and Immunomodulatory xenobiotics)
.For modulation of Th cell responses, peptides offere several advantages over intact Ags as immunogens or tolerogens (either as altered peptide ligands (APLs), competitors, or vaccines). First, peptides require less stringent degradative conditions than native Ags. Second, with a smaller determinant, there is less likelihood of cross-reactivity between the peptide and other self-proteins. And third, peptides offer exquisite specificity over native Ags. Despite these advantages, the use of peptides has remained fairly limited, because they are rapidly cleared from the circulation and poorly taken-up by APCs (by pinocytosis only) : effectiveness of taking up can be increased by coupling peptides to ligands (eg. transferrin (Tf)) specific for cell surface receptors found on APCs (eg. CD71 / TfR).
)
are used for chronic infections and cancers
vaccine : if given within 4 days of exposure to virus, it halts the disease's
advance.
vaccine : intramuscular (2.5 IU
= 1 mL) or intradermic (0.1 mL) injections of at day 0, 3, 7, 14,
28 and 90. Effective for 2-3 years. The treatment
of rabies exposure has come a long way since the time a person had to take
21 shots in the stomach. Today, most people need 5 rabies vaccinations
in the arm spaced out over a 28-day period, as well as a one-time dose
of rabies immune
globulin (RIG).
The WHO recommends 2 economical intradermal regimens for post-exposure
rabies prophylaxis (PEP) :;
purified
chick embryo cell vaccine (PCECV)
and purified
duck embryo vaccine (PDEV).
The other recommended vaccine, purified
vero cell rabies vaccine (PVRV),
has only 0.5 ml per ampoule and it has not been tested with the 8-site
regimen, but if it were used the logical dose would be 0.05 ml per site,
or alternatively 0.1 ml in half the number of sites. (A whole ampoule of
vaccine must be used on day 0 in every case). The 2-site id regimen was
designed for use with PVRV. It requires a dose of 0.1 ml per site for PVRV
and 0.2 ml per site for the other 3 vaccines. Full details
of the economical regimens and practical advice are availableon the
WHO website. The advantages of the 8-site method are a rapid induction
of neutralising antibody, making it the treatment
of choice when no rabies immunoglobulin is available (>95% of post-exposure
treatments in the third world); a wide margin of safety, because if up
to half of the 8 id injections on day 0 are not truly id, the immune response
will still be adequate; and using a wholeampoule on day 0, which avoids
the need to share ampoules of vaccine between patients during the emergency
treatment and eliminates any risk of contamination or wastage of vaccine.
Finally, giving a large antigenic stimulus on day 0 gives the best chance
of survival to patients who are 'low responders' to the vaccine and to
those who fail to return on time for subsequent doses, a common problem
in the rural tropics. The complexity of these treatments demonstrates the
urgent need for animproved, single, simplified regimen that is suitable
for use with all the WHO recommended vaccines. Efforts are being made to
achieve this goal. Meanwhile there is reluctance to use these vaccines
economically. This deprives many patients of excellent treatment and perpetuates
double standards of treatment in the third
world. Under current CDC guidelines, post-exposure prophylaxis for
persons not pre-exposure vaccinated currently consists of a
vaccine : V-1
Immunitor (V1) (Remune©; Immune
Response Corp., Carlsbad, CA, USA, and Trinity Medical Group Co., Ltd.
of Bangkok, Thailand) is a low-cost, polyvalent
therapeutic
mucosal
AIDS
vaccine against core antigens based on the Salk Immunogen, which is derived
from an HIV isolate which has been inactivated by chemical depletion of
gp120 (clade A/G) : it is composed of an HIV-1 isolate (HZ321) from serum
collected from a patient in Zaire in 1976. This virus has been sequenced
and classified as clade A envelope and clade G Gag and grown in the Hut
78 T-cell lineref1,
ref2,
ref3.
It was developed and manufactured in Thailand and has been licensed
by the Thai FDA as a orally available dietary
supplement and investigational R&D drug. It has
shown effectiveness against wasting syndromeref1,
ref2,
ref3.
A phase II double-blind controlled clinical trial using HIV-1 immunogen
was undertaken in Thailand in 1995 in conjunction with phase III approval
in the USA. In most instances, immune response to the virus was induced.
The results from this study led to the extension of further trials over
the following 4 years. At present, the Thai Food and Drug Administration
(FDA) is in the process of reviewing the files on all information to assess
the effectiveness of this therapeutic vaccine as well as reviewing the
proposal to conduct a phase III trial to further confirm previous efficacy
results from the phase II trialsref.
vaccine : the incidence and severity of
herpes zoster and postherpetic neuralgia increase with age in association
with a progressive decline in cell-mediated immunity to VZV. 38,546 adults
> 60 years of age were enrolled in a randomized, double-blind, placebo-controlled
trial of an investigational live attenuated Oka/Merck VZV vaccine ("zoster
vaccine"). Herpes zoster was diagnosed according to clinical and laboratory
criteria. The pain and discomfort associated with herpes zoster were measured
repeatedly for 6 months. The primary end point was the burden of illness
due to herpes zoster, a measure affected by the incidence, severity, and
duration of the associated pain and discomfort. The secondary end point
was the incidence of postherpetic neuralgia. > 95% of the subjects continued
in the study to its completion, with a median of 3.12 years of surveillance
for herpes zoster. A total of 957 confirmed cases of herpes zoster (315
among vaccine recipients and 642 among placebo recipients) and 107 cases
of postherpetic neuralgia (27 among vaccine recipients and 80 among placebo
recipients) were included in the efficacy analysis. The use of the zoster
vaccine reduced the burden of illness due to herpes zoster by 61.1%, reduced
the incidence of postherpetic neuralgia by 66.5% (P<0.001), and reduced
the incidence of herpes zoster by 51.3%. Reactions at the injection site
were more frequent among vaccine recipients but were generally mild. The
zoster vaccine markedly reduced morbidity from herpes zoster and postherpetic
neuralgia among older adults.)
: on the basis of the successes of attenuated pathogen vaccines and owing
to the initial lack of defined tumour antigens, the first cancer vaccines
were composed of whole cancer cells removed during surgery, killed by irradiation
and re-administered together with non-specific adjuvants.
(modified tumour cells / transfectoma)
: hapten-treated autologous melanoma cells,
then killed by irradiation and administered intradermallyref
(good responses)ref1,
ref2,
ref3,
ref4
(good responses)ref1,
ref2ref1,
ref2ref
in patients with recurrent prostate
adenocarcinoma.
A single-institution phase I/II trial was done in hormone therapy-naive
patients with PSA relapse following radical prostatectomy and absence of
radiologic metastases. Treatments were administered weekly via intradermal
injections of 1.2 x 108 GM-CSF gene-transduced, irradiated,
cancer cells (6 x 107 LNCaP cells and 6 x 107 PC-3
cells) for 8 weeks. 21 patients were enrolled and treated. Toxicities included
local injection-site reactions, pruritus, and flu-like symptoms. 1 patient
had a partial PSA response of 7-month duration. At 20 weeks post first
treatment, 16 of 21 (76%) patients showed a statistically significant decrease
in PSA velocity (slope) compared with prevaccination (P < 0.001). Injection
site biopsies showed intradermal infiltrates consisting of CD1a+
dendritic cells and CD68+ macrophages, similar to previous clinical
trials using autologous GM-CSF-transduced cancer cells. Posttreatment,
patients developed new oligoclonal antibodies reactive against at least
five identified antigens present in LNCaP or PC-3 cells. A high-titer antibody
response against a 250-kDa antigen expressed on normal prostate epithelial
cells was induced in a patient with partial PSA remission; titers of this
antibody decreased when treatment ended, and subsequent PSA relapse occurredrefref1,
ref2
is an allogeneic melanoma tumor cell lysate combined with the adjuvant
Detox-PC, often in regimens containing pretreatment with low-dose cyclophosphamide
and IFN-a2bref1,
ref2
: phase I and II trials in stage IV patients showed a 10-20% response rate
(clearing of some metastatic sites) and in another 10-20% of patients disease
was stabilized (no progression for various periods of time of tumours that
were growing at the start of the vaccine protocol. It may have its greatest
benefit in a large subset of melanoma patients who express either the HLA-A2
and/or HLA-C3ref.
and colon cancer : irradiated allogeneic cells induce IgG and IgM immune
responses to glycoprotein melanoma-associated antigen (TA90) after surgical
resection, but only the IgM response is correlated with improved survival
: endogenous immune responses to TA90 have also been reportedref.
It is a polyvalent vaccine (PV) containing > 20 tumor antigens. In vitro
cellular immune response to Canvaxin cells directly correlates with survival
after subsequent initiation of immunotherapy for AJCC stage III melanomaref.
Canvaxin is currently undergoing a multi-centre phase III randomized clinical
trials for melanoma and a phase II trial for colorectal
carcinoma.
and colorectal
carcinomas.
as adjuvants would activate DCs to prime immunity to many autoantigens
(otherwise subject to peripheral tolerance) other than the tumor antigens
: the use of whole tumour cells or complex mixtures o tumour-derived material
undermines one unique advantage that immunotherapy has over other forms
of therapy - that is, specificity. Recently, in spite of the availability
of well-defined tumour antigens, development in the cancer-vaccine field
has focused again on the use of whole tumour cells or whole cell lysates
as antigens : the reason being that these complex mixtures will contain
unique
tumour antigens that are expressed only by an individual tumour
that, by analogy to unique antigens of mouse carcinogen-induced tumours
might be more immunogenic and promote a better anti-tumour immune response.
But experiments carried out in mice transgenic for shared
tumour antigenshave shown that these antigens can elicit equally
strong antitumour immunity and tumour rejection.
vaccine : conventional envelope-based antibody-inducing vaccines
do not appear to hold promise, and broadly-neutralizing antibodies are
now being searched as an alternative to the failed approach with subunit
vaccines. The current consensus is that cellular immune responses, especially
those mediated by CD8+ CTL and CD4+ Th
lymphocytes, are needed to control HIV. Vaccines capable of inducing cell-mediated
responses are, therefore, considered critical for controlling the spread
of HIV. DNA-based vaccines triggering CTL reaction are currently thought
to be an answer, but will they fulfill the promise?
vaccine : P. insidiosum-antigen
(PIA) 100-200 ml of PIA (2.0 mg/ml), at a 14-day
intervalref1,
ref2,
ref3-antigen
therapeutic vaccines (see also immunooncology). |
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has developed as a powerful technique to select ligands to essentially
any chosen target from diverse repertoires of peptides, proteins, or Abs
displayed on the surface of a phage particle. Previous application has
mainly focused on Ab libraries and constrained or linear peptide libraries.
This technology has also been used to identify immunogenic targets or epitopes
recognized by Abs; for example, selection of peptide libraries on mAbs
and complex sera of patients with disease has led to the isolation of immunoreactive
peptide epitopes. Although mapping sera with peptide phage libraries has
had some success with small viral genomes such as HBV,
the selection of peptide repertoires appears not to be a suitable approach
when profiling the complexity of the immune response in the context of
the human genome. One of the drawbacks is that often predominantly Ag
mimotopes are recovered, for which further complicated analysis is
required to recover the original Ag that elicited the immune response.
A more successful approach is to display cDNA expression libraries on filamentous
phage. In contrast to the use of peptide display repertoires, there have
been significantly fewer reports on cDNA display due to technical challenges
in their construction and expression. For example, due to stop codons inherent
to cDNA, cDNA display libraries cannot be fused to the N terminus of the
popular phage anchor protein pIII. As such, anchoring of the cDNA product
on the phage coat has to be performed via indirect linkage to pIII or by
direct fusion to the C terminus of another phage coat protein, pVI. In
the case of low titer IgG response that predominate in immunosuppressed
conditions such as cancer, early studies using autologous selection of
phage-displayed primary tumor cDNA repertoires were unsuccessful, although
selections with both mAbs and polyclonal rabbit sera did recover specific
ligands. Selections performed with immobilized autologous cancer patient
IgG using and anti-human IgG capture Ab resulted in the preferential recovery
of only IgG transcripts that were present due to tumor-infiltrating B cells.-associated
tumors provide target Ags for immunotherapy,
lung,
bladder,
ovarian
carcinomas,
hepatocellular
carcinoma (HCC)
(MAGE-1, SSX-1, CTp11 and HCA587 => polyvalent vaccinationsref),
breast
carcinomas
and multiple myeloma.
T-cell immune responses to CGAg have been identified in patients with solid
tumors and multiple myelomaref.
They do not belong to the immunological self because they are expressed
only in immunoprivileged
sites.
Since germ line cells do not carry HLA molecules on their surface they
cannot present antigens to T cells. Their immunogenicity can be enhanced
if they are expressed (together with both allogeneic and (transfected)
syngeneic and MHC determinants) by highly antigenic (e.g. non-self)
cells after DNA transfection.
Supporting the role of DNA methylation in generating the intratumor heterogeneity
of CTA, the DNA hypomethylating agent 5-aza-2'-deoxycytidine (5-AZA-dCyd)
invariably induced their expression in all CTA-negative clones of human
cutaneous melanomaref)
: a MAGE-3 antigen presented by HLA-A1 has been used in several vaccination
trials on metastatic melanoma patients. Only a small minority of patients
have shown evidence of tumor regression. Attempts to correlate the tumor
rejections with the cytotoxic T lymphocyte (CTL) response against the vaccine
have been hampered by the low level of these responses. In noncancerous
individuals, the frequency of the T cell precursors against antigen MAGE-3.A1
is approximately 4 x 10-7 CD8 T cells. The diversity of the
TcR repertoire of these anti-MAGE-3.A1 precursors was analyzed in one individual.
The results indicate that it is very likely that the repertoire comprises
>100 clonotypes. On this basis, it is possible to use not only the frequency
of CTL precursors in the blood but also the presence of dominant clonotypes
to ascertain in patients the existence of anti-MAGE-3.A1 responses as low
as 10-6 of CD8. With this approach, we observed a correlation
between tumor regression and anti-MAGE-3.A1 CTL responses in patients vaccinated
with a recombinant virus encoding the antigen and also in patients vaccinated
with peptide-pulsed dendritic cells. In contrast, for patients showing
tumor regression after vaccination with peptide alone, CTL responses were
almost never observed. It is possible that even those CTL responses that
are below our present detection level can trigger a sequence of events
that leads to tumor regressionref.
MAGE-A3 peptides presented by HLA class I molecules have been identified
using CD8 lymphocytes stimulated with cells that either expressed gene
MAGE-A3 or were pulsed with candidate peptides. One antigen identified
with the latter method is peptide MAGE-A3195-203 IMPKAGLLI,
presented by HLA-A24 molecules. It has been used to vaccinate advanced
cancer patients. HLA/peptide tetramers were used to detect T cells recognizing
this peptide. Their frequency was estimated to be 2 x 10-8 of
the blood CD8 cells in non-cancerous HLA-A24+ individuals, which
is 10-fold lower than the reported frequencies of T cells against other
MAGE peptides. In the blood of a patient vaccinated with MAGE-A3, the estimated
frequency was 5 x 10-7. Anti-MAGE-3.A24 cytolytic T cell clones
were derived, that lysed peptide-pulsed cells with half-maximal effect
at the low concentration of 500 pM. However, these CTL did not recognize
a panel of HLA-A24+ tumor cells that expressed MAGE-A3 at levels
similar to those found in HLA-A1+ tumor cells recognized by
anti-MAGE-3.A1 CTLs. Furthermore, 293-EBNA cells transfected with MAGE-A3
and HLA-A24 constructs were hardly recognized by the anti-MAGE-3.A24 CTL
clones. These results suggest that peptide MAGE-A3195-203 is
poorly processed and is not an appropriate target for cancer immunotherapyref.,
lung,
bladder,
and head and neck
cancers)
: the peptide NYKRCFPVI, which corresponds to amino acids 143 to151 of
the MAGE-4 protein, can be presented by HLA-A24 molecules, which are widely
expressed in different ethnic groups. A cytolytic T cell clone that recognized
the MAGE-4 peptide was isolated from the blood cells of a donor without
cancer and lysed specifically A24 carcinoma cells expressing MAGE-4. The
antigenic peptide is processed more efficiently in tumor cells pre-treated
with IFN-gref.
respectively. Both CT antigens were more frequently expressed in squamous
cell carcinoma (SCC) than in adenocarcinoma. An inverse correlation was
found between MAGE-A4 expression and patient survival in advanced stage
cancers. Combined infiltration of both CD4+ and CD8+ T cells into tumor
nest predicted better survival. There was no correlation, however, between
lymphocyte infiltration and antigen expression in the tumor. MAGE-A4 expression
in advanced group and T cell infiltration may provide prognostic information.
Lastly, these CT antigens, especially MAGE-A4, may represent potential
targets for cancer immunotherapy in patients with NSCLCref.
and acute myeloid
leukemias)ref)
is a highly immunogenic protein and the observation that > 90% of vasectomized
males develop immunity against Sp17 suggests the opportunity and safety
of Sp17 for tumor vaccines. Although Sp17 transcripts could be detected
by RT-PCR in some normal tissues, the levels of expression were <2%
of those in normal testis. In contrast, Sp17+ myeloma cells
expressed 3-18% of normal testis levels of Sp17 transcript. IHC using 2
Sp17 murine mAbs, each directed at a non-overlapping B-cell epitope, showed
Sp17 protein to be expressed only in testis and not any other normal tissues.
Specificity of binding of the antibodies to testis was also confirmed in
competitive binding assaysref.
Recent works by other workers suggest a low level of expression of Sp17
in some normal tissues, and investigators have questioned whether Sp17
is in fact a suitable target for immunotherapyref.
Sp17-specific HLA class I-restricted CTL generation was successful in all
4 patients, irrespective of the Sp17 status of their tumors : most importantly,
the CTLs were able to lyse autologous tumor cells that expressed Sp17ref.
Sp17-specific HLA class I-restricted CTLs can also be easily generated
from the peripheral blood of healthy donor. Because CTLs are able to kill
HLA-matched fresh myeloma cells, it may be possible to generate and administer
myeloma-specific donor T cells to MM patients following allogeneic stem
cell transplantation to enhance graft-versus-myeloma (GVM) without inducing
graft-versus-host disease (GVHD)ref1,
ref2.
Sp17 was found to be expressed in the primary tumor cells from 70% of the
patients with ovarian
carcinoma.
HLA class I-restricted Sp17 specific CTLs were generated successfully from
the peripheral blood of 3 patients with ovarian carcinoma at the time of
disease presentation. These CTLs were able to lyse autologous EBV-transformed
lymphoblastoid cells in a Sp17-dependent manner. Target lysis was HLA class
I-dependent and could be blocked by antibodies against monomorphic HLA
class I but not HLA class II molecules. The CTLs also lysed Sp17+
autologous tumor cells, suggesting that Sp17 is processed and presented
in association with the HLA class I molecules in Sp17+ tumor
cells in a concentration and configuration that could be recognized by
recombinant protein-primed CTLs. Tumor cell killing by the CTLs appeared
to be mediated through the perforin pathwayref,
3 of 11 chronic
lymphocytic leukemia (CLL),
4 of 14 chronic
myeloid leukemia,
and 6 of 17 multiple myeloma.
In contrast, they were not detected in corresponding specimens from any
healthy donors. SLLP1 exhibited a very restricted normal tissue expression,
being found only in testis/spermatozoa. SLLP1 was expressed in some tumor
cells at a level of >25%. High titer IgG antibodies against SLLP1 were
also detected in the sera of some of these patients. CONCLUSIONS: SLLP1
is a novel cancer-testis antigen in hematologic malignancies and is capable
of eliciting B-cell immune responses in vivo in cancer-bearing individualsref
and other hematologic malignancies such as chronic
lymphocytic leukemia (CLL),
chronic
myeloid leukemia,
and acute myeloid
leukemia.
It is not expressed in any normal tissue except testis. High-titer IgG3
and IgG2 against SPAN-Xb were detectable in the sera of
these patients. In contrast, SPAN-Xb mRNA or antibodies could not be detected
in any of the healthy donors. There was a good correlation between SPAN-Xb
gene expression and B-cell immune responses. These results suggest the
in vivo immunogenicity of the SPAN-Xb protein. The presence of high-titer
IgG responses suggests that the B-cell responses are likely to have been
generated with CD4 T-cell cognitive helpref)ref
have antibody responses to SCP-1, but not to other cancer testis antigens
(GAGE, LAGE-1a,-1b, CT-7, NY-ESO-1, SSX-1-5). Antibody response correlated
with the expression of SCP-1 in the primary tumor of the respective patient
as shown by RT-PCR, IHC and Western blot. In contrast, no serological response
to CTAgs was observed in healthy donors. The humoral immune response against
SCP-1 was associated with the size of tumor, but not with other clinico-pathological
parameters such as histology, stage, presence of lymph node metastases,
grading, age, gender or gemcitabine treatmentref
is highly expressed in 80% of hepatocellular
carcinoma (HCC)ref1,
ref2,
ref3,
ref4,
ref5,
but AFP immunization alone still resulted in lower levels of specific response
and poorly reproducible protective immunityref1,
ref2,
ref3,
ref4,
ref5
: the immunogenicity of AFP could be improved by presenting to APCs through
HSP70 moleculesref1,
ref2,
ref3,
ref4,
ref5.
Sequential immunization by i.m. injection of a recombinant DNA plasmid
vaccine encoding AFP and HSP70 fragments could generate effective AFP-specific
T cell responses and induce definite antitumor effects on AFP-producing
tumors, which may be suitable for some clinical testing as a vaccine for
HCCref.
The recombinant expression vectors PEGF-N3/AFP1 and PEGF-N3/AFP2 with or
without signal peptide were constructed successfully. AFP2/DCs can specifically
activate T lymphocytes in vitro and effectively induce antitumor
immune responseref.
: C57BL/6 and BALB/c mice were immunized with 2 different plasmids (p91023B
and pKCEA66) encoding different forms of the TAA CEA. The wild type form
of CEA (p91023B), which is expressed at the cell surface, induces stronger
anti-CEA IgG response after DNA-plasmid immunizations than the modified
intracellular form of CEA (pKCEA66), which was designed to mount strong
cellular responses. Boosting with recombinant CEA (rCEA) increased the
anti-CEA IgG response significantly. In the tumor protection model used,
where SCID mice are challenged with human tumor cells mixed with splenocytes
from immunized mice, both innate and specific immune responses are responsible
for the protective effectref.
Exosomes from heat-stressed CEA+ tumor cells (CEA+/HS-Exo)
contained CEA and more HSP70 and MHC-I and significantly induced dendritic
cell maturation. Immunization of HLA-A2.1/K(b) transgenic mice with CEA+/HS-Exo
was more efficient in priming a CEA-specific CTL, and the CTL showed antitumor
effect when adoptively transferred to SW480-bearing nude mice. Moreover,
in
vitro incubation of lymphocytes from HLA-A*0201+ healthy
donors and HLA-A*0201+CEA+ cancer patients with CEA+/HS-Exo-pulsed
autologous dendritic cells induces HLA-A*0201-restricted and CEA-specific
CTL response. CEA+/HS-Exo has superior immunogenicity than CEA+/Exo
in inducing CEA-specific CTL responseref.
CEA, the most widely used human tumor marker, is a heavily glycosylated
protein over-expressed by a wide range of tumors. It has been indicated
that CEA might be a useful target for human anti-tumor immunotherapy. CEA
assay for research as well as clinical trials demands a continuous source
of CEA protein preparations. In a multi-purpose research program to provide
a reliable source for large production of CEA, the membrane-bound carcinoembryonic
antigen was converted into a secretory protein by site-specific mutagenesis.
The secretory CEA protein was made by introducing a new stop codon at 99
bp upstream of the original stop codon in CEA cDNA by PCR-based mutagenesis.
The glycosylation of recombinant CEA proteins, especially those destined
for administration to human trials is crucially important. To produce CEA
with the same glycosylation pattern and immunogenicity as the native CEA
expressed by human tumors in vivo, the truncated CEA cDNA which
does not encode the last C-terminal 33-amino acid hydrophobic domain was
transfected into HT29, a human colon carcinoma cell line by the calcium
phosphate method. Stable transfectants were selected and pooled. CEA secretion
from the cells was verified by analysis of the transfectant culture supernatant
for CEA protein. As determined by ELISA, 16 mg/L
of recombinant CEA was secreted per 106 transfectants within
48 hrs, an increase over 40 times relative to the untransfected cells.
The size of the recombinant CEA secreted by HT29 transfectants in our experiment
is identical to that of reference CEA secreted from tumors and is fully
antigenic. It seems that the C-terminal truncation does not affect CEA
glycosylation in HT29 cells. It is predicted that human cancer immunotherapy
using recombinant CEA expressed in this system would be more effective
than the commercial protein which is usually prepared from bacterial or
other heterologous expression systemsref.
The existence of a murine model transgenic for human CEA allows CEA vaccination
efficacy to be studied in a physiologically tolerant context. Immunization
of CEA-transgenic mice with an adenoviral vector coding for CEA induced
a significant CD8+ T-cell response specific to CEA but failed
to induce CEA-specific CD4+ T cells and antibodies. To overcome
CD4+ T-cell tolerance, the effect of adjuvants inducing in
vivo DC maturation were explored. 2 different TLR ligands, monophosphoryl
lipid A (MPL) and CpG motif-containing oligodeoxynucleotides (CpG-ODN),
were tested. CD4+-mediated IFN-g
production was induced in the CEA-transgenic mice only when the genetic
immunization was performed in the presence of these adjuvants. Moreover,
CpG-ODN had a greater effect than MPL in inducing CD4+ T-cell
response and enabling anti-CEA antibody productionref.
in juvenile
myelomonocytic leukemia
are, again, unfolded. Characteristically, expression follows the same tissue
presentation as during embryogenesis. Clinical paternal disomies, i.e.
trophoblastic
diseases,
and their maternal counterpart, i.e. ovarian
teratomas,
are associated with apparent relaxation of imprinting once they turn malignant.
Paediatric neoplasms, like Wilms'
tumor (WT)
and rhabdomyosarcoma,
often express IGF2 and H19. Recently H19 expression has been found in invasive
urothelial cancer. Evidently, imprinted genes display an oncodevelopmental
mode of expression, very much like the classical oncofetal proteins AFP
and CEA.
for adenocarcinomas (certain breast
carcinomas,
...) is overexpressed and modified by tumor cells in > 50% of all cancer
cases. Animal data show that it can be a target for immune-mediated tumor
rejection, and finally, preliminary clinical results show that vaccine-based
immunotherapy with MUC1 does have an impact on the therapy of cancerref.
BLP25
liposome (L-BLP25®) vaccine (source : Biomira)
is a cancer vaccine that targets the exposed core peptide of the MUC1 tumor-associated
antigen. Studies in advanced-stage NSCLC
demonstrate that L-BLP25 vaccine has the potential to extend the survival
of patients with Stage IIIB locoregional NSCLC and maintain quality of
life for longer : it is now undergoing phase III clinical trial. L-BLP25
vaccine also shows promise for prostate
cancer
patients, having the potential to prolong PSA doubling time in men with
biochemical failure post prostatectomy. These clinically meaningful results
with a relatively nontoxic therapeutic vaccine are very encouraging and
suggest potential for L-BLP25 to fulfill an unmet medical needref.
patients received Tn(c)-KLH vaccine containing either 3, 7, or 15
mg
of Tn(c) per vaccination. 10 patients received 100 mg
of Tn(c)-palmitic acid (PAM). QS21 was included in all vaccines. 5 vaccinations
were administered subcutaneously during 26 weeks with an additional booster
vaccine at week 50. Tn(c), when given with the carrier molecule KLH and
QS21, stimulated the production of high-titer IgM and IgG antibodies. Inferior
antibody responses were seen with T(c)-PAM. There was no evidence of enhanced
immunogenicity with increasing doses of vaccine. An antitumor effect in
the form of a decline in posttreatment versus pretreatment PSA slopes was
also observed. A safe synthetic conjugate vaccine in a trimer formation
was developed that can break immunologic tolerance by inducing specific
humoral responses. It seemed to affect the biochemical progression of the
disease as determined by a change in PSA log sloperef.
and MUC-1 (pNGVL-hFLex-MUC-1) is able to induce antigen-specific CTL immunity
in vivo, resulting in a potent anti-tumour response, and the Flt-3L
component is essential to the efficacy of the DNA vaccine. Moreover, the
route of immunization is critical in determining the type of immune response
generated; intramuscular (i.m.) immunization with pNGVL-hFLex-MUC-1 conferred
tumour protection in contrast to poor response with hydrodynamic-based
intravenous delivery. Post-i.m. immunization, a massive infiltration of
mononuclear cells to the injection site was observed, comprised predominantly
of CD11c+/CD8a- DC. Flt-3L
acts as an adjuvant to recruit DC, thereby priming the anti-tumour response.
However, systemic expansion of DC prior to immunization did not enhance
the specific cellular response, suggesting that it is in situ recruitment
or expansion of DC that is critical for pNGVL-hFLex-MUC-1 potencyrefref1,
ref2
(expressed in > 85% of human cancers) : the main risk of targeting dominant
TERT epitopes is induction of autoimmunity in activated B cells. Immunity
against low-affinity epitopes is induced with heteroclitical variants.
The CTL repertoire against high-affinity epitopes of murine TERT (mTERT)
is partially tolerized, while that against low-affinity epitopes is composed
of frequent CTLs with high avidity. The high-affinity p797 and p545 mTERT
epitopes are not able to protect mice from a lethal challenge with the
mTERT-expressing EL4-HHD tumor. In contrast, mice developing CTL responses
against the p572 and p988 low-affinity epitopes exhibit potent antitumor
immunity and no sign of autoimmune reactivity against TERT-expressing normal
tissuesref.
Endogenous in vivo priming of T cells against hTERT is a rare event
in multiple myeloma
patients. These results suggest that strategies targeting hTERT may have
to utilize techniques to induce T cell responses from a naive precursor
frequencyref.
In an attempt to develop an effective vaccine against most human cancers,
a chemotactic-hTERT vaccine has been constructed. 2 hTERT fragments encoding
multiple cytotoxic T lymphocyte and T helper cell epitopes were fused as
a tumor antigen (named Te). The plasmid based DNA vaccine (pCCL21-Te-Fc)
was constructed by linking human CCL21 and IgG Fc gene sequences to each
end of Te. In poorly immunogenic B16F10 mouse melanoma model, DNA (pCCL21-Te-Fc)
vaccination significantly inhibited tumor growth and all of the mice were
dead by day 52. The immunization with pCCL21-Te-Fc-modified tumor cells
(B16/CCL21-Te-Fc) resulted in a higher antitumor effect than DNA vaccination
and 25% of tumor-bearing mice achieved long-term survival (>120 days).
The combined therapy of B16/CCL21-Te-Fc plus anti-4-1BB MAbs further enhanced
the immune response, resulting in 75% of tumor-bearing mice achieved long-term
survival (>120 days) in subcutaneous model and few lung nodules in pulmonary
metastasis model. Rechallenge experiment showed that a persistent memory
response was successfully induced by the combined therapy. In vivo depletion
of lymphocytes indicated that CD8+ T cells were essential in
the antitumor activity induced by B16/CCL21-Te-Fc + anti-4-1BB MAbs, whereas
NK cells and CD4+ T cells played substantial roles. The CTL
activity induced by pCCL21-Te-Fc-transfected PBMCs specifically lysed a
variety of HLA-matched and hTERT-positive human tumor cells, suggesting
pCCL21-Te-Fc could serve as a vaccine against most human cancersref.
HLA-B*0702-restricted peptides from hTERT were selected by using a method
of epitope prediction and tested for their immunogenicity in human (in
vitro) and HLA-B*0702 transgenic mice (in vivo). All the 6 hTERT
peptides that were predicted to bind to HLA-B*0702 molecule were found
to induce primary human CTL responses in vitro. The peptide-specific CD8+
CTL lines were tested against various hTERT+tumor cells. Although
differences were observed according to the tumor origin, only 3 CTL lines
specific for p277, p342, and p351 peptides exhibited cytotoxicity against
tumor cells in a HLA-B*0702-restricted manner. In addition, this cytotoxicity
was inhibited by the addition of peptide-loaded cold target cells and indicated
that these epitopes are naturally processed and presented on the tumor
cells. Further, in vivo studies using humanized HLA-B*0702 transgenic
mice showed that all the candidate peptides were able to induce CTL responses
after peptide immunization. Furthermore, vaccination with a plasmid DNA
encoding full-length hTERT elicited peptide-specific CTL responses, indicating
that these epitopes are efficiently processed in vivo. Together
with previously reported hTERT epitopes, the identification of new CTL
epitopes presented by HLA-B*0702 increases the applicability of hTERT-based
immunotherapy to treating cancerref
: mutations leading to functional inactivation of the p53 tumour suppressor
protein occurs in ~50% of human cancers. Subsequent accumulation of mutant
p53 in nuclei and cytopasm is associated with high-level presentation of
p53-derived peptides by MHC class I molecules on malignant cells. Due to
the diversity of p53 mutations in different tumours, peptides representative
of wild type, as opposed to mutant p53 sequences, were proposed to serve
as a universal TAA for a CTL-based immunotherapy of cancer. Increased susceptibility
of malignant cells to lysis by wt p53-specific CTLs has been demonstrated
to be linked rather to a high turnover rate than to a high steady-state
level of p53 expression. A major barrier to the design of broad-spectrum
p53-specific immunotherapeutics, however, has been the observation that
low-level expression of wt p53 peptides by non-transformed tissues and
cells results in self-tolerance of T lymphocytes with high avidity for
self-MHC class I/self-p53-peptide complex. Although the peripheral T cell
repertoire is mostly devoid of such high-avidity wt p53 epitope-specific
CTLs due to self-tolerance, this does not necessarily preclude the possibility
of developing wt p53-directed vaccination strategies. Low-avidity and even
residual high-avidity CTLs have been demonstrated to partially escape the
induction of wt p53-specific self-tolerance : these can be primed and amplified
in
vivo.
for breast
carcinomas
and oral carcinomas.
HER-2 gene is highly overexpressed in approximately 25% to 30% of invasive
breast carcinomasref1,
ref2,
and in comparison to normal breast epithelial cells, tumor cells may have
up to a 100-fold greater expression of HER-2ref,
with both dominant-negative and a gain-of-function properties, display
early aggressive and metastasizing parotid
tumors.
Multiple acinar and ductal hyperplasia foci overexpressing the HER-2/neu
gene product are evident at wk 5 and progress to poorly differentiated
carcinoma by wk 7. Mice die before wk 18 with invasive carcinomas and multiple
metastases that no longer express HER-2/neu. A combination of repeated
electroporations of plasmids coding for the extracellular and transmembrane
domains of the rat HER-2/neu receptor with systemic IL-12 administrations
started when the parotids that present diffuse hyperplasia protected all
female and 50% of the male mice until the close of the experiment at wk
40. This combined treatment began when multifocal in situ carcinomas
that were already present cured 33% of the females and 25% of the males.
The most prominent immunologic features associated with the antitumor protection
were the production of high titers of anti-HER-2/neu Abs and the nonappearance
of cell-mediated cytotoxic reactivity. In conclusion, anti-HER-2/neu vaccination
combined with systemic IL-12 control parotid carcinomas as far as p53 mutation
makes their growth independent of HER-2/neu expressionref.
and melanoma
in humans, and it was useful as a novel tumor marker. The preimmunization
of BALB/c mice with dendritic cells pulsed with the H-2Kd-restricted
mouse GPC3298-306 (EYILSLEEL) peptide prevented the growth of
tumor-expressing mouse GPC3. Because of similarities in the peptide binding
motifs between H-2Kd and HLA-A24 (A*2402), the GPC3298-306
peptide therefore seemed to be useful for the immunotherapy of HLA-A24+
patients with HCC and melanoma. The GPC3(298-306) peptide could induce
GPC3-reactive CTLs from the peripheral blood mononuclear cells (PBMC) of
HLA-A24 (A*2402)+ HCC patients. In addition, HLA-A2.1 (HHD)
transgenic mice were used to identify the HLA-A2 (A*0201)-restricted GPC3
epitopes to expand the applications of GPC3-based immunotherapy to the
HLA-A2+ HCC patients. The GPC3144-152 (FVGEFFTDV)
peptide could induce peptide-reactive CTLs in HLA-A2.1 (HHD) transgenic
mice without inducing autoimmunity. In 5 out of 8 HLA-A2+ GPC3+
HCC patients, the GPC3144-152 peptide-reactive CTLs were generated
from PBMCs by in vitro stimulation with the peptide and the GPC3298-306
peptide-reactive CTLs were also generated from PBMCs in 4 of 6 HLA-A24+
GPC3+ HCC patients. The inoculation of these CTLs reduced the
human HCC tumor mass implanted into nonobese diabetic/severe combined immunodeficiency
miceref
is expressed and extremely sensitive, easily detectable and highly correlated
with breast cancer. A gene vaccine was developed using a plasmid vector
to deliver the six copies of 10-amino acid residues of b-hCG
109-118 and b hCG C-terminal 37-amino acid (CTP37).
BALB/c female mice were immunized with a combination of pCR-HBc-X6-betahCGCTP37
DNA vaccine and HSP-X6-betahCGCTP37 protein vaccine. pCR-HBc-X6-betahCGCTP37
DNA vaccine were injected intramuscularly 3 times, on days -46,-25 and
-11 and HSP-X6-bhCGCTP37 protein were applied
2 times, 21 and 14 days before tumor cell challenge. A combined DNA and
protein vaccine was assessed for its effect of against murine EMT6 mammary
tumor cells. In this study, animals vaccinated DNA vaccination boosting
with the repeat b-hCG C-terminal peptide carried
by mycobacterial heat-shock protein HSP65 induced higher avidity antibodies
and effectively inhibited the growth of tumor, compared with treatment
using DNA alone or BCG priming HSP-X6-bhCGCTP37
protein boosting. The data presented demonstrate that improve immunogenicity
of DNA vaccination by boosting with the repeat b-hCG
C-terminal peptide carried by mycobacterial heat-shock protein HSP65, which
should prove useful in the development of new DNA vaccine against growth
factors for cancer immunotherapyref
is overexpressed in a variety of solid tumors : HLA-A*0201-binding peptides
and a naturally processed and presented T-cell epitope capable of inducing
CYP1B1-specific CTLs in HLA-A2 transgenic miceref.
Immunization of HLA-A2/Kb transgenic mice with human CYP1B1
encoding plasmid DNA formulated in poly(lactide-co-glycolide) (PLG) microparticles
elicits CD8+ T cells that respond to human CYP1B1+
target cells. PLG-encapsulated DNA therapeutic elicits durable immune responses
to CYP1B1, the responses are dependent on repeat immunization, and that
the formulation is well toleratedref.
Endogenous in vivo priming of T cells against CYP1B1 is a rare event
in multiple myeloma
patients. These results suggest that strategies targeting CYP1B1 may have
to utilize techniques to induce T cell responses from a naive precursor
frequencyref
cells but not in normal cells. To explore the potential of HM1.24 as a
target for cellular immunotherapy, 4 HM1.24-derived peptides that possess
binding motifs for HLA-A2 or HLA-A24 were selected by using 2 computer-based
algorithms. The ability of these peptides to generate cytotoxic T lymphocytes
(CTLs) was examined in 20 healthy donors and 6 patients with MM by a reverse
immunologic approach. Dendritic cells (DCs) were induced from peripheral-blood
mononuclear cells of healthy donors or peripheral-blood stem-cell (PBSC)
harvests from patients with MM, and autologous CD8+ T cells
were stimulated with HM1.24 peptide-pulsed DCs. Both IFN-g-producing
and cytotoxic responses were observed after stimulation with either HM1.24-126
or HM1.24-165 peptides in HLA-A2 or HLA-A24 individuals. The peptide-specific
recognition of these CTLs was further confirmed by tetramer assay and cold
target inhibition assay. Importantly, HM1.24-specific CTLs were also induced
from PBSC harvests from patients with MM and these CTLs were able to kill
MM cells in an HLA-restricted manner. These results indicate the existence
of functional DCs and HM1.24-specific CTL precursors within PBSC harvests
and provide the basis for cellular immunotherapy in combination with autologous
PBSC transplantation in MMref.
The HM1.24 sequence was scanned for immunogenic peptides using the HLA-binding
prediction software SYFPEITHI and BIMAS. Peripheral blood mononuclear cells
from HLA-A2+ healthy volunteers/blood donors (ND) were stimulated
with autologous HM1.24-peptide-loaded dendritic cells, and expanded in
vitro. Activation of T cells was assessed by ELISpot and cytotoxicity
by 51Cr-release assays. T2-cells pulsed with irrelevant peptide,
the HM1.24-/HLA-A2+ breast carcinoma cell line MCF-7
and the HM1.24+/HLA-A2- myeloma cell line RPMI-8226
were used as controls. Expression of the HM1.24 gene (BST2) was assessed
using purified plasma cells and Affymetrix-U133A+B microarrays. Frequency
of peptide-specific CD8+ T cells was detected using the flow-cytometric
tetramer technique. Of 8 nona-peptides with the highest probability of
binding to HLA-A2, the HM1.24 aa22-30 peptide (LLLGIGILV) showed
the most frequent activation of CD8+ T cells in healthy volunteers
(specific activation in 8 of 11 [73%] ND; compared with 5-19% for the 7
other HM1.24 peptides). Antigen recognition by the HM1.24 aa22-30-specific
CD8+ T cells was HLA-A2-restricted (ELISpot with HLA-A2-blocking
antibodies: median, 15; range, 14-18 spots/well; isotype-control antibodies:
median, 47; range, 44-48). HM1.24-aa22-30-specific CD8+ T cells
lysed HLA-A2+ myeloma-derived cell lines (51Cr-release
assay: XG-1 vs MCF-7, 91% vs 0%; U266 vs MCF-7, 38% vs 4.2%; IM-9 vs RPMI-8226,
22% vs 0%). Using the cross-reactive Neisseria meningitidis peptide LLSLGIGILV-specific
CD8+ T cells recognizing target cells loaded with the HM1.24
aa22-30 peptide (LLLGIGILV) as well as the myeloma-derived cell line U266
could be expanded from MM patients. The HM1.24 gene was expressed at comparable
levels by plasma cells from 65 MM patients, 7 patients with MGUS, and 7
ND. HM1.24 aa22-30 is a newly identified HLA-A2-restricted T-cell epitope
that is processed and presented by major histocompatibility complex class
I. Specifically activated CD8+ T cells are able to lyse MM cell
lines. HM1.24 aa22-30 represents a suitable candidate target for a specific
peptide-based immunotherapy of MMref.
offers an attractive commercial opportunity for the development of TroVax.
Studies show that the 5T4 antigen is present on > 90% of RCC samples; current
treatment options are ineffective or have serious side effects; and TroVax
could benefit from orphan drug and fast track designations in this indication.
A Phase II trial in RCC with TroVax alongside high dose interleukin-2 is
underway in the United States. Preliminary immunology data are expected
around mid-2005. In breast cancer, the Southwest Oncology Group, which
is a US clinical trials consortium, is expected to start a Phase II trial
in late stage patients, in 2005. Clinical response data from Oxford BioMedica’s
Phase II trials are unaudited. They are based on patients that are evaluable
and use the industry standard Response Evaluation Criteria in Solid Tumours
(RECIST). The data from the trials of the chemotherapy agents alone, cited
in this press release, are derived from evaluable patients, using clinical
response criteria set by the World Health Organisation (WHO). The criteria
defined by RECIST and WHO are similar but not identical.
-melanocyte
differentiation antigens (MDAs) / melanoma-associated antigens (MAAs))
present on T-cells and dendritic cells is important in activating and regulating
immunity. The effectiveness of CCL21 as an adjuvant with an HVJ anionic
liposomal TRP2 DNA (plasmid) vaccine to enhance anti-TRP2 Ab, cytokines,
delayed-type hypersensitivity, T-cell responses, and tumor protection against
B16 melanoma cells was investigated. Induction of anti-TRP2 immunity depended
mainly on cell-mediated immunity, which was regulated by timing and route
of CCL21 administration with DNA vaccine. The optimum protocol was to administer
CCL21 im 24 h before DNA vaccine at the same vaccination site. 2 vaccinations
(prime/boost) were essential for induction of strong anti-TRP2 cell-mediated
immunity. CCL21 administration 3 days before or 24 h after DNA vaccine,
simultaneous with DNA vaccine, or at different sites (iv, opposite leg)
was not effective. This study demonstrated that CCL21 was an effective
adjuvant to enhance TRP2-specific immunity induced by a plasmid DNA cancer
vaccineref
by using melanoma Ags common to normal melanocytes (when vaccine is injected
i.d. but not when injected i.m.ref
: see danger theory)
and risk for subacute demyelinating polyneuropathy using melanoma
Ags common to normal Schwann cells. 26 patients with advanced melanoma
were randomly assigned to vaccination with a mixture of 4 gp100 and tyrosinase
peptides restricted by HLA-A1, HLA-A2, and HLA-A3, plus a tetanus helper
peptide, either in an emulsion with GM-CSF
and Montanide ISA-51 adjuvant, or pulsed on MDDCs. Systemic low-dose IL-2
was given to both groups. Anti-melanoma cell responses are observed in
the sentinel immunized lymph node (SIN) that drain the vaccine site in
80% of patients in the GM-CSF group, but only 13% of the patients that
are vaccinated with peptide-loaded DCs. Favourable outcomes, including
tumour shrinkage, are observed in 31% of patients who received the GM-CSF-based
vaccine, and 2 patients experienced stable disease for several months.
Favourable outcomes were only observed in 15% of patients in the DC group,
and only 1 patient experienced stable disease. 2 patients who received
te GM-CSF-containing vaccine also developed vitiligoref
for prostate
adenocarcinoma.
Side effect : autoimmune prostatitis.
Low doses of the pPSA vaccine were not capable of inducing tumor protection,
but when pIL-18
was co-administered, complete tumor protection was observed in all mice.
Tumor protection was mediated by both CD4+ and CD8+
T cells. Detailed analysis of the immune response in mice immunized with
either pPSA or pPSA/pIL-18 demonstrated that pIL-18 skewed the PSA-specific
immune response toward Th1. More importantly, stronger CD4+
and CD8+ T cell responses developed in the pPSA/pIL-18-immunized
mice, with faster kinetics. These results suggest that IL-18 is a powerful
adjuvant molecule that can enhance the development of antigen-specific
immunity and vaccine efficacyref.
In a mouse model, the CD8+ T lymphocyte response to a prostate
cancer DNA vaccine encoding PSA were evaluated after intradermal electroporation.
A significantly increased gene expression (100- to 1000-fold) and higher
levels of PSA-specific T cells, compared to DNA delivery without electroporation,
was demonstrated. Interestingly, investigation of a panel of different
electroporation conditions showed that only some conditions that induce
high levels of gene expression additionally induced cellular immunity.
This suggests that electroporation parameters should be carefully optimized,
not only to enhance transfection efficiency, but also to enhance the immune
response to the vaccineref.
Several DNA fragments encoding multiple CTL and T helper cell epitopes
were selected from human prostate-specific membrane antigen (hPSM), mouse
prostatic acid phosphatase (mPAP), and human prostate-specific antigen
(hPSA). These DNA fragments were ligated together to form a novel fusion
gene, termed the 3P gene. The secondary lymphoid tissue chemokine (SLC),
3P and human IgG Fc genes were inserted into pcDNA3.1 to construct a DNA
vaccine, designated pSLC-3P-Fc. After vaccination, the DNA is taken up
by cells that produce and secrete the SLC-3P-Fc fusion proteins, termed
chemotactic antigen (chemo-antigen). The secreted chemo-antigens,
in addition to promoting the co-localization of naive, non-polarized
memory T cells and DCs, are efficiently captured and processed by dendritic
cells via receptor-mediated endocytosis and then cross-presented to both
major histocompatibility complex class I and class II in a cognate manner.
The results of this study demonstrate that vaccination with pSLC-3P-Fc
by gene gun inoculation induced a strong antitumour response in a mouse
tumour model, which significantly inhibited tumour growth and prolonged
the survival time of the tumour-bearing mice. In vitro, the secreted
SLC-3P-Fc fusion protein can attract lymphocytes from human peripheral
blood mononuclear cells (PBMC); when human lymphocytes were stimulated
by pSLC-3P-Fc-transfected autologous PBMC, CTLs were induced which could
specifically kill hPSM-, hPAP-, or hPSA-expressing tumour cells. These
observations provide a new vaccine strategy for cancer therapy through
promoting the co-localization of lymphocytes and the concomitant enhancement
of antigen-specific CD4+ helper and CD8+ cytotoxic
T-cell responses against tumourref.ref.
A phase I study of a DNA vaccine targeting PAP in patients with stage d0
prostate cancerref,
with highest levels of expression observed in high-grade cancers, metastatic
disease, and androgen-independent tumorsref1,
ref2,
ref3,
ref4.
The membrane-associated character and correlation between expression levels
and disease grade have enabled PSMA to become an important biomarker for
prostate cancer, and antibodies to PSMA are currently being developed for
the diagnosis and imaging of recurrent and metastatic prostate cancer as
well as for the therapeutic management of malignant diseaseref1,
ref2,
ref3,
ref4.
The LNCaP prostate cancer cell line had an estimated 180,000 molecules
of PSMA per cell
chain is a primary binding and internalization subunit for a Th2-derived
immune regulatory cytokine, IL-13. Although extremely high levels of IL-13Ra2
chain are expressed on a variety of human tumor cells and specimens. D5
melanoma cells were stably transfected with the human IL-13Ra2
gene (D5a2) to assess the effect
of an IL-13Ra2 DNA vaccine in immunocompetent
animals. Prophylactic immunization of mice with the IL-13Ra2
DNA vaccine resulted in protection against D5a2
tumor development. In vivo depletion experiments in C57BL/6 and
RAG-2-/- mice indicated that both T and B cells, but not NK
cells, were required for the tumor protection. In addition, antibody induced
by the IL-13Ra2 DNA vaccine showed
a modest but significant inhibitory effect on D5a2
cells in vitro, suggesting that the antibody is biologically functional.
The IL-13Ra2 DNA vaccine also exhibited
antitumor activity against established D5a2
tumors in mice. Histologic analysis of regressing tumors identified infiltration
of CD4+ and CD8+ T cells and the expression of CXCL9
chemokine in tumorsref
for 90% of primary prostate
adenocarcinoma
and lung
spindle cell carcinomas
and 50% of primary breast
carcinomas
: presented on HLA-A*02.01
produced an immune response in 60% of patients tested - or 20 of 33 evaluable
patients - with active or relapsed AML,
refractory CML
or high-risk MDS.
Patients only get the vaccine 3 times, yet an immune response has been
measured in patients 4 years after treatment. Of those 20 patients who
mounted an immune response against their cancer, 14 had an overall survival
of 4 years, and none of the 13 patients who did not have such a response
lived that long. 3 patients are in "molecular remission" in which there
is no evidence of the disease remaining.
with mutations in APC,
axin
1 and b1-catenin
genes.ref.
It is a member of the secretoglobin protein familyref
successfully used for DNA immunization in murine models by researchers
at Washington University School of Medicine and the Siteman Cancer Center
in St. Louis
is expressed at high levels in leukemic blast cells in most acute
myeloid leukemias
and acute lymphoblastic leukemias.
In myelodysplastic syndrome,
WT1 mRNA expression levels increase along with disease progression; thus,
WT1 mRNA is a tumor marker for leukemic blast cells. WT mRNA is also expressed
at high levels in various types of solid cancers, including cancers of
the lung, breast, colon and pancreas. Patients with WT1-expressing tumors
produce antibodies and cytotoxic T-lymphocytes against WT1 protein, indicating
that WT1 protein is highly immunogenic and a promising tumor antigen. MHC
class I-restricted CTL and class II-restricted helper epitopes of WT1 protein
were identified, and clinical studies of cancer immunotherapy using these
CTL epitope peptides were performed without significant adverse effect
and with clinical results promising enough to encourage further clinical
trials. The clinical efficacy of cancer immunotherapy targeting the WT1
protein should be clarified by a large-scale clinical studyref.
The expression of WT1 was examined in 494 cases of human cancers, including
tumors of the gastrointestinal and pancreatobiliary system, urinary tract,
male and female genital organs, breast, lung, brain, skin, soft tissues
and bone by immunohistochemistry using polyclonal (C-19) and monoclonal
(6F-H2) antibodies against WT1 protein. Staining for C-19 and 6F-H2 was
found in 35-100 and 5-88% of the cases of each kind of tumor, respectively.
WT1-positive tumors included tumor of the stomach, prostate, and biliary
and urinary systems, and malignant melanomas. A majority of the positive
cases showed diffuse or granular staining in the cytoplasm, whereas ovarian
tumors and desmoplastic small round cell tumors frequently showed nuclear
staining. Glioblastomas, some of soft tissue sarcomas, osteosarcomas, and
malignant melanomas of the skin showed extremely strong cytoplasmic staining
as compared with other tumors. Western blot analysis showed that WT1 protein
was predominantly expressed in the cytoplasm of the tumor cells in 2 cases
of lung adenocarcinoma, supporting the intracytoplasmic staining for WT1
using immunohistochemistryref.
is a pivotal regulator of apoptotic cell death and it is overexpressed
in many cancers. Consequently, the Bcl-2 protein is an attractive target
for drug design, and Bcl-2-specific antisense oligonucleotides or small-molecule
Bcl-2 inhibitors have shown broad anticancer activities in preclinical
models and are currently in several clinical trials. The clinical application
of immunotherapy against cancer is rapidly moving forward in multiple areas,
including the adoptive transfer of anti-tumor-reactive T cells and the
use of "therapeutic" vaccines. The overexpression of Bcl-2 in cancer and
the fact that immune escape by down-regulation or loss of expression of
this protein would impair sustained tumor growth makes Bcl-2 a very attractive
target for anticancer immunotherapy. Spontaneous T-cell reactivity against
Bcl-2 in peripheral blood from patients suffering from unrelated tumor
types (ie, pancreatic
cancer,
breast
cancer,
acute
myeloid leukemia (AML),
and B-cell
chronic lymphocytic leukemia)
has been described : these Bcl-2-reactive T cells are indeed peptide-specific,
cytotoxic effector cells. Thus, Bcl-2 may serve as an important and widely
applicable target for anticancer immunotherapeutic strategies (eg, in the
combination with conventional radiotherapy and chemotherapy)ref.
and melanomaref.
The recombinant HSP105 did not induce DC maturation, and the mice vaccinated
with HSP105-pulsed BM-DCs were markedly prevented from the growth of subcutaneous
tumors, accompanied with a massive infiltration of both CD4+
T cells and CD8+ T cells into the tumors. In depletion experiments,
we proved that both CD4+ T cells and CD8+ T cells
play a crucial role in anti-tumor immunity. Both CD4+ T cells
and CD8+ T cells specific to HSP105 were induced by stimulation
with HSP105-pulsed DCs. As a result, vaccination of mice with BM-DCs pulsed
with HSP105 itself could elicit a stronger tumor rejection in comparison
to DNA vaccinationref.
: WT1124-138 and WT1247-261 were shown to induce
peptide-specific anti-tumor CD4+ helper T lymphocytes (HTL),
which were restricted by frequently expressed HLA class II alleles. Both
peptides-reactive HTL lines were capable of recognizing naturally processed
antigens presented by DCs pulsed with tumor lysates or directly by WT1+
tumor cells that express MHC class II molecules. Interestingly, the 2 WT1
HTL epitopes described here are closely situated to known MHC class I-restricted
CTL epitopes, raising the possibility of stimulating CTL and HTL responses
using a relatively small synthetic peptide vaccineref-melanocyte
differentiation antigens,
a 16.5kDa TAA, is the smallest member of the inhibitor of apoptosis family
that is abundantly expressed during development but essentially absent
in normal adult tissues. Interestingly, survivin expression is up-regulated
in virtually all types of cancers studied, as well as in vascular endothelial
cells during tumor associated angiogenesis. Survivin links apoptosis to
cell cycle progression and plays a pivotal role in regulation of cell proliferationref.
These characteristics make survivin a potentially promising generic target
for cancer immunotherapy. Overexpression of survivin in tumors is linked
to decreased patient survival, increased tumor recurrence, and resistance
to therapyref.
Spontaneous immune responses against survivin were recently demonstrated
in a variety of patients with cancerref.
Survivin-based DNA vaccines were also shown to induce T cell-mediated antitumor
responses without severe toxicities in preclinicalref
and clinical trialsref.
Hence, a genetic immunization strategy to induce tumor-specific immune
responses against human survivin in a pre-clinical animal model was developed.
In initial studies, BALB/c mice were immunized by intramuscular injection
with DNA coding for human survivin (pcDNA3.1/hSurv). In addition, a construct
encoding a secreted version of survivin (pSecTag2B/hSurv) was designed.
A plasmid coding for murine GM-CSF was co-injected in both cases as a molecular
adjuvant. Expression of survivin following transfection in mouse cells
was corroborated. Humoral responses against human survivin were detected
in mice sera using 2 immunization protocols (injections at 2- or 3-week
intervals). The humoral response was markedly improved by secretion of
survivin and co-expression of GM-CSF. The predominant antibody subclass
detected in responsive mice was IgG2a, suggesting that a Th1-CD4+
cellular response had been induced. Furthermore, DNA immunization with
survivin encoding vectors generated an effective CD8+ T cell
response measured as an increase of cytotoxic IFN-g
secreting CD8+ T cells. In conclusion, intramuscular genetic
immunization of mice with human survivin encoding plasmids induced a survivin-specific
humoral as well as cellular immune response in recipient mice. Secretion
of survivin and co-injection of GM-CSF as a genetic adjuvant appear to
be more important in generating an humoral than a cellular immune responseref
on recipient cells in allogeneic
HSCT
recipients mediate the GvL effect. For minor histocompatibility antigens
HA-1 and HA-2, normal cell expression is restricted to hemopoietic cells,
and boosting the immune response to these antigens may potentiate GvL effect
without accompanying GvHD. To increase efficacy, expansion of HA-1- or
HA-2-specific CTL before transplantation is desirable. However, primary
HA-1- or HA-2-specific CTL expanded in vitro are often of low avidity.
An alternative approach is to prime specific CTL responses in vivo by vaccination.
Clearly, donor vaccination must be safe and specific. DNA fusion vaccines
able to induce high levels of epitope-specific CTL using linked CD4+
T-cell help have been developed. The vaccines incorporate a domain of tetanus
toxin (DOM) fused to a sequence encoding a candidate MHC class I binding
peptide. This design generates antitumor CD8+ T-cell responses
and protective immunity in preclinical models. For clinical application,
vaccines encoding HLA-A*0201-restricted peptides from human HA-1 and HA-2,
which were fused to DOM, were constructed and tested performance in HLA-A*0201-transgenic
mice. Priming induced epitope-specific, IFN-g-producing
CD8+ T cells with cytotoxic function boosted to high levels
with electroporation. Strikingly, these mouse T cells efficiently killed
human lymphoblastoid cell lines expressing endogenous HA-1 or HA-2. High
avidity is indicated by the independence of cytolysis from CD8/MHC class
I interaction. These safe epitope-specific vaccines offer a potential strategy
to prime HA-1- or HA-2-specific CTL in transplant donors before adoptive
transferref
on recipient cells in the setting of allogeneic
HSCT
mediate graft-versus-leukemia effect by T-cells. For minor histocompatibility
antigens HA-1 and HA-2, normal cell expression is restricted to hemopoietic
cells, and boosting the immune response to these antigens may potentiate
graft-versus-leukemia effect without accompanying graft-versus-host disease.
To increase efficacy, expansion of HA-1- or HA-2-specific CTL before transplantation
is desirable. However, primary HA-1- or HA-2-specific CTL expanded in
vitro are often of low avidity. An alternative approach is to prime
specific CTL responses in vivo by vaccination. Clearly, donor vaccination
must be safe and specific. DNA fusion vaccines able to induce high levels
of epitope-specific CTL using linked CD4+ T-cell help have been
developed. The vaccines incorporate a domain of tetanus toxin (DOM) fused
to a sequence encoding a candidate MHC class I binding peptide. This design
generates antitumor CD8(+) T-cell responses and protective immunity in
preclinical models. For clinical application, vaccines encoding HLA-A*0201-restricted
peptides from human HA-1 and HA-2 fused to DOM were constructed and their
performance was tested in HLA-A*0201-transgenic mice. Priming induced epitope-specific,
IFN-g-producing CD8+ T cells with
cytotoxic function boosted to high levels with electroporation. Strikingly,
these mouse T cells efficiently killed human lymphoblastoid cell lines
expressing endogenous HA-1 or HA-2. High avidity is indicated by the independence
of cytolysis from CD8/MHC class I interaction. These safe epitope-specific
vaccines offer a potential strategy to prime HA-1- or HA-2-specific CTL
in transplant donors before adoptive transferref.
for B
cell lymphomas
: Idiotypic (Id) vaccination has shown promising results in patients with
follicular
lymphoma.
With 2 ongoing, phase-III clinical trials enrolling newly-diagnosed FL
patients, idiotypic vaccinationref
is approaching the final stage of its clinical development, that of demonstrating
a possible benefit to patients. However, even in the event that either
or both ongoing clinical trials succeed, a number of relevant questions
would still remain unanswered, in particular whether idiotype (Id) vaccines
may be feasible for most if not all relapsed FL and for some other B-cell
malignancies. All Id vaccine production attempts were carried out by the
same personnel and always using the same fusion partner (K6H6/B5, i.e.
ATCC number: CRL-1823), according to the standard tumor/hetero-hybridoma
fusion-based method previously describedref1,
ref2,
ref3,
ref4.
Patients with FL, mantle cell lymphoma and small lymphocytic lymphoma were
supposed to receive Id vaccine treatment if they achieved either complete
(CR) or partial (PR) response, while patients with either diffuse large
cell or Burkitt's lymphoma were supposed to receive Id vaccine treatment
only if they achieved a CR. The feasibility of Id vaccination in relation
to induction treatment (if the salvage therapy did not induce a response
sufficient to proceed with Id vaccination) was markedly different, being
80-100% in indolent NHL subtypes and 40-50% in aggressive ones. This difference
was not, however, due to an overall lack of efficacy of the respective
chemotherapy regimens, but rather to the different eligibility criteria
for Id vaccination following chemotherapy. In fact, the overall response
to induction treatment for aggressive NHL was 75-80% (CR+PR), but in this
group only patients achieving CR were considered eligible to receive Id
vaccination. A far more important factor that halted treatment was Id-secreting
hybridoma production. Fusion experiments were successful in most FL cases
at the very first attempt, whereas in other NHL cases, irrespective of
the ultimate Id production outcome, as many as 5 attempts had to be carried
out most of the time. Similarly, in most FL cases, the average number of
successful fusion wells per 96-well plate was >> 15, whereas that of most
of the other NHL cases was typically < 5. Statistically significant
differences in fusion-related and overall feasibility were found between
cases of FL and those of all other NHL, indolent and aggressive lymphoma,
respectively. Both fusion-related and overall feasibility of the Id vaccine
treatment for FL in first relapse (87%) were comparable with those already
described for both newly-diagnosed and relapsed patients with the same
diseaseref1,
ref2,
ref3.
Interestingly, the Id vaccine production success rate was substantially
low in cases of mantle cell lymphoma,
as opposed to what has been preliminarily described with the very same
method in newly-diagnosed MCL patients (Neelapu SS, Wilson WH, Baskar S,
White T, Frye R, Pennington R, et al. Induction of T-cell responses by
tumor antigen vaccination in mantle cell lymphoma following rituximab-based
treatment. J Clin Oncol 2003; 22 Suppl 1:663a). This apparent discrepancy
could be due, at least in theory, to MCL cells at first relapse biologically
resembling those of aggressive NHL rather than FL clones, with obvious
possible repercussions on the fusion process. The main cause for Id vaccine
production failure was poor hybridoma survival (83% of cases), which accounted
for 100% of failures in aggressive lymphoma (9/9 cases). Loss of
Id secretion by growing hybridomas accounted for only 17% of cases
of Id vaccine production failure. In the vast majority of cases, the feasibility
of idiotypic vaccination for patients with first-relapse B-cell malignancies
strictly depends on the ultimate ability to produce a viable Id vaccine
rather than on the probability of inducing a clinical response suitable
for subsequent idiotypic vaccination. In this respect, first-relapse FL
clearly appears more suitable for hybridoma-based Id vaccine production
than any other first-relapse NHL subtype tested. As a major consequence
of these data, clinical trials on Id vaccination for B-cell lymphomas other
than FL in first relapse have been closed. However, it is possible that
alternative methods to produce the Id protein, particularly those based
on molecular techniquesref1,
ref2,
ref3,
ref4,
may prove far more efficient than the traditional approachref.
Anti-idiotypic antibodies that bind to the BcR complex appear to induce
apoptosis. In a fraction of patients treated with anti-idiotypic antibodies,
recurrence is associated with loss of the relevant idiotypic determinants
as genes encoding the cell surface BcR continue to undergo somatic
hypermutation.
Vaccination with Id-pulsed dendritic cells (DCs) or with soluble Id-KLH
in adjuvant induced immune responses that eliminated both B-cell lymphoma
and myeloma in tumor-bearing mice; however, the 2 vaccination regimens
resulted in distinct immune responses. Whereas soluble Id plus adjuvant
induced high levels of anti-Id antibodies, the Id-pulsed DCs did not induce
anti-Id or any antitumor antibodies. Immunization with Id-pulsed DCs induced
a significant increase in the frequency of Id-reactive T cells. Depletion
studies in DC-vaccinated mice showed that the predominant effector cells
responsible for tumor rejection were of the CD8 subset. The finding that
DC-based Id vaccines elicit tumor protection, which is entirely based on
cell-mediated effector mechanisms, is of particular importance for plasma
cell tumors because these tumors do not express Id on the surface and hence
do not bind anti-Id antibodiesref.
(MyVax© / GTOP99 personalized
immunotherapy; source : Genitope Corporation)
for follicular lymphoma
is in phase 3 trial : 8 cycles of CVP
=> if at least partial response, => 7 s.c. vaccinations given over 24 weeks
(wk 0, 4, 8, 12, 16, 20 and 24) + 4 injections of GM-CSF each cycle
: with a median follow-up of 9.2 years, 45% of patients remained
in continuous remission, OS = 95%, DFS = 96.5 months.ref|
|
epitopes presentable by MHC class I molecules of |
|
|
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|
|
|
|
|
|||||||
|
|
|
|
|
|
|
plasmid priming + AAV boosting | ||||
| BCL1 | anti-Id antibodies-dependent
(30-80%) |
anti-Id antibodies-dependent (30-100%) | ||||||||
| 38C13 (Href, Lref) | anti-Id antibodies-independent
(30%) |
no anti-idiotype antibodies induced (exception : )
(40%) |
no anti-Id antibodies induced
(80%) |
|||||||
| JLms (sIg-) | CD8+ T cell-dependent
(20%) |
CD8+ T cell-dependent
(50%) |
no anti-Id antibodies induced
CD8+ T cell-dependent |
|||||||
cell lines expressing the epithelial marker, epithelial cell adhesion molecule
(EpCAM). VH gene transcripts were identifiable by nested reverse
transcription-PCR either as single or dual VDJ rearrangements in 4 of 6
lines, most being potentially functional. V(D)J transcripts were observed
in sequential cultures, indicating stable expression. To exclude coexisting
lymphocytes, each cell line was shown to be EBV negative, with CD19/CD20
and cytoplasmic/surface immunoglobulin also absent by flow cytometry. Identified
VH transcripts were then sought in individual tumor cells, isolated
as EpCAM+ single cells by flow cytometry. Importantly, in 3
of 3 selected cell lines, VH genes were identifiable in a significant
fraction (approximately 32%) of single cells. In 5 of 6 identified VH
genes, somatic mutations were apparent with no intraclonal variation, indicating
cessation of mutational activity. VH transcripts were pre- and
post-isotype switch, with activation of switch events evident from expressed
germ-line switch transcripts in two of six lines. Strikingly, 6 of 6 cell
lines expressed activation-induced cytidine deaminase (AID) essential for
mutational and switch activity. These data suggest either a de novo
rearrangement and modification of VH genes in epithelial tumor
cells or assimilation of lymphocyte-derived chromatin. Constitutive AID
activation in malignant epithelial cells further raises a potential for
inducing aberrant mutational activityref
ref
for T
cell lymphomas
:.
Favrille uses a similar approach with its lead product candidate FavId
for the treatment of B-cell NHL. In January the Company completed enrollment
in a pivotal Phase 3 clinical trial of FavId following induction therapy
with rituximab, the current standard of care. Favrille has received Fast
Track designation from the FDA for FavId.
gene in the 5' end fused to a GDP-L-fucose:b-D-galactoside-2-a-L-fucosyltransferase
(FUT1) in an antisense orientation in the 3' end.
in acute promyelocytic leukemia
(APL)
: lipo ATRA was highly effective in inducing durable molecular remissions
in immunocompetent mice [C57BL/6 x C3H F(1) (B6C3HF1)]; arsenic therapy
was much less effective, and did not clearly synergize with Lipo ATRA to
increase the remission rate in immunocompetent mice. The survival of Lipo
ATRA-treated severe combined immunodeficient (SCID) animals (lacking functional
T and B cells) was inferior to that of immunocompetent B6C3HF1 recipients
(40% vs. 88% survival at 1 y, P < 0.001). These data suggest that adaptive
immunity cooperates with pharmacologic therapy to induce or maintain remissions
in murine APL. It also implies that immunosuppressive anti-leukemia therapies
could paradoxically blunt effective anti-leukemia immune responses that
are important for clearing small numbers of residual tumor cells after
chemotherapy-mediated cytoreductionref.
oncogene to tetanus fragment C (FrC) sequences can have a pronounced effect
on survival in mice with an animal model of APL, both alone and when combined
with all-trans retinoic acid (ATRA). The survival advantage is concomitant
with time-dependent antibody production and an increase in IFN-g.
ATRA therapy on its own triggers an immune response in this model. When
DNA vaccination and conventional ATRA therapy are combined, they induce
protective immune responses against leukemia progression in mice and may
provide a new approach to improve clinical outcome in human leukemiaref./c-Abl
in CML
: the junctional sequences represent potentially immunogenic TSAs..
No significant toxic effects were observed. In 14 of 14 patients, delayed-type
hypersensitivity (DTH) and/or CD4 proliferative responses developed after
beginning vaccinations, and 11 of 14 patients showed IFN-g
release by CD4 ELISPOT at one or more time points. These responses were
CD4+CD45RO+. A peptide-specific CD8+ IFN-g
ELISPOT was found in 4 patients. 4 patients in hematologic remission had
a decrease in Philadelphia chromosome (Ph) percentage (3 concurrently receiving
IFN-a
and 1 on imatinib mesylate), and 3 patients in molecular relapse after
allogenic transplantation became transiently PCR- after vaccination;
2 of these patients received concurrent donor
lymphocyte infusion (DLI). All 5 patients on IFN-a
ultimately reached a complete cytogenetic remission. A relationship between
the clinical responses and vaccination cannot be determined from this trialref
as an immunological adjuvant. Delayed-type hypersensitivity (DTH), humoral
responses, and unprimed ex vivo autologous proliferation (3H-thymidine
incorporation) and cytotoxicity (chromium-51 release) responses were measured.
All 68 vaccinations were well tolerated without significant adverse effects.
In 3 of the 6 patients treated at the 2 highest dose levels of vaccine,
peptide-specific, T-cell proliferative responses (n = 3) and/or DTH responses
(n = 2) were generated that lasted up to 5 months after vaccination. CTLs
have not been identifiedref,
and no further reduction of residual disease for at least 6 months preceding
enrolment were recruited and given 6 vaccinations with a peptide vaccine
derived from the sequence p210-b3a2 + molgramostim
and QS-21
as adjuvants (CMLVAX100) before assessment of immunological and
disease response, which included detecting amounts of b3a2 transcripts
by standardised quantitative real-time RT-PCR. Of 10 patients on imatinib,
9 started CMLVAX100 having had a median of 10 months' stable cytogenetic
disease (median 10% Ph+ metaphases), whereas one started in
stable CCR. All patients' cytogenetic responses improved after 6 vaccinations,
with 5 reaching CCR. Notably, 3 of these 5 patients also had undetectable
amounts of b3a2 transcript (BCR-ABL:ß2 microglobulin ratio
<0.00001). 6 patients on IFN-a treatment
with a median of 17 months' stable residual disease (median 13% Ph+
cells) were also vaccinated. All but one had improved cytogenetic responses,
and 2 reached CCR. Overall, peptide-specific delayed-type hypersensitivity
was recorded in 11 of 16 patients, CD4 cell proliferation in 13 of 14 assessed,
and interferon production in 5 of 5 assessed. Addition of CMLVAX100 to
conventional treatment in patients with CML might favour further reduction
of residual disease and increase the number of patients reaching a molecular
responseref.
Imatinib
does not impair specific antitumor T-cell immunity in patients with CMLref
or chemotherapy
:
and breast
cancer
cells that can serve as targets for immune recognition and treatment strategies.
Globo H has been synthesized, conjugated to KLH, and administered with
the immunologic adjuvant QS-21 as a vaccine for patients with prostate
cancer who have relapsed after primary therapies such as radiation or surgery.
The vaccine, given as 5 subcutaneous vaccinations over 26 weeks, has been
shown to be safe and capable of inducing specific high-titer IgM antibodies
against globo H. Its immunogenicity was confirmed in prostate cancer patients
with a broad range of stages and tumor burdens. Observations of several
patients who had evidence of disease relapse restricted to a rising biochemical
marker, PSA, indicated that a treatment effect could occur within 3 months
after completion of the vaccine therapy. This effect was manifested as
a decline of the slope of the log of PSA concentration vs. time plot after
treatment compared with values before treatment. 5 patients continue to
have stable PSA slope profiles in the absence of any radiographic evidence
of disease for > 2 yearsref.
An efficient route to complex synthetic oligosaccharides attached to an
affinity matrix for identifying and isolating antibodies elicited against
such a carbohydrate-based vaccine in humans has been reported. Pre- and
postvaccination profiles from serum samples of patients immunized with
globo H-KLH were compared. All anti-globo H antibody activity was efficiently
separated from other serum constituents. The isolated antibodies were readily
quantified, and their specificities were analyzed. Since no comparable
data were available on antibodies resulting from the vaccination of other
cancer patients, we compared the observed levels with those quoted in studies
with bacterial polysaccharide vaccines that had been quantified. Remarkably,
cancer patients immunized with globo H-KLH produce anti-globo H antibody
levels often exceeding those formed by immunization with bacterial polysaccharides.
In addition, substantial quantities of both IgG and IgM antibodies were
elicited, clearly indicating a class switch to IgGref.,
mammary, sebaceous and some ovarian
carcinomas.
Cancer cells frequently undergo incomplete glycosylation resulting in the
appearance of precursor structures that normally would be absent like the
case with the T antigen. T antigen can be detected by several different
reagents including monoclonal antibodies and several plant lectins-e.g.,
Arachis
hypogea (peanut agglutinin)(particle-mediated
immunotherapeutic delivery (PMID)-peptide
complex-based vaccinesref
(produced only for 90% of patients) (in all stage IV patients, median survival
improved by more than 50%; median survival in the Oncophage-treated arm
was 20.9 months, versus 12.8 months in the physician's choice armref1,
ref2);
in phase I/II trials at the Istituto dei Tumouri, Milan (in association
with Sigma-Tau), Italy. Preliminary data from the phase II trial for melanoma
was presented at the AACR-NCI-EORTC International Conference in Florida,
USA, in October 2001. In phase I/II trial in the USA (36 patients)ref
(produced for 100% of patients) : in phase I/II trials (30 patients) at
the Istituto dei Tumouri, Milan (in association with Sigma-Tau), Italy
(produced for 98% of patients) : in a pivotal phase III clinical trial
for renal cancer at 80 clinical sites worldwide. The trial is enrolling
at least 500 patients who are randomised to receive surgical removal of
the primary tumour followed by out-patient treatment with Oncophage((R))
or surgery only. This study was initiated on the basis of results from
a pilot phase I/II study and preliminary results from a phase II study
in patients with renal cell cancer. In October 2001, Oncophage was designated
as a fast-track product by the Food and Drug Administration in the US for
the treatment of renal cell carcinoma. Oncophage is also in a phase I/II
(42 patients) and a phase II trial (84 patients) in the US for renal cell
cancer
: phase II trial in the USA (35 patients)
: specific CTL lines were obtained after repeatedly stimulating T cells
with autologous, HLA-A*0201+ DCs pulsed with gp96 derived from
HLA-A*0201+ human myeloma cell line (HMCL) U266 or primary myeloma
cells. hese T cells lysed not only gp96-pulsed DCs, U266, and other HLA-A*0201+
HMCLs IM-9 and XG1 but also effectively killed HLA-A*0201+ primary
myeloma cells from patients. No killing was observed against unpulsed dendritic
cells, dendritic cells pulsed with control gp96, HLA-A*0201- HMCLs, and
primary myeloma cells, or HLA-A*0201+ nonmyeloma cells. Cytotoxicity
was mainly MHC class I/HLA-A*0201 restricted, suggesting that the CTLs
recognized gp96-chaperoned peptides on HLA-A*0201 that were derived from
shared myeloma antigens and that myeloma cells naturally present these
peptides in the context of their surface MHC molecules. Upon antigen stimulation,
these T cells secreted IFN-g and TNF-a,
indicating that they belong to Th1 subsetsref.
(produced only for 71% of patients) : phase I/II trials at Johannes Gutenberg-University
Hospital, Mainz, Germany (30 patients)
(produced only for 30% of patients because of the high concentration of
proteases in that tissue type) : phase I/II trials at Johannes Gutenberg-University
Hospital, Mainz, Germany. A pilot phase I trial in patients with pancreatic
cancer began in the US in 1997 with 5 patients enrolled. In November 2000,
Antigenics announced that this trial had been expanded to a phase I/II
study which would now include survival as an endpoint and would enroll
5 additional patients.-peptide
complex-based vaccines
fusion protein
fusion protein (such an approach is likely to be more effective in targeting
antigens to macrophages than to DCs, because macrophages have higher levels
of FcR's)
mAb) fusion protein
of APCs with tumour DNA encoding the full ORF of the tumor antigen
by using
genetically modified
can be exploited for clinical use in cancer immunotherapy by targeting
autologous tumour vaccines to APC, thereby increasing their immunogenicity.
Autologous intact tumour cells from haematological malignancies, or autologous
tumour cell membranes from solid tumours are processed to express a-gal
epitopes by incubation with neuraminidase, recombinant a1,3GT
and with UDP-galactose. Subsequent immunization with such autologous tumour
vaccines results in in vivo opsonization by anti-Gal IgG binding
to these a-gal epitopes. The interaction of
the Fc portion of the vaccine-bound anti-Gal with FcgRs
of APC induces effective uptake of the vaccinating tumour cell membranes
by the APC, followed by effective transport of the vaccinating tumour membranes
to the regional lymph nodes, and processing and presentation of the TAA
peptides. Activation of tumour-specific T cells within the lymph nodes
by autologous TAA peptides may elicit an immune response that in some patients
will be potent enough to eradicate the residual tumour cells that remain
after completion of standard therapy. A similar expression of a-gal
epitopes can be achieved by transduction of tumour cells with an adenovirus
vector (or other vectors) containing the a1,3GT
gene, thus enabling anti-Gal-mediated targeting of the vaccinating transduced
cells to APC. Intratumoral delivery of the a1,3GT
gene by various vectors results in the expression of a-gal
epitopes. Such expression of the xenograft carbohydrate phenotype is likely
to induce anti-Gal-mediated destruction of the tumour lesion, similar to
rejection of xenografts by this antibody. Opsonization of the destroyed
tumour cell membranes by anti-Gal IgG further targets them to APC, thus
converting the tumour lesion, treated by the a1,3GT
gene, into an in situ autologous tumour vaccineref.)
under the control of tissue-specific promoters : their apoptosis enhances
antigen uptake and presentation by professional APCs.
transfected with tumour antigen-encoding gene(s)
=> autologous
bone marrow transplantation (BMT)
+ Flt3
ligands + anti-CD40 mAb to overcome induction
of central tolerance, which could occur following repopulation of the thymus
with antigen-expressing DCs differentiated in vivo. This strategy
is more successful than administration of ex vivo generated transduced
DCs, resulting in the long-term survival of 50% of treated mice
mobilization into peripheral blood :,
which binds to CCR1 and CCR5ref-mobilized
peripheral blood monocytes as a cell source of DCs : previous investigations
have suggested that G-CSF-mobilized peripheral blood monocytes produce
reduced levels of proinflammatory cytokines such as IL-12 and TNF-a.
These Th1-type cytokines are thought to promote antitumor immune
response. The functional abilities of DCs generated from G-CSF-mobilized
monocytes obtained from 13 patients with CEA-positive advanced solid cancers
were assessed. Peripheral blood mononuclear cells were obtained from leukapheresis
products collected before and after systemic administration of G-CSF (subcutaneous
administration of high-dose [5-10 mg/kg] human
recombinant G-CSF for 5 consecutive days). In vitro cytokine production
profiles after stimulation with lipopolysaccharide (LPS) were compared
between monocytes with and without G-CSF mobilization. DCs generated from
monocytes were also examined with respect to cytokine production and the
capacity to induce peptide-specific T cell responses. Administration of
G-CSF was found to efficiently mobilize peripheral blood monocytes. Although
G-CSF-mobilized monocytes (G/Mo) less effectively produced Th1-type
cytokines than control monocytes (C/Mo), DCs generated from G/Mo restored
the same level of IL-12 production as that seen in DCs generated from C/Mo.
T cell induction assay using recall antigen peptide and phenotypic analyses
also demonstrated that DCs generated from G/Mo retained characteristics
identical to those generated from C/Mo. G-CSF mobilization can be used
to collect monocytes as a cell source for the generation of DCs for cancer
immunotherapy. DCs generated in this fashion were pulsed with HLA-A24-restricted
CEA epitope peptide and administered to patients safely; immunological
responses were induced in some patientsref
starting from hematopoietic
progenitor stem cells (HPSC)
using different long and short term culture systems :
starting from,
because monocytes represent a readily accessible source of DC precursors
and can easily be obtained in relatively large numbers from patients. Methods
for enrichment of monocytes collected in a leukapheresis procedureref
:
target was induced. Specific cytotoxicity was augmented when cultures were
boosted continuously with IL-2 (20 U/mL biological response modifier program)
plus tumor lysate stimulation. These results suggest that nonadherent PBMCs
may participate in enhancing DC maturation. Besides the simplicity of this
culture technique, bulk DC cultures potentially may be used with the same
efficiency as conventional purified DCs. Furthermore, bulk culture-derived
DCs may be used directly in vivo as a tumor vaccine, or for further
ex
vivo expansion of co-cultured CTLs to be used for adoptive immunotherapyref.
The number of DC obtained with 2% human serum albumin (HSA)-supplemented
cultures are consistently higher than in cultures with various concentrations
of autologous plasma or serum. DC prepared with HSA acquire the ability
to uptake dextran, and express high levels of MHC and co-stimulatory molecules
similar to DC cultured with autologous plasma or serum. Although DC cultured
with autologous plasma or serum consist of CD1a+ and CD1a-
populations, DC differentiated in the presence of HSA express CD1aref.
: CB and APB monocyte-derived ex vivo expanded DC express similar
DC markers CD83 (31.27+ 11.7% versus 34.0+ 5.2%, CB versus APB), CD1a (23.4+
4.2% versus 27.6+ 6.3%), and CD80 (21.97+ 12.01% versus 27.7+ 5.95). IHC
shows that cells with DC morphology express CD1a but not CD14. Neither
FLT-3L nor TGF-b
enhance DC expansion. Addition of 10% autologous plasma to CB cultures
promotes greater cell survival and a 150% increase in CD1a+/CD80+
cell recovery. CB DC were 62% as effective stimulators of adult allogeneic
T-cells as APB DC (p < .05) in allogeneic MLRref.
Cord blood mononuclear cells were cultured in serum-free medium in the
presence of GM-CSF, SCF and Flt-3 ligand (FL), to generate DC from their
precursors, and thereafter transfected with the eGFP reporter gene by electroporation.
Maturation of DC was induced by stimulation with GM-CSF, SCF, FL and phorbol
myristate acetate (PMA). Transfected DC strongly expressed EGFP, but transfection
efficiency of DC, defined as HLA-DR+ cells lacking lineage-specific
markers, did not exceed 2.5%. Expression of the reporter gene was also
demonstrated in the DC generated from transfected, purified CD34+
cord blood cells, by stimulation with GM-CSF, SCF, FL, and TNF-a.
Transfection of CD34+ cells was very efficient, but proliferation
of the transfected cells was much reduced as compared to the untransfected
cells. Therefore, the yield of transgene-expressing DC was relatively low.
In conclusion, nonviral transfection of cord blood DC proved feasible,
but considering the requirements for immunotherapy in cancer patients,
transfection of differentiated DC or generation of DC from transfected
hematopoietic stem cells provide only a limited number of DC expressing
the transgeneref
is based on culture of adherent cells in medium containingref
:
... at 37°C, 5% CO2. The addition of GM-CSF + IL-4 at the
onset of culture proved disadvantageous and is, therefore, delayed for
24 h. DC development from apheresis cells occurs faster than from fresh
blood or buffy coat, and is complete after 6-7 daysref.
and IL-3
for differentiation and the TLR3
ligand poly I:C for activation before pulsing with the TSA. Such a NA17.A2-loaded
DC vaccine was well-tolerated in 8 stage III or IV melanoma with only minor
and transient flu-like symptoms and inflammatory reactions at the injection
sites. In most patients, isotopic imaging documented DC migration from
the intradermal injection site to the draining lymph nodes. Finally, mixed
lymphocyte-peptide culture under limiting dilution conditions followed
by tetramer labeling indicated that 3 out of 8 patients mounted a CD8 T
cell response against the NA17.A2 antigenic peptideref
cells are poorly immunogenic and inhibit T-cell function. AML-derived
dendritic cells (AML-DCs) have better antigen-presentation capacity
than undifferentiated leukemic blasts, but may not be fully competent to
stimulate T cells previously inhibited by leukemic cells. IL-12
produced by AML-DCs plays a critical role in counteracting the inhibitory
activity of LCs on T-cell function. IL-12 gene can be successfully expressed
into AML-DCs defective in endogenous IL-12 production by using a novel
nonviral method that does not modify their phenotypical, cytogenetic, and
functional features. Genetically modified AML-DCs restore a near normal
T-cell functionref.
Serum levels of ANA
were examined with an ELISA kit. One patient with gastric carcinoma (patient
1, DC/tumour-fusion vaccine alone), one patient with breast cancer (patient
2, DC/tumour-fusion vaccine alone) and one patient with ovarian cancer
(patient 3, DC/tumour-fusion vaccine + rhIL-12) showed significant elevations
of serum ANA levels during treatment. In patient 1 malignant ascitic effusion
resolved and serum levels of tumour markers decreased. Patients 2 and 3
remained in good physical condition during treatment for 24 and 9 months,
respectively. Immunoblot analysis indicated antibody responses to autologous
tumour cells after vaccination in patient 2. None of the treated patients
showed clinical symptoms suggesting autoimmune disease. Patients with elevated
serum levels of ANA had significantly longer treatment periods than those
without it. Elevated serum levels of ANA after DC/tumour-fusion cell vaccine
might be associated with anti-tumour immune response induced by the vaccinationref.alone
significantly suppresses the growth of primary 4T1 murine mammary tumors
and the incidence of lung metastases in the prophylactic setting but is
less effective against pre-established tumors. However, celecoxib administered
after primary tumor establishment synergizes with tumor lysate-pulsed DC
and the adjuvant, GM-CSF,
to improve the antitumor immune response by suppressing primary tumor growth
and markedly reducing the occurrence of lung metastases. This triple combination
therapy elicits a tumor-specific immune response evidenced by elevated
IFN-g and IL-4 secretion by CD4+
T cells and results in increased infiltration of CD4+ and CD8+
T cells to the tumor site. In addition, dietary celecoxib inhibits angiogenesis
evidenced by decreased vascular proliferation within the tumor and serum
vascular endothelial growth factor levelsref
freely traverse membrane barriers. NY-ESO-1, an immunogenic antigen by
itself highly expressed in 60% of high-risk myeloma patients, was purified
to near homogeneity both alone and as a recombinant fusion protein with
a PTD, derived from HIV-Tat. Efficient entry of PTD-NY-ESO-1 into DCs,
confirmed by microscopy, Western blotting, and intracellular flow cytometry,
was achieved without affecting dendritic cell phenotype. Experiments with
amiloride, which inhibits endocytosis, and N-acetyl-l-leucinyl-l-norleucinal,
a proteasome inhibitor, confirmed that PTD-NY-ESO-1 entered dendritic cells
by protein transduction and was degraded by the proteasome. Tetramer analysis
indicated superior generation of HLA-A2.1, CD8+ T lymphocytes
specific for NY-ESO-1157-165 with PTD-NY-ESO-1 compared with
NY-ESO-1 control protein (44% versus 2%, respectively). NY-ESO-1-specific
T lymphocytes generated with PTD-NY-ESO-1 secreted IFN-g
indicative of a Tc1-type cytokine response. Thus, PTD-NY-ESO-1
accesses the cytoplasm by protein transduction, is processed by the proteasome,
and NY-ESO-1 peptides presented by HLA class I elicit NY-ESO-1-specific
T lymphocytesref
(obtainable even when only tiny amounts of tumour tissue are available)
by DOTAPref
is superior to lipofection and passive pulsing of mRNA and to electroporation
of plasmid cDNA. 0.5 mL of the cell suspension is mixed with 20 mg
of IVT mRNA, and electroporated in a 0.4-cm cuvette using an Easyject Plus
device (EquiBio, Kent, United Kingdom)ref1,
ref2.
Alternatively cells are electroporated in 2-mm cuvettes (200 ml
of DC (5 x 106 cells) at 300 V for 500 ms
using an Electro Square Porator ECM 830 (BTX, San Diego, CA). The amount
of in vitro transcribed RNA used was 2 mg/106
cellsref.
::
still a challenge because of low transduction efficiency and gene silencing
in DCs. An efficient method to prepare such DCs by in vitro differentiation
of hematopoietic progenitor cells transduced with chicken ovalbumin (OVA)
cDNA via the gene-silencing-resistant retroviral vector GCDNsap
packaged in vesicular stomatitis virus G (VSV-G) protein has been established.
When c-KIT+/lineage- cells were transduced with OVA
followed by expansion and differentiation, more than 90% of mature DCs
expressed the transgene. Mice inoculated with those cells completely rejected
the OVA-expressing tumor E.G7-OVA, and the anti-tumor effects were stronger
than those observed in mice inoculated with the same number of OVA peptide-pulsed
DCs. The mice harbored more cytotoxic T lymphocytes (CTLs) against E.G7-OVA
and produced antibody against OVA, suggesting the generation of multiple
CTLs recognizing different OVA epitopes and OVA-specific CD4+
T cells. Successive inoculations of the transduced DCs in a therapeutic
setting eradicated preexisting E.G7-OVA and prevented the progression of
retransplanted tumors. Thus, this vaccine therapy may represent a potent
immunotherapeutic approach for various malignant tumors that express suitable
TAAsref.
: lentivirally transduced DC have been shown to present antigenic peptides,
prime transgene-specific T cells in vitro and elicit a protective
cytotoxic T-lymphocyte (CTL) response in animal modelsref
: one of the major advantages of these vectors was reported to be the absence
of deleterious immune responses following gene transfer. However, recent
studies have shown that AAV vectors elicit humoral and cellular responses
against the transgene productsref.,
previously reported to enhance AAV gene expression. rAAV transduction of
freshly purified MO followed by 7 days of culture with cytokines to generate
DCs, and subsequent sorting for coexpression of DC markers CD1a and CD40,
shows robust transgene expression as well as evidence of nuclear localization
of the rAAV genome in the DC population. Phenotypic analyses using multiple
markers and functional assays of one-way allogeneic MLRs indicates that
rAAV-transduced MO-derived DCs are as equivalent to nontransduced DCsref.)
(this is required for presentation by CD40-B cells as they do not appear
to efficiently process and present exogenous Ags through the class I MHC
pathway). The poly-epitope response is usually more effective than the
response against a single epitope, as peptides may represent mutant as
well as wild-type portions of the protein. APC transduced with viral vectors
have been reported to have impaired functions in terms of stimulatory capacity.
: this would result in autocrine DC stimulation in vivo after re-injecion.
in culture. Co-culture of DCs with autologous T cells leads to an increase
in both the number of Tregs, as well as the expression of FOXP3 protein
per cell both in healthy donors and myeloma patients. DC mediated expansion
of FOXP3high Tregs is enhanced by endogenous but
not exogenous IL-2, and DC-T cell contact, including the CD80/86 membrane
costimulatory molecules. DCs also stimulate the formation of Tregs
from CD25- T cells. The efficacy of induction of Tregs
by DCs depends on the nature of the DC maturation stimulus, with inflammatory
cytokine treated DCs (Cyt-DC) being the most effective Treg
inducers. DCs induce Tregs using cells from both healthy
donors as well as myeloma patients, and the induced Tregs effectively
suppress T cell responses in the mixed leukocyte reaction (MLR). A single
injection of Cyt-DCs led to rapid enhancement of FOXP3+ Tregs
in vivo in 3/3 myeloma patients. These data reveal a role for DCs in
increasing the number of functional FOXP3high Tregs
in humansref.
ref
cells that generate prostatic
acid phosphatase : D9902B
/ Sipuleucel-T (APC8015)ref
(Provenge©). As part of a Phase III trial for FDA approval
they gave the vaccine treatment to 82 men with metastatic prostate cancer
unresponsive to hormone therapy, while 45 got a placebo treatment. On average
the men who got the vaccine lived 26 months, while those who got a placebo
lived 22 months (18% longer). 3 years later, 34% of vaccine patients were
still alive, compared 11% of unvaccinated patients. A phase III study was
undertaken to evaluate the safety and efficacy of sipuleucel-T in a placebo-controlled
study. A total of 127 patients with asymptomatic metastatic hormone refractory
prostate cancer (HRPC) were randomly assigned in a 2:1 ratio to receive
three infusions of sipuleucel-T (n = 82) or placebo (n = 45) every 2 weeks.
On disease progression, placebo patients could receive APC8015F, a product
made with frozen leukapheresis cells. Of the 127 patients, 115 patients
had progressive disease at the time of data analysis, and all patients
were followed for survival for 36 months. The median for time to disease
progression (TTP) for sipuleucel-T was 11.7 weeks compared with 10.0 weeks
for placebo (P = .052, log-rank; hazard ratio [HR], 1.45; 95%CI, 0.99 to
2.11). Median survival was 25.9 months for sipuleucel-T and 21.4 months
for placebo (P = .01, log-rank; HR, 1.70; 95%CI, 1.13 to 2.56). Treatment
remained a strong independent predictor of overall survival after adjusting
for prognostic factors using a Cox multivariable regression model (P =
.002, Wald test; HR, 2.12; 95%CI, 1.31 to 3.44). The median ratio of T-cell
stimulation at 8 weeks to pretreatment was eight-fold higher in sipuleucel-T-treated
patients (16.9 v 1.99; P < .001). Sipuleucel-T therapy was well tolerated.
While the improvement in the primary end point TTP did not achieve statistical
significance, this study suggests that sipuleucel-T may provide a survival
advantage to asymptomatic HRPC patients. Supportive studies are underwayref.
(APC8024©)
: APC-80200 (Mylovenge©)
and colorectal cancer
: Eladem© : autologous dendritic cells pulsed with recombinant
human PSAref))))
: phase II trial is starting in November 2004. The patients will receive
4 initial immunizations over a 6 week period with 1.25 mg HER-2 protein
AutoVac formulated in Alhydrogel adjuvant followed by booster immunizations
every 4 weeks for up to 26 weeks. The trial will be carried out at 5-10
cancer centers in Poland and Hungary. Pharmexa is also currently preparing
an application for another phase II trial with the vaccine formulated in
a stronger adjuvant (an immunostimulatory substance contained in all vaccines).
This trial is also planned to take place in Poland and Hungary and to include
up to 50 patients. The first trial, which includes up to 50 breast cancer
patients, will start in Hungary and is expected to conclude by mid 2006.)
to mature
DCs
: DC-poietins
increase protection in mice on immunizationrefref
gene either linked to an Ag-encoding generef
as a bicistronic plasmidref
or as a separate plasmid mixed with the Ag encoding plasmidref1,
ref2,
ref3,
ref4,
ref5,
ref6.
It has a paracrine effect at the injection site that recruits a mixed cellular
infiltrate including neutrophils, macrophages, immature DCs, eosinophils
(and T ad B cellsref1,
ref2,
ref3)
capable of recognizing tumour Ags at metastatic sites : separation of injections
with pGM-CSF and Ag-expressing plasmid plasmid into different sites does
not enhance immune responseref.
injected locally or systemicallyref1,
ref2,
ref3,
ref4,
ref5
: systemic toxicityref and flaring of autoimmune diseases have
been reportedref1, ref2.)
and G-CSFR,
was shown to be highly effective in expanding DC in vivo. In an
HLA-A2.1 transgenic mouse system, ProGP-4-generated DC were found to be
approximately equivalent in presenting a CTL peptide to a CTL line in
vitro compared with bone marrow (BM)-derived DC and >20-fold more efficient
than macrophages or B cells, and >100-fold better than BM-DC, macrophages,
or B cells at presenting PADRE, a universal helper T cell epitope, to a
T cell clone. The heightened epitope presentation by ProGP-4 DC was paralleled
in vivo inasmuch as a >6-fold increase in CTL induction was observed
compared with other APC populations following ex vivo loading with
peptide. The in vitro and in vivo CTL responses stimulated
by ProGP-4 DC could be further augmented by either culturing with TNF-a
or co-loading with PADRE. Peptide-loaded ProGP-4-generated DC demonstrate
potent antigenicity and immunogenicity for CTL, making them an attractive
component of epitope-based vaccinesref.
:
ligand(s)
to the cytokine cocktail further enhances the yield, maturation, migratory
and immunostimulatory capacity of the DC generated. More importantly, culture
conditions also influence the outcome of the T cell response induced.
/IL-1b/IL-6
or MCM alone induce CD4+ T cells that release intermediate levels
of IFN-g
and no IL-4
or IL-10.
Production of IFN-g
is significantly induced by addition of PGE2,
while no effect on production of IL-4
or IL-10
is observed.low,
IL-4neg
cells, TNF-a/IL-1b/IL-6promoted
growth of IFN-gintermediate,
IL-44neg
CD8+ T cells. Addition of PGE2
again only further polarized this pattern enhancing IFN-g
production by alloreactive CD8+ T cells in both cultures without
inducing type 2 cytokines./IL-1b/IL-6
can substitute for MCM and that addition of PGE2further
enhances the yield and quality of DC generated. TNF-a/IL-1b/IL-6
+ PGE2-cultured
DC seem to be optimal for generation of IFN-gintermediate-producing
CD4/CD8+ T cellsref.
Yet, despite rigorous criteria used for their characterization in vitro,
it is not clear whether the chosen conditions for maturation result in
DC with optimal in vivo functions, including migration to lymph
nodes and activation of Th1 CD4+ and CD8+
CTL. The above-reported ex vivo maturation protocol is suboptimal
and leads to a partially matured DC, which is refractory to optimal maturation
stimuli encountered in situ, such as imiquimod or Adjuprimeref.
In addition, reagents used for DC maturation using this or other protocols
can be expensive and not always readily available for clinical use.
co-culture of day 7 monocyte-derived DC generated with GM-CSF and IL-4
induces increased numbers of CD40+54+80+83-.
Also, IFN-a maturation leads to an increase
in IP-10 and MCP-1 chemokine secretion, but only a minor increase in IL-12p40
secretion. In line with this, maturation with IFN-a
has only a small effect on induction of autologous T-cell stimulatory capacity
of the DC. However, an increase in DC allogeneic T-cell stimulatory capacity
was observed. These data suggest that IFN-a
has a potential as a maturation agent used in DC-based cancer vaccine trials,
but not as a single reagentref
ligand(s)
cream can act as an adjuvant for gene
gun
DNA vaccination as good as rGM-CSFref.
When administered by subcutaneous injection at the vaccination site immediately
after particle-mediated immunotherapeutic delivery (PMID) of plasmid DNA
using a gene gun, resiquimod produces a similar Th1
biased immune response with a 10-fold reduced dose compared with imiquimodref:
resiquimod 50 nM moderately enhances IFN-g production
per cell (as measured by a peptide-based ELISpot assay), and produces a
several-fold increase in the stimulation index of antigen-specific T-cell
proliferation, increased antibody titer, and strong Th1 cytokine
biasrefmarkedly
enhances the targeting of antigen to DEC-205+ DCs but fails
to produce sustained antigen-specific immunity because of the failure to
activate DCs in vivo. The addition of an activating CD40-specific
antibody to anti-DEC-205-antigen conjugate did result in sustained immunity,
showing the importance of combining MHC-targeting and APC-activation strategies.
and CD70 / CD27L
along with an allogenic MHC class I (H-2Kd) molecule expression
on melanoma cells (B16F10, H-2b) to favor the antitumor immune
response. B16F10 tumor growth slows significantly when CD40L and CD70 are
coexpressed by tumor cells and the association with the allogenic molecule
(H-2Kd) enhances this effect. Growth kinetics of mock and CD40L-CD70-H-2Kd-expressing
B16F10 tumors in immunocompetent versus nu/nu and beige mice suggested
that CD8+ T lymphocytes and NK cells were involved in this antitumor
immunity. A delay in mock tumor growth was observed when CD40L-CD70-H-2Kd-expressing
B16F10 cells and mock tumor cells were injected simultaneously and contralaterally.
It was also shown that in vivo immunization of immunocompetent mice with
CD40L-CD70-H-2Kd B16F10 tumor cells improved the generation
of cytotoxic lymphocytes against the wild-type melanoma cells expressing
the syngenic MHC Class I molecule H-2Kb (B16K1). These observations
lay a path for new immunotherapeutic trials using semiallogenic fibroblasts
expressing costimulatory molecules and tumor-associated antigensref.
secreted by DCs and pulsed with synthetic peptides subserve the DC
function in MHC class I-restricted, peptide-specific CTL priming in
vitro and in vivoref
with E1-, E3-deleted adenoviral vector
would enhance the T cell-mediated cellular immune response with a consequent
induction of anti-tumor immunity to suppress tumor growth. To prove this
hypothesis, established tumors of different mouse cancer cells (B16-F10
melanoma, H-2b, and Colon-26 colon adenocarcinoma, H-2d)
were treated with intratumoral injection of bone marrow-derived DC that
had been modified in vitro with an RGD fiber-mutant adenovirus vector
expressing mouse fractalkine (Ad-FKN). In both tumor models tested, treatment
of tumor-bearing mice with Ad-FKN-transduced DC gave rise to a significant
suppression of tumor growth along with survival advantages in the treated
mice. Immunohistochemical analysis of tumors treated with direct injection
of Ad-FKN-transduced DC demonstrated that the treatment prompted CD8+
T cells and CD4+ T cells to accumulate in the tumor milieu,
leading to activation of immune-relevant processes. Consistent with the
finding, the intratumoral administration of Ad-FKN-transduced DC evoked
tumor-specific CTLs, which ensued from in vivo priming of Th1
immune responses in the treated host. In addition, the anti-tumor effect
provided by intratumoral injection of Ad-FKN-transduced DC was completely
abrogated in CD4+ T cell-deficient mice as well as in CD8+
T cell-deficient mice. These results support the concept that genetic modification
of DC with a recombinant fractalkine adenovirus vector may be a useful
strategy for cancer immunotherapy protocolsref.
,
and head and neck
cancersref
molecules into recombinant DNA and viral vaccine vectors for antigen-specific
vaccination that directly transduce or infect APCs. Even though DCs naturally
express B7 molecules, the increased level of expression of B7 as well as
altered patterns or ratios of expression of the different B7 family members
could significantly modify the outcome of T-cell priming in vivo
(iCD40) receptor permits temporally controlled, lymphoid-localized, DC-specific
activation. iCD40 is comprised of a membrane-localized cytoplasmic domain
of CD40 fused to drug-binding domains. This allows it to respond to a lipid-permeable,
high-affinity dimerizer drug while circumventing ectodomain-dependent negative-feedback
mechanisms. These modifications permit prolonged activation of iCD40-expressing
DCs in vivo, resulting in more potent CD8+ T-cell effector
responses, including the eradication of previously established solid tumors,
relative to activation of DCs ex vivo (P < 0.01), typical
of most clinical DC protocols. In addition, iCD40-mediated DC activation
exceeded that achieved by stimulating the full-length, endogenous CD40
receptor both in vitro and in vivo. Because iCD40 is insulated
from the extracellular environment and can be activated within the context
of an immunological synapse, iCD40-expressing DCs have a prolonged lifespan
and should lead to more potent vaccines, perhaps even in immune-compromised
patientsref,
TNF-a,
and IL-12
between mice and humans, with mice tolerating 50-300-fold higher serum
concentrations. So, doses of these cytokines that induce tumour regression
in mice are lethal in humans. Indeed the primary motivation is to maximize
the expression of the cytokine at the site of antigen delivery. Considering
that the cytokine gene-transduced tumour vaccines are typically administered
intradermally, subcutaneously or intramuscularly, whereas T-cell priming
generally occurs in the draining lymph node, it is not surprising that
tumour-cell vaccines engineered to produce APC-targeted (particularly DC-targeted)
cytokines might ultimately be more effective in priming immune responses.
Targeting of cytokines to areas of tumour metastasis, to which T lymphocytes
would subsequently traffic and carry out their effector functions, is achieved
by chimeric antibody-cytokine fusion molecules, in which cytokines such
as IL-2
are linked to the Fc regions of antitumour antibodies. Incorporation of
genes encoding T lymphocyte costimulatory or proliferative cytokines into
recombinant DNA or viral vaccines quantitatively enhance T-cell responses
and alters the differentiation pattern of antigen-specific T lymphocytes.
ligands : in syngeneic mice, ectopic expression of NKG2D ligands causes
NK cell-mediated rejection of transfected tumor cellsref1,
ref2.
It also primes cytotoxic T cells (CTLs), which are responsible for rejecting
subsequent challenges with tumor cells lacking NKG2D ligand expressionref.
Engagement of the murine NKG2D receptor (by using a DNA vaccine encoding
one of its ligands, H60) enhanced innate and adaptive immune responses
induced by a survivin-based DNA vaccineref.
This, in turn, augmented the vaccine's antitumor efficacy in prophylactic
and therapeutic settings against tumors of different origin and NKG2D expression
levelsref.
Engaging the NKG2D receptor markedly improved the efficacy of a survivin-based
DNA vaccine, delivered by attenuated S typhimurium. The combination
vaccine encoding both the NKG2D ligand H60 and survivin activates both
innate and adaptive antitumor immunity, resulting in better protection
against tumors of different origin and NKG2D expression levels. The enhanced
vaccine efficacy is in part attributable to increased crosstalk between
lymphocytes. Depletion of CD8 T cells during priming reduces the vaccine-induced
activation of DCs and NK activity. Depletion of NK cells during priming
leads to reduced DC activation and CTL activity. However, depletion of
CD4+ T cells results in activation of DCs, NK and CD8+
T cells and enhances NK cell activity. The pH60/survivin vaccine also increases
DC and NK cells, but decreases CD4+ T cells homing to Peyer's
patches, presumably as a result of changes in the their homing receptor
profile. Thus, by preferentially activating and attracting positive regulators,
and reducing negative regulators in Peyer's patches, this dual function
DNA vaccine induces a microenvironment more suitable for NK cell activation
and T cell primingref.
inhibitors prevent Trp starvation.
Inevitably, this approach will restrict DC traffic and, therefore, confine
the vaccine effect to one or a few regional LNs. Unexpectedly, the state
of maturation at the time of injection had no influence on the proportion
of DC that localized to draining lymph nodes, as labeled immature and mature
DC were detected in equal numbers. Immature DC that trafficked to lymph
nodes underwent a significant up-regulation of CD86 expression indicative
of spontaneous maturation. Moreover, immature DC exited completely from
the dermis within 36 h of injection, whereas mature DC persisted in large
numbers associated with a marked inflammatory infiltrate. In vitro
maturation is not a requirement for effective migration of DC in vivo
and administration of Ag-loaded immature DC that undergo natural maturation
following injection may be preferred for DC-based immunotherapyref.
Intradermal immunization with MART-1 peptide-pulsed DCs results in an increase
in circulating IFN-g-producing, antigen-specific
T cells, but the frequency of these cells did not correlate with responseref.
Intradermal administration results in about a 3-fold higher migration to
lymph nodes than subcutaneous administration, while mDC showed, on average,
a 6-to 8-old higher migration than iDC. The first DC are detected in lymph
nodes 20-60 min after inoculation and the maximum concentration was reached
after 48-72 href.)
that respond by cytokine secretion to peptide-pulsed targets and de
novo DTH compared with other routes in melanomaref1,
ref2,
ref3
and cutaneous
T-cell lymphoma (CTCL)ref
patients, indicating that IN may be the preferred ROA for mature DC vaccinesref.
Inevitably, this approach will restrict DC traffic and, therefore, confine
the vaccine effect to one or a few regional LNs.ref.
The inability of mature DCs to home to LNs from the blood is an important
obstacle to the use of antigen-pulsed DCs as vaccines. . DC vaccination
might be improved if DCs could be manipulated to home to LNs throughout
the body after intravenous injection. For this, DCs were retrovirally transformed
with a chimeric E-selectin-L-selectin molecule - a substitute for L-selectin
- that recognizes PNADref.
This molecule was chosen because expression of L-selectin does not change
when DCs are retrovirally transfected with the full-length molecule, presumably
because the ectodomain is proteolytically removedref.
Importantly, L-selectin is clustered on the tips of microvilli on leukocytes,
which greatly enhances its ability to form tethers under high shear flowref.
Because E-selectin-L-selectin fusion protein contains the transmembrane
and intracellular domain of L-selectin, it is also targeted to tips of
microvilli. The ectodomain derived from E-selectin is not cleaved, mediates
slow rolling of DCs through PNAD interactions, and enables DCs to extravasate
directly from blood through LN HEVsref.
So by reconstituting a missing component of the LN-homing cascade,
it is possible to target circulating leukocytes to the LNs
blockade
blockade
responses, but these may measure CD4 T cells instead of CD8 T cells
assays allow identification and quantification of numbers of cells producing
cytokines such as IFN-g when encountering target
antigens, but cytokine production may not correlate with tumor cell killing
may not correlate directly with tumor burden
to estimate the number of circulating tumor cells in peripheral blood
: patients with cancer in whom cancer vaccines are presently being tested
are, almost without exception, of advanced age (65-80 years), many decades
after the thymus has stopped producingnaive T lymphocytes. Therefore, the
generation of an effector-cell population in response to a vaccine depends
on the recognition of the vaccine antigen by one or more memory cells in
the T-cell repertoire of the patient. In mouse models, it can be clearly
shown that young mice make stronger primary responses than old mice. There
is an important discrepancy between preclinical studies in mouse models
and clinical trials of cancer vaccines : few studies, if any, use old mice.
Those that do, report an age-related increase in susceptibility to cancer
due to changing patterns of T-cell subsets, as well as difficulty in the
induction of effective antitumour immune responses. Hence elderlies could
need more adjuvant(s).
in first remission. Humoral antigen- and tumor-specific responses were
detectable but delayed, and they correlated with peripheral blood B-cell
recovery. In contrast, vigorous CD4+ and CD8+ antitumor
Th1 cytokine responses were induced in most individuals in the
absence of circulating B cells. Analysis of relapsing tumors showed no
mutations or change in expression of target antigen to explain escape from
therapy. These results show that severe B-cell depletion does not impair
T-cell priming in humans. Based on these results, it is justifiable to
administer vaccines in the setting of B-cell depletion; however, vaccine
boosts after B-cell recovery may be necessary for optimal humoral responsesref.
is a Th1-type
cytokine that has demonstrated potential as a biological adjuvant in murine
tumor models. The anti-tumor potential and mechanism of action of simultaneous
Apoptin and IL-18 gene transfer in C57BL/6 mice bearing Lewis lung carcinoma
(LLC). The growth of established tumors in mice immunized with pAPOPTIN
in conjunction with pIL-18 was significantly inhibited compared with the
growth of tumors in mice immunized with the empty vector (EV) or pAPOPTIN
alone. Furthermore, the immunization of mice with pAPOPTIN in conjunction
with pIL-18 elicited strong NK activity and LLC tumor-specific CTL responses
in vitro. In addition, T cells from lymph nodes of mice vaccinated
with pIL-18 or pAPOPTIN + pIL-18 secreted high levels of the Th1
cytokine IL-2 and IFN-g, indicating that the
regression of tumor cells is related to a Th1-type dominant
immune response. These results demonstrate that vaccination with Apoptin
together with IL-18 may be a novel and powerful strategy for cancer immunotherapyref.
,
mucin-1
and a triad of costimulatory molecules (TRICOM), which include B7.1, ICAM-1
and LFA-3. Vaccination is administered by subcutaneous injection followed
by 4 days of local recombinant adjuvant granulocyte-macrophage colony-stimulating
factor at the vaccination site. The vaccine has been developed for patients
with advanced pancreatic cancer and has now entered a randomized Phase
III clinical trial. Currently in phase III studyref.
19 peptide sequences with HLA-A*0201 motifs were selected by computer-based
algorithms. 5 peptides (KDR82-90, KDR288-297, KDR766-774, KDR1093-1101,
or KDR1035-1044) stimulated specific cytotoxic T lymphocytes (CTLs) from
PBMCs of three HLA-A*0201 donors. The decapeptide KDR288-297 was efficient
in sensitizing target cells for recognition by a CTL clone at a concentration
of 10 nM. More importantly, KDR288-297-specific CTLs lyzed target cells
transfected with HLA-A2/KDR cDNAs and a range of HLA-matched KDR+
angiogenic endothelial cells (aECs) and recognized CD34+ endothelial
progenitor cells as well. The specificity of CTLs was further confirmed
by tetramer assay and cold-target inhibition assay. In addition, ex vivo
exposure of aECs to the inflammatory cytokines enhanced CTL reactivity,
which is in keeping with upregulated KDR and HLA class I expression. In
Matrigel assays, recognition of aECs by specific CTLs triggered an anti-vascular
effect. These findings provide the first proof of the antigenic property
of KDR protein, and may be useful for devising new immunotherapeutic approaches
to human cancersref.
(stabilises pericyte-endothelial interactions during angiogenesis and is
highly expressed on endothelium during several diseases, including arthritis,
age-related macular degeneration and cancer) : a region unique to Tie-2
(amino acids 1-196) that is homologous in humans and mice. Using computer
algorithms, several HLA-A*0201 epitopes that are identical in mice and
humans were predicted within this region; however, binding assays showed
that the majority of these epitopes were of low affinity. Modification
of the anchor residues of 4 epitopes enhanced HLA binding. These epitopes
linked via alanines to a known helper epitope from HBVc128-140
(TPPAYRPPNAPILAAFLPATLTMV) were incorporated by site-directed mutagenesis
into a Tie-2 DNA construct. As FcR has been shown to increase antigen uptake
and delivery to both class I and II a DNA Fc-fusion vaccine using the amino
acids 1-196 of Tie-2 was designed. Immunisation of HLA*0201 transgenic
mice with one of the modified Tie-2 constructs stimulated CTLs that recognised
both wild-type and modified peptide-pulsed target cells. In contrast, no
CTLs were generated in mice immunised with wild-type Tie-2 construct, demonstrating
that the modified epitope was necessary in the generation of CTLs. Moreover,
CTLs from mice immunised with the modified construct killed HLA-A*0201
endothelial cells overexpressing Tie-2ref.
fusion vaccine to break the immune tolerance of self antigens associated
with angiogenesis. PCR amplification of mature b-defensin
2 (Def) and extracelluar segment of Vascular Endothelial Cadherin (Cad)
was conducted. (GGGS)3 was used as linker peptide for fusion
plasmid(Def-Cad). All the 3 segments were inserted into pSecTag2B plasmid
(pSec), and CT26 tumor cells were transfected. Expression was verified
by RT-PCR and chemotaxis assay. The alginate bead model was used to study
the antiangiogenic effect of fusion plasmid vaccination. All constructions
were verified by sequencing and were found expressed in mammalian cell.
The b-defensin 2 fusion protein had similar
capacity to chemoattract DCs as compared with nonfusion protein (P>0.05).
The alginate bead assay revealed that the tumor-induced angiogenesis in
fusion plasmid immunized mice was significantly suppressed when compared
with that in nonfusion plasmid (P<0.01). An active immunization strategy
based on b-defensin 2 fused with angiogenesis
related antigen of endothelial can inhibit angiogenesis and may be a useful
approach for cancer therapyref.
or MMP-14.
To enhance the immunogenicity of the mRNA-translated FAP product, a lysosomal
targeting signal derived from LAMP-1
was fused to the COOH terminus of FAP to redirect the translated product
into the class II presentation pathway. DCs transfected with mRNA encoding
the FAP-LAMP fusion product stimulated enhanced CD4+ and CD8+
T-cell responses. The growth of solid tumors is accompanied by tissue remodeling
that appears to involve interactions between the tumor cells and the stromaref.
The reactive stromal fibroblasts are not transformed, but they are distinguished
from normal fibroblasts by their production of certain extracellular matrix
proteins and proteasesref1,
ref2.
Among the latter, FAP appears to be consistently expressed as a cell-surface
protein in the stroma of malignant solid tumorsref.
The presence of FAP in 2 malignant melanoma cell linesref1,
ref2
also suggests that FAP expression can be induced in the cancer cell itself
as a result of malignant transformation. FAP is a serine protease that
is closely related to another type II integral membrane protein, CD26/DPP-IVref1,
ref2.
The catalytic site in CD26/DPP-IV and FAP contains the characteristic catalytic
triad of Ser630/624, Asp708/702, His740/734
(residues are numbered according to human CD26/DPP-IV and FAP respectively),
and the active serine is situated in a nucleophile elbow in the sequence
Gly-Trp-Ser-Tyr-Glyref1,
ref2,
ref3.
Both enzymes cleave NH2-terminal dipeptides from polypeptides
with the sequence NH2-Xaa-Pro/Ala, where Xaa can be any natural amino acidref1,
ref2,
ref3.
In addition, FAP was found to possess gelatinase activity in vitro,
and this observation gave rise to the notion that the enzyme might play
a role in tissue remodeling by digesting type I collagen of the extracellular
matrixref1,
ref2.
Rat CD26/DPP-IV has also been reported to possess gelatinase activityref,
and although this activity is seldom attributed to CD26/DPP-IV, the ability
of CD26/DPP-IV to form a complex with FAP at invadopodia of migratory fibroblasts
has suggested a possible role for the coordinated activity of both enzymesref
. The amino boronic dipeptide, Val-boro-Pro (PT-100), appeared to be an
interesting drug candidate in this context. PT-100 competitively inhibits
the dipeptidyl peptidase (DPP) activity of FAP and CD26/DPP-IV, and there
is a high-affinity interaction with the catalytic site due to the formation
of a complex between Ser630/624 and the boron of PT-100ref1,ref2.
The potential of PT-100 as an antitumor agent was intriguing in the light
of our earlier observation that PT-100, apparently through its interaction
with FAP, could stimulate hematopoiesis via the increased production of
cytokines [including G-CSF] both in vitro in stromal cell supported
human bone marrow cultures and in vivo in miceref.
The cytokines involved in the stimulation of hematopoiesis by PT-100 are
also known to promote innate as well as T-cell-mediated antitumor responses.
In particular, G-CSF gene transfection of the colon adenocarcinoma C-26
cell line has been shown to inhibit tumor take in mice and cause regression
of an established tumor in sublethally irradiated mice inoculated with
the transfected cellsref
. The release of G-CSF by the tumor cells appeared to promote tumor infiltration
by polymorphonuclear leukocytes expressing IL-1b
and TNF-a mRNA followed, sequentially, by macrophages
and T cells. PT-100 abrogates tumor growth as a single agent and augments
antibody-dependent cell-mediated cytotoxicity. Similarly to hematopoietic
stimulation, the antitumor activity did not require an interaction with
CD26. The data suggest that, via the up-regulation of cytokine and chemokine
expression in both the tumor and the draining lymph nodes, PT-100 can stimulate
immune responses against tumors involving both the innate and adaptive
branches of the immune systemref.
Mice immunized against FAP using DCs transfected with FAP mRNA inhibit
growth of melanoma, carcinoma, and lymphoma models and the magnitude of
the antitumor response was comparable to that of vaccination against tumor
cell-expressed antigens. Both s.c. implanted tumors and lung metastases
were susceptible to anti-FAP immunotherapy. The antitumor response could
be further enhanced by augmenting the CD4+ T-cell arm of the
anti-FAP immune response, achieved by using a lysosomal targeting sequence
to redirect the translated FAP product into the MHC class II presentation
pathway, or by covaccination against FAP and a tumor cell-expressed antigen,
tyrosinase-related protein 2 (TRP2). No morbidity or mortality was associated
with anti-FAP vaccination except for a small delay in wound healingref.
Fibroblasts are also the primary source of collagen type I, which contributes
to decreased chemotherapeutic drug uptake in tumors and plays a significant
role in regulating tumor sensitivity to a variety of chemotherapies. To
specifically kill tumor-associated fibroblasts, an oral DNA vaccine targeting
FAP, which is specifically overexpressed by fibroblasts in the tumor stroma,
was constructed. Through CD8+ T cell-mediated killing of tumor-associated
fibroblasts, the vaccine successfully suppressed primary tumor cell growth
and metastasis of multidrug-resistant murine colon and breast carcinoma.
Furthermore, tumor tissue of FAP-vaccinated mice revealed markedly decreased
collagen type I expression and up to 70% greater uptake of chemotherapeutic
drugs. Most importantly, pFap-vaccinated mice treated with chemotherapy
showed a 3-fold prolongation in lifespan and marked suppression of tumor
growth, with 50% of the animals completely rejecting a tumor cell challenge.
This strategy opens a new venue for the combination of immuno- and chemotherapiesref.
and cancer immunotherapyref
vaccine (Norelin©; source :
YM
BioSciences, Inc.) for androgen-dependent
prostate
adenocarcinoma
: a double-chain miniprotein with each chain containing 3 linear repeats
of the self-peptide gonadotropin-releasing hormone (GnRH3),
the hinge region of human IgG1 (hinge), and a T-helper epitope
from the measles virus protein (MVP). The GnRH3-hinge-MVP miniprotein was
conjugated to purified recombinant Hsp65 of Mycobacterium bovis and
used to immunize rats primed with subcutaneous injections of BCG in the
absence of adjuvantsref
vaccine : CTP37 (Avicine©;
source : AVI BioPharma, Inc./SuperGen
Corporation) contains 37 amino acids from the carboxyl terminus of
hCG. Vaccination result in 2 distinct antibody responses to separate epitopes
within the peptideref
immunogen (G17DT) (Gastrimmune©; source : Aphton
Corp.) neutralises the gastrin 17 hormone and the Gly-G17 hormone.
Gastrin 17 is a growth factor for pancreatic, stomach and colorectal
carcinomas,
and a potent stimulator of gastric acid secretion. The anti-gastrin immunogen,
G17DT, consists of a large carrier protein, called diptheria toxoid (DT).
A synthetic peptide, which is similar to a portion of the gastrin 17 hormone
(GT), is attached to the carrier protein. When administered to patients,
G17DT induces an immune response producing antibodies, which cross-react
and neutralise the target hormone thus preventing its interaction with
disease-causing or -participating cells : the antibodies both reduced G17
stimulated gastric acid secretion and inhibited gastrin from interacting
with the CCK-2 receptor. Therapeutic efficacy of both passive and active
immunisation with G17DT has been established in a number of tumour systems
including both primary and metastatic disease. Furthermore, additive effects
with 5-fluorouracil (5-FU)/leucovorin have been confirmed in both colon
and gastric tumour models. Phase I/II studies in advanced gastrointestinal
(GI) malignancies have shown no systemic or autoimmune reactions to active
immunisation with G17DT. Use of an optimised dose has yielded a high proportion
of responders (> 80%), with minimal side effects and antibody titres measurable
within 2-4 weeks. Taken together these results suggest that the G17DT immunogen
is a promising agent for the treatment of GI cancer and Phase III trials,
currently underway, will definitively evaluate this early promiseref.
Aphton has entered into an agreement with Aventis Pasteur for the marketing
of G17DT in all human cancer indications in North America and Europe. Under
the terms of the agreement, Aphton is responsible for product development,
clinical trials and regulatory agency approvals. The agreement was originally
with Pasteur Merieux Connaught, a subsidiary of Rhone-Poulenc. However,
in December 1999, Rhone-Poulenc merged with Hoechst to form Aventis. As
a result of the merger, Pasteur Merieux Connaught underwent a name change
to Aventis Pasteur. In July 2002, Aphton announced that it would negotiate
with companies wanting to licence the vaccine in territories other than
North America and Europe. In February 2003, Aphton announced it had received
fast track designation for G17DT in combination with cisplatin
and fluorouracil
for use in stage IV gastric cancer to improve overall survival. In July
2002, the US FDA granted G17DT orphan drug status for the treatment of
gastric
cancerref,
an indication that was broader than the indication Aphton originally sought.
The vaccine was also granted orphan drug status in Australia for gastric
cancer in December 2002. In July 2002, Aphton announced that the US FDA
had granted G17DT orphan drug status for the treatment of adenocarcinoma
of the pancreas. In September 2002, the US FDA also granted G17DT, used
in combination with gemcitabine,
fast track status for the treatment of pancreatic
cancer
patientsref.
In addition, the vaccine was also granted orphan drug status in Australia
for pancreatic cancer in December 2002. In March 2003, Aphton announced
that the Committee for Orphan Medicinal Products had recommended to the
European Commission that G17DT be granted orphan drug status for both pancreatic
and gastric cancer in the EU. Aphton expects this will be approved and
ratified within a few weeks. In August 2002, Aphton filed an IND with the
US FDA, as well as the European equivalent, to conduct clinical trials
for patients suffering from gastro-esophophageal
reflux disease (GERD).
The trials will assess, among other endpoints, the efficacy of G17DT in
providing symptomatic relief to patients with GERD. In November 2002, Aphton
announced that it had received approval to initiate a clinical trial in
Europe. Additionally, Aphton has agreed with the FDA to conduct toxicology
and other preclinical studies prior to initiation of a trial in the US.
Patient recruitment for the European study was scheduled to begin in January
2003ref
vaccines currently are tested on men whose cancers are growing and no longer
responsive to hormone therapy, so that < 20% of men respond to vaccines
aloneref.
To date, the cancer vaccines that have been tested definitively in randomized
phase 3 clinical trials did not show statistically significant disease-free
or overall survival, the ultimate test of clinical efficacyref.
Clinical studies using adoptive transfer of T cells activated and expanded
ex
vivo
have unequivocally demonstrated proof of principle that cancer
patients possess tumor-specific T cells capable of mediating anticancer
activity
in vivo, as measured by regression of metastatic tumorsref1,
ref2.
Translation of these findings into a standardized, reproducible and exportable
therapy is still many years away. First and foremost, the natural state
of endogenous tumor-reactive T cells appears to be characterized by general
hyporesponsiveness or anergy. This is likely to result from a number of
mechanisms that tumors utilize to induce antigen-specific immune toleranceref1,
ref2.
Although a number of the newer-generation vaccines can effectively transfer
antigen to and activate dendritic cells in vivo, T cell tolerance
remains a major barrier that is difficult to overcome by therapeutic vaccination.
Preclinical models demonstrate that for poorly immunogenic tumors, therapeutic
vaccines alone are ineffective at curing animals with a significant tumor
burden, particularly once tolerance has been established. Combinations
of cancer vaccines administered in conjunction with inhibitors of immunologic
checkpoints and agonists for TLRs or T cell costimulatory pathways can
overcome tolerance and generate significant antitumor responses even in
cases of metastatic cancerref1,
ref2,
ref3.
Treatment of patients with an antagonist antibody to a powerful immunologic
checkpoint, CTLA-4, resulted in antitumor responses in a number of melanoma
patients, but was associated with a high incidence of severe autoimmune
responsesref.
These and other studies emphasize that antigen-driven responses such as
vaccines will need to be combined with manipulation of costimulatory and
checkpoint pathways to achieve sufficient therapeutic potency with
acceptable toxicity. Because various of these costimulatory pathways and
immunologic checkpoints work at distinct points in the evolution of the
effector immune response, it is likely that the most potent antitumor immune
responses will be generated by combination strategies that affect multiple
regulatory points. Complex whole cell-derived vaccines have given clinically
superior responses compared to vaccines containing well-defined antigens,
such as peptides or gangliosides; however, well-defined vaccines are theoretically
more desirable because of their reproducibility. Therapeutic cancer vaccines
are expected to boost an already existing, albeit weak, immune response
rather than prime new responses : if the existing response was primed against
a tumour that originated at a mucosal site, this response might be more
effectively boosted by a mucosal rather than systemic route of immunization.
Barriers to cancer vaccine development :
,
which has a 2-year mortality rate of >97%. This lack of division based
on disease severity can lead to an imbalance in risk-benefit judgments
in regulating the development of novel approaches to treat lethal cancers.This effort should clearly be integrated with RAID's mission of developing
academically held immunotherapeutics using the BRB as a resource for coordination
of the production of these materials. Such an enterprise cannot be successful
unless its budget is at least doubled and its physical plant significantly
upgraded. These increases represent a tiny fraction of the total NCI budget
and would go a long way to fulfilling the stated mission: significant reduction
in cancer morbidity and mortality over the next decade. Resources might
also come through the NIH Roadmap for Medical Research, as this endeavor
is clearly aligned with the NIH's vision of creating translationally relevant
core technologies and making their outputs broadly available to the community.
The FDA must take responsibility for better educating itself and the oncology
community regarding the cutting-edge immunotherapeutics currently being
developed. This would be most efficiently accomplished by significantly
expanding the scope of BRMAC's composition and advisory role. In addition,
the FDA could develop an online database of immunotherapeutics that covers
mechanisms of action, potential and actual toxicities, contacts (within
academia, companies and the FDA) with expertise in a particular area and
regulatory points-to-consider for investigational new drug applications.
This would alleviate much of the confusion and misperceptions about regulatory
issues regarding specific classes of agents. As suggested above, the FDA
has great power to facilitate the development of combinations of experimental
biologic agents by promoting the message that combinatorial experimental
therapies with a sound scientific basis are encouraged rather than discouraged.
This message can only come with the confidence of a well-educated FDA that
communicates more effectively with both academia and industry. In the case
of cancer therapeutics, creation of separate centers for the evaluation
of therapeutics for acutely lethal diseases may facilitate a more accommodating
atmosphere for novel combinatorial immunotherapeutics by reorientation
of risk-benefit judgments. Only with concerted measures such as these will
the tremendous opportunities that immunologic science has provided over
the past decade be effectively tapped for the sake of the most effective
therapy for patients with cancer.
Thus far, the emphasis has been on the activation of one arm of the
immune system, in the majority of cases T-cell immunity, since T cells
are believed to be the major players in tumor control. The available experimental
evidence argues that activation of multiple arms of the immune system is
a must to enhance the ability of immunotherapy to control tumor growth