 min. The cells were rinsed and plated in wells of a 24-well plate at 3 x 106 cells/well in 10% autologous serum-RPMI-PSF with 10 ng/ml IL-7 (Biosource, Camarillo, CA) and 5 µg/ml keyhole limpet hemocyanin (Sigma, St. Louis, MO) (1.5 ml total volume). The nonadherent cells were restimulated weekly with fresh, autologous, peptide-pulsed and irradiated PBMC at a 1:1 ratio and supplemented with IL-2 (Hoffman-La Roche, Nutley, NJ) at 10 U/ml every 3–4 days.
CTL generation from AdV-transduced DC
DC (prepared as described above from HLA-A2.1 or A2.4 donors) were transduced with AdVMART1 at an moi of 1000 for 2 h. The transduced DC were irradiated and plated at 2–5 x 105 cells/well in a 24-well plate to serve as stimulators for CTL generation. Autologous nonadherent cells were depleted of CD4, CD14, CD19, and CD56+ cells by magnetic bead depletion (Dynal, Lake Success, NY) to prepare CD8+-enriched responder cells. The CD8+ cells were plated with the AdVMART1-transduced DC at 2 x 106 cells/well, in 5% autologous medium plus IL-7 at 10 ng/ml to generate MART-1 CTL. These cultures were supplemented with IL-2 at 10 U/ml every 3–4 days. The CD8+ CTL were restimulated weekly with fresh, autologous AdVMART1-transduced DC at a ratio of 1 DC to 5–10 CD8+ CTL. Most cultures were phenotyped for CD4+ and CD8+ cells on a weekly basis.
In the experiments presented in Fig. 5
, CTL cultures were generated
with PBL responder cells and both AdVMART1 and RR5 AdV-transduced
autologous DC (DC:PBL ratio, 1:10) stimulator cells in the presence of
50 U/ml rIL-2.
|
Cytotoxicity assays were performed as follows. Target cells were harvested, washed, counted (and T2 cells were peptide pulsed at 50 µg/ml), and chromated with 100 µCi of Na251CrO4 (Amersham, Arlington Heights, IL), with shaking incubation at 37°C for 1.5-2 h. CTL were washed, counted, and diluted to desired concentrations in RPMI-10% AB and plated in triplicate wells at 100 µl/well in a round-bottom 96-well plate (Costar). Target cells were washed three times, diluted to 5 x 104 cells/ml, and plated with CTL at 100 µl/well. To control for nonspecific lysis by NK cells, a 10–50-fold excess of K562 cells was added to each target population (except "K562 alone" samples) before adding to CTL (21). For class I inhibition, 20–30 µl of either anti-ß2-microglobulin (PharMingen) or W6/32 Ab were added per well. The plates were spun briefly at 1000 rpm and incubated for 4 h. Supernatants (100 µl/well) were harvested and counted in a gamma counter. Triplicate wells were averaged and % specific lysis was calculated as [(sample - spontaneous release)/(maximum release - spontaneous release)]. For spontaneous release, targets were plated without CTL in 10% AB medium. For maximum release, targets were plated with 5% Nonidet P-40 (IGEPAL; Sigma) detergent.
For cytokine release studies, CTL were washed and plated at 1 x
105 cells/well in a flat-bottom 96-well plate (Costar) in
150 µl of RPMI/10% AB. Stimulator cells were harvested, washed, and
plated at 1 x 105 cells per well in 100 µl. IL-2
was added at 300 U/ml, and the plates were incubated at 37°C. After
24 h, plates were spun briefly to pellet cells, and 100 µl of
duplicate supernatants were assayed for TNF-
or IFN- release in
an ELISA. Ab pairs and cytokine standards were purchased from
PharMingen (anti-human TNF- capture and biotinylated Abs are
clones Mab1 and Mab11, respectively; anti-human IFN- capture and
biotinylated detection Abs are clones NIB42 and 4S.B3, respectively),
and the ELISA assays were performed according to the manufacturer’s
instructions. Plates were read at 450 nm in a Spectra ELISA plate
reader (SLP Lab Instruments, Salzburg, Austria) within 30 min of
assay completion.
Statistical analysis
SE values are presented as error bars in graphs or listed in the tables. Comparisons between experimental groups were performed by Student’s t test to interpret the significance of differences observed, and p values are listed in the figure legends or table footnotes.
|
Results |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
We have previously reported that AdV transduction is an efficient
method for transgene expression in DC (19). Transduction of human DC
with AdVMART1 results in synthesis of MART-1 mRNA. The amount of mRNA,
as detected by semiquantitative RT-PCR, increases with increasing moi
(Fig. 1A). We have previously
determined that an moi of 1000 generates high levels of transgene
expression in human DC (19) with minimal to no cytopathic effect.
MART-1 mRNA expression in these transduced DC was detectable for the
entire 8-day culture period (Fig. 1B).
|
). The MART-1-positive M202 melanoma
cell line has strong MART-1 staining (not shown), and untransduced DC
and AdVRR5-transduced DC have only very faint background staining (not
shown and Fig. 2A, respectively). DC transduced with
AdVMART1 show strong positive staining in >90% of the DC (Fig. 2B). Untransduced or AdVRR5-transduced HepG2 cells show only
faint background staining, whereas AdVMART1-transduced HepG2 cells are
strongly positive for MART-1 protein expression (data not shown).
|
The E3/19K gene product of AdV has been reported to down-regulate
cell surface class I expression by affecting ER transport (22, 23). To
assess any effect from E3-intact AdVMART1 transduction,
HLA-A2+ DC or HepG2 cells were transduced with AdVMART1 and
assessed for class I (W6/32 Ab) and HLA-A2 (BB7.2 Ab) surface
expression by flow cytometry. Large, granular DC were gated on as shown
in the dot plot (Fig. 3A).
There is only a minor decrease in total class I or HLA-A2 expression in
DC transduced with AdVMART1 (15% decease in class I, average of two
donors; 7% decrease in HLA-A2, average of two donors) (Fig. 3, B and C). Modulation of class I expression was
also assessed in the HepG2 cell line to investigate any potential
change after AdV transduction when these cells were used as
cytotoxicity target cells. HepG2 showed a decrease in total class I
(40%) or HLA-A2 (33%) after AdVMART1 transduction as well as a
decrease after AdVRR5 transduction (59% decrease in class I; 30%
decrease in HLA-A2). A control AdVLacZ transduction of HepG2
cells confirmed the percentage of cells transduced at the moi used
(>95%, data not shown).
|
MART-127–35 peptide pulsed PBMC were used to generate
peptide-specific CTL in vitro from normal donors. After 3–4 wk of
culture, these cells were tested for cytotoxic activity against a
variety of targets. As shown in Fig. 4,
these bulk cell populations (generally 30–40% CD8+ by
phenotypic analysis) demonstrate MART-127–35 peptide
specificity by lysis of MART-1 pulsed T2 cells in a 4-h chromium
release assay, but no lysis of Flu M1 peptide-pulsed T2. These cultures
also show specific killing of the MART-1+ M202 melanoma
cells, indicating that MART-127–35-specific CTL recognize
the endogenously processed and presented MART-1 epitope. The results of
the cytotoxicity data are also reflected by cytokine release data (not
shown). Both TNF- and IFN- were released in increased amounts by
CTL stimulated by T2 cells pulsed with the specific MART-1 peptide, not
by the control Flu peptide-pulsed cells.
|
B demonstrates that the transduction of HepG2
cells by AdVMART1 renders them sensitive to lysis by the
MART-127–35 CTL, while transduction with AdVRR5 does not. MART-127–35 TIL clone recognition of AdVMART1- transduced DC
A MART-127–35-specific melanoma TIL clone, generated
from an HLA-A2.1 melanoma patient was used to further demonstrate that
AdVMART1 transduction results in proper processing and presentation of
the immunodominant MART-127–35 peptide. The TIL clone
lysed MART-1 peptide-pulsed T2 cells and not control targets. The TIL
line also secreted TNF- upon recognition of MART-127–35
peptide-pulsed T2 cells and not a control peptide (MAGE 3)-pulsed T2
cells or T2 cells alone, confirming MART-127–35 peptide
specificity (Table I). Normal HLA-A2.1
donor DC were transduced with AdVMART1 or RR5 and cocultured with the
MART-127–35-specific TIL. The TIL line secreted IFN-
and TNF-ß specifically in response to DC transduced with AdVMART1,
and not DC transduced with the control AdVRR5 (Table II). This indicates that the
AdVMART1-transduced cells process and present the same
MART-127–35 peptide epitope present on the original
melanoma from the patient from which the TIL line was derived.
|
|
DC were generated from healthy HLA-A2.1 (donor 1) and A2.4 donors
(donor 2), transduced with AdVMART1, and used as APCs to generate
MART-1-specific CTL from CD8+ T cell-enriched PBMC.
Specific killing of A2+/MART-1+ melanoma cells
(M202) was detected in as little as 1 wk of culture (Table III). This cytotoxicity was increased
with IFN- treatment of the targets and partially inhibited by
anti-class I blocking Abs. The results of weekly assays over a 3-wk
culture period from one donor and the first assay from a second donor
are shown in Table III. Killing of either HLA-mismatched (A2-negative,
M201, or M207) or A2+/MART-1-negative melanoma cell lines
(M237 or M238) was not observed. Four of four A2+ normal
PBMC donors could generate M202-killing CTL within 1 wk of culture with
this DC/AdV-based culture system. Both HLA-A2.1 and A2.4 donors
generated similar responses against the T2 cell (not shown) and
M202 melanoma targets (Table III).
|
). Therefore, any
AdV Ag reactivity generated in these CTL cultures appears to be weak
and does not dampen MART-1-specific reactivity.
To demonstrate MART-127–35 reactivity, cocultures were set
up with total PBL from a normal donor and AdVMART1- or RR5-transduced
autologous DC. When tested for cytotoxicity on day 7 (Fig. 5A), the cocultures with
AdVMART1/DC generated MART-127–35 peptide-specific CTL.
The AdVMART1/DC-stimulated effector cells also synthesized TNF- upon
specific recognition of MART-127–35 peptide (Fig. 5B). The RR5/DC-stimulated PBMC showed only background
TNF- secretion to the targets tested.
Lastly, to further assess AdV Ag reactivity in the transduced DC CTL
cultures, RR5-transduced DC were cultured with
CD8+-enriched T cells. The RR5 AdV, which contains no
insert in the plasmid backbone, stimulated cultures that did produce
CTL reactive to RR5-transduced targets (Fig. 5C) compared
with untransduced targets.
|
Discussion |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
DC have been shown to be efficient cells for in vitro CTL generation in several reports (10, 29, 30, 31, 32, 33, 34, 35, 36, 37). CTL specific for several melanoma Ags (MART-1, gp100, tyrosinase) have been generated utilizing peptide-pulsed DC (10, 38). In addition, use of DC has uncovered subdominant epitopes from gp100 that were not detected utilizing other APC (31). Infection of DC with influenza virus was used to generate Flu-reactive CD8+ CTL in as few as 7 days (39). A MART-1-expressing retrovirus has been used to transduce CD34+ hemopoietic progenitor cells (40) which, after differentiation into DC, could successfully generate anti-MART-1 CTL. Vaccinia viral vectors have also been used as DC transduction vectors with melanoma tumor Ags. A tyrosinase-vaccinia viral vector (41) was used to transduce melanoma patient APC; both CD4+ and CD8+ tyrosinase-reactive T cells were generated after limiting dilution cloning. This viral vector was also used to successfully generate tyrosinase-specific CTL from two of four normal donors tested. A MART-1 vaccinia virus was used to transduce melanoma patient DC (42) and successfully generated CD8+ CTL that recognized MART-1 targets in 6 of 6 patients within 7–14 days of culture. Likewise, a MART-1 expressing AdV was reported (43) that sensitized targets cells to MART-1-specific lysis. These reports support the continued investigation of virally encoded tumor Ags for the genetic engineering of DC. These viruses have in common the potential to express a variety of known and unknown tumor Ag epitopes on the surface of DC and without HLA restriction. This would also potentially reduce the problem of Ag loss variants that could evade the immune response generated (44, 45). In addition, processing and presentation of peptides through endogenous routes may be more efficient for cell surface display than exogenous loading of synthetic peptides (46).
An important finding of this work was the demonstration that
MART-1-transduced DC generate CTL in vitro. We have not observed
significant AdV Ag reactivity in the CTL cultures generated with
AdVMART1-transduced DC, despite the high transduction efficiency with
this virus (Fig. 2B). This may be due to the low levels of
AdV gene transcription after removal of the E1
trans-activating region. There is, most likely, some low
level of AdV protein synthesis, given that the CTL generated from the
empty control AdV RR5 recognize AdV-transduced cells to a somewhat
greater degree than nontransduced cells (Fig. 5C). As a
potential improvement on this strategy, we are pursuing a
third-generation, helper-dependent, adenoviral vector that will have
all viral genes deleted (47, 48) to possibly improve the Ag-specific
reactivity, eliminate the low level of class I expression
down-regulation, and reduce viral gene immunogenicity.
|
Acknowledgments |
|---|
|
Footnotes |
|---|
2 Address correspondence and reprint requests to Dr. James S. Economou, Division of Surgical Oncology, 54-140 CHS, UCLA Medical Center, 10833 Le Conte Avenue, Los Angeles, CA 90095-1782. E-mail address:
3 Abbreviations used in this paper: TIL, tumor-infiltrating lymphocytes; DC, dendritic cell; MART-1, MART-1/Melan-A; AdV, adenovirus; AdVMART1, adenovirus encoding MART-1 cDNA; moi, multiplicity of infection; PSF, penicillin-streptomycin-Fungizone.
Received for publication March 10, 1998. Accepted for publication July 20, 1998.
|
References |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
This article has been cited by other articles:
|
|
 |
|
  Y. Liu, S. Daley, V. N. Evdokimova, D. D. Zdobinski, D. M. Potter, and L. H. Butterfield Hierarchy of {alpha} Fetoprotein (AFP)-Specific T Cell Responses in Subjects with AFP-Positive Hepatocellular Cancer J. Immunol., July 1, 2006; 177(1): 712 - 721. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 L. Chapatte, M. Ayyoub, S. Morel, A.-L. Peitrequin, N. Levy, C. Servis, B. J. Van den Eynde, D. Valmori, and F. Levy Processing of Tumor-Associated Antigen by the Proteasomes of Dendritic Cells Controls In vivo T-Cell Responses. Cancer Res., May 15, 2006; 66(10): 5461 - 5468. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 W. Li, J. Li, D. L. J. Tyrrell, and B. Agrawal Expression of hepatitis C virus-derived core or NS3 antigens in human dendritic cells leads to induction of pro-inflammatory cytokines and normal T-cell stimulation capabilities J. Gen. Virol., January 1, 2006; 87(1): 61 - 72. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
T. P. Moran, M. Collier, K. P. McKinnon, N. L. Davis, R. E. Johnston, and J. S. Serody A Novel Viral System for Generating Antigen-Specific T Cells J. Immunol., September 1, 2005; 175(5): 3431 - 3438. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
Y.-P. Liao, C.-C. Wang, L. H. Butterfield, J. S. Economou, A. Ribas, W. S. Meng, K. S. Iwamoto, and W. H. McBride Ionizing Radiation Affects Human MART-1 Melanoma Antigen Processing and Presentation by Dendritic Cells J. Immunol., August 15, 2004; 173(4): 2462 - 2469. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 H. Mukouyama, N. K. Janzen, J. M. Hernandez, J. S. Lam, R. Caliliw, A. Y. Wang, R. A. Figlin, A. S. Belldegrun, and G. Zeng Generation of Kidney Cancer-Specific Antitumor Immune Responses Using Peripheral Blood Monocytes Transduced With a Recombinant Adenovirus Encoding Carbonic Anhydrase 9 Clin. Cancer Res., February 15, 2004; 10(4): 1421 - 1429. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
A. Tuettenberg, H. Jonuleit, T. Tuting, J. Bruck, V. Biermann, S. Kochanek, J. Knop, and A. H. Enk Early Adenoviral Gene Expression Mediates Immunosuppression by Transduced Dendritic Cell (DC): Implications for Immunotherapy Using Genetically Modified DC J. Immunol., February 1, 2004; 172(3): 1524 - 1530. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
M. R. Parkhurst, C. DePan, J. P. Riley, S. A. Rosenberg, and S. Shu Hybrids of Dendritic Cells and Tumor Cells Generated by Electrofusion Simultaneously Present Immunodominant Epitopes from Multiple Human Tumor-Associated Antigens in the Context of MHC Class I and Class II Molecules J. Immunol., May 15, 2003; 170(10): 5317 - 5325. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
J. M. Hernandez, M. H. T. Bui, K.-r. Han, H. Mukouyama, D. G. Freitas, D. Nguyen, R. Caliliw, P. I. Shintaku, S. H. Paik, C.-L. Tso, et al. Novel Kidney Cancer Immunotherapy Based on the Granulocyte- Macrophage Colony-stimulating Factor and Carbonic Anhydrase IX Fusion Gene Clin. Cancer Res., May 1, 2003; 9(5): 1906 - 1916. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
L. H. Butterfield, A. Ribas, V. B. Dissette, S. N. Amarnani, H. T. Vu, D. Oseguera, H.-J. Wang, R. M. Elashoff, W. H. McBride, B. Mukherji, et al. Determinant Spreading Associated with Clinical Response in Dendritic Cell-based Immunotherapy for Malignant Melanoma Clin. Cancer Res., March 1, 2003; 9(3): 998 - 1008. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 A. Burchert, S. Wolfl, M. Schmidt, C. Brendel, B. Denecke, D. Cai, L. Odyvanova, T. Lahaye, M. C. Muller, T. Berg, et al. Interferon-alpha , but not the ABL-kinase inhibitor imatinib (STI571), induces expression of myeloblastin and a specific T-cell response in chronic myeloid leukemia Blood, January 1, 2003; 101(1): 259 - 264. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
Y. Ikuta, N. Katayama, L. Wang, T. Okugawa, Y. Takahashi, M. Schmitt, X. Gu, M. Watanabe, K. Akiyoshi, H. Nakamura, et al. Presentation of a major histocompatibility complex class 1-binding peptide by monocyte-derived dendritic cells incorporating hydrophobized polysaccharide-truncated HER2 protein complex: implications for a polyvalent immuno-cell therapy Blood, May 15, 2002; 99(10): 3717 - 3724. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
A. Ribas, L. H. Butterfield, S. N. Amarnani, V. B. Dissette, D. Kim, W. S. Meng, G. A. Miranda, H.-J. Wang, W. H. McBride, J. A. Glaspy, et al. CD40 Cross-Linking Bypasses the Absolute Requirement for CD4 T Cells during Immunization with Melanoma Antigen Gene-modified Dendritic Cells Cancer Res., December 1, 2001; 61(24): 8787 - 8793. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
C. Meyer zum Buschenfelde, J. Metzger, C. Hermann, N. Nicklisch, C. Peschel, and H. Bernhard The Generation of Both T Killer and Th Cell Clones Specific for the Tumor-Associated Antigen HER2 Using Retrovirally Transduced Dendritic Cells J. Immunol., August 1, 2001; 167(3): 1712 - 1719. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
D. Rea, M. J. E. Havenga, M. van den Assem, R. P. M. Sutmuller, A. Lemckert, R. C. Hoeben, A. Bout, C. J. M. Melief, and R. Offringa Highly Efficient Transduction of Human Monocyte-Derived Dendritic Cells with Subgroup B Fiber-Modified Adenovirus Vectors Enhances Transgene-Encoded Antigen Presentation to Cytotoxic T Cells J. Immunol., April 15, 2001; 166(8): 5236 - 5244. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
L. H. Butterfield, W. S. Meng, A. Koh, C. M. Vollmer, A. Ribas, V. B. Dissette, K. Faull, J. A. Glaspy, W. H. McBride, and J. S. Economou T Cell Responses to HLA-A*0201-Restricted Peptides Derived from Human {{alpha}} Fetoprotein J. Immunol., April 15, 2001; 166(8): 5300 - 5308. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
C. J. Kirk, D. Hartigan-O’Connor, B. J. Nickoloff, J. S. Chamberlain, M. Giedlin, L. Aukerman, and J. J. Mulé T Cell-dependent Antitumor Immunity Mediated by Secondary Lymphoid Tissue Chemokine: Augmentation of Dendritic Cell-based Immunotherapy Cancer Res., March 1, 2001; 61(5): 2062 - 2070. [Abstract] [Full Text]
|
|
|
|
|
|
P. J. Frost, L. H. Butterfield, V. B. Dissette, J. S. Economou, and B. Bonavida Immunosensitization of Melanoma Tumor Cells to Non-MHC Fas-Mediated Killing by MART-1-Specific CTL Cultures J. Immunol., March 1, 2001; 166(5): 3564 - 3573. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
M. V. Dhodapkar, J. W. Young, P. B. Chapman, W. I. Cox, J. F. Fonteneau, S. Amigorena, A. N. Houghton, R. M. Steinman, and N. Bhardwaj Paucity of Functional T-Cell Memory to Melanoma Antigens in Healthy Donors and Melanoma Patients Clin. Cancer Res., December 1, 2000; 6(12): 4831 - 4838. [Abstract] [Full Text]
|
|
|
|
|
|
B. W. Anderson, G. E. Peoples, J. L. Murray, M. A. Gillogly, D. M. Gershenson, and C. G. Ioannides Peptide Priming of Cytolytic Activity to HER-2 Epitope 369-377 in Healthy Individuals Clin. Cancer Res., November 1, 2000; 6(11): 4192 - 4200. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
C. Meyer zum Buschenfelde, N. Nicklisch, S. Rose-John, C. Peschel, and H. Bernhard Generation of Tumor-Reactive CTL Against the Tumor-Associated Antigen HER2 Using Retrovirally Transduced Dendritic Cells Derived from CD34+ Hemopoietic Progenitor Cells J. Immunol., October 1, 2000; 165(7): 4133 - 4140. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 S. Gnjatic, Y. Nagata, E. Jager, E. Stockert, S. Shankara, B. L. Roberts, G. P. Mazzara, S. Y. Lee, P. R. Dunbar, B. Dupont, et al. Strategy for monitoring T cell responses to NY-ESO-1 in patients with any HLA class I allele PNAS, September 26, 2000; 97(20): 10917 - 10922. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
A. Ribas, L. H. Butterfield, B. Hu, V. B. Dissette, W. S. Meng, A. Koh, K. J. Andrews, M. Lee, S. N. Amar, J. A. Glaspy, et al. Immune Deviation and Fas-mediated Deletion Limit Antitumor Activity after Multiple Dendritic Cell Vaccinations in Mice Cancer Res., April 1, 2000; 60(8): 2218 - 2224. [Abstract] [Full Text]
|
|
|
|
 |
|
 A. Ribas, L. H. Butterfield, and J. S. Economou Genetic Immunotherapy for Cancer Oncologist, April 1, 2000; 5(2): 87 - 98. [Abstract] [Full Text]
|
|
|
|
|
|
G. P. Linette, S. Shankara, S. Longerich, S. Yang, R. Doll, C. Nicolette, F. I. Preffer, B. L. Roberts, and F. G. Haluska In Vitro Priming with Adenovirus/gp100 Antigen-Transduced Dendritic Cells Reveals the Epitope Specificity of HLA-A*0201-Restricted CD8+ T Cells in Patients with Melanoma J. Immunol., March 15, 2000; 164(6): 3402 - 3412. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 D. Rea, F. H. E. Schagen, R. C. Hoeben, M. Mehtali, M. J. E. Havenga, R. E. M. Toes, C. J. M. Melief, and R. Offringa Adenoviruses Activate Human Dendritic Cells without Polarization toward a T-Helper Type 1-Inducing Subset J. Virol., December 1, 1999; 73(12): 10245 - 10253. [Abstract] [Full Text]
|
|
|
|
|
|
E. Ranieri, W. Herr, A. Gambotto, W. Olson, D. Rowe, P. D. Robbins, L. S. Kierstead, S. C. Watkins, L. Gesualdo, and W. J. Storkus Dendritic Cells Transduced with an Adenovirus Vector Encoding Epstein-Barr Virus Latent Membrane Protein 2B: a New Modality for Vaccination J. Virol., December 1, 1999; 73(12): 10416 - 10425. [Abstract] [Full Text]
|
|
|
|
|
|
C. M. Vollmer Jr., F. C. Eilber, L. H. Butterfield, A. Ribas, V. B. Dissette, A. Koh, L. D. Montejo, M. C. Lee, K. J. Andrews, W. H. McBride, et al. {{alpha}}-Fetoprotein-specific Genetic Immunotherapy for Hepatocellular Carcinoma Cancer Res., July 1, 1999; 59(13): 3064 - 3067. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
L. H. Butterfield, A. Koh, W. Meng, C. M. Vollmer, A. Ribas, V. Dissette, E. Lee, J. A. Glaspy, W. H. McBride, and J. S. Economou Generation of Human T-cell Responses to an HLA-A2.1-restricted Peptide Epitope Derived from {{alpha}}-Fetoprotein Cancer Res., July 1, 1999; 59(13): 3134 - 3142. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
K. R. Prilliman, K. W. Jackson, M. Lindsey, J. Wang, D. Crawford, and W. H. Hildebrand HLA-B15 Peptide Ligands Are Preferentially Anchored at Their C Termini J. Immunol., June 15, 1999; 162(12): 7277 - 7284. [Abstract] [Full Text] [PDF]
|
|
| |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
