Cancer Urotherapy – An Update
Joseph
Eldor, MD
Theoretical
Medicine Institute
P.O.Box 12142, Jerusalem, 91120,Israel
1. Cancer antigens
2. Cancer vaccines
3. Cancer immunotherapy
4. Dendritic cells
5. Cancer antibodies
6. Genetic immunotherapy
7. Adjuvant immunotherapy
8. Cancer urotherapy
Malignant tumors express antigens that may stimulate and serve as targets for
antitumor immunity. Virally induced tumors usually contain integrated proviral
genomes in their cellular genomes and often express viral genome-encoded
proteins that may stimulate specific host immune responses. Antigens unique to
individual tumors that stimulate specific rejection of transplanted tumors have
been demonstrated only in experimental animals. Other tumor antigens that
potentially can stimulate immune responses are shared by different tumors.
These include products of mutated or rearranged oncogenes or tumor-suppressor
genes. Tumors may also overexpress tissue differentiation antigens or embryonic
antigens, which also have the potential to be recognized by the immune system.
The recent identification of tumor antigens recognized by cytotoxic T cells
opens up new possibilities for constructing chemically defined antigens for
specific immunotherapy. Treatment of malignant tumors in humans by immunologic
approaches, although theoretically attractive, has not yet succeeded on a large
scale. Important progress in immunotherapy of cancer is emerging with several
different treatment modalities (1).
Recent studies have identified new melanoma antigens that are recognised by
CD4(+) T cells. Analysis of tumour-specific CD4(+) T-cell responses may lead to
the development of optimal anti-cancer vaccines that can induce an orchestrated
effort of tumour-specific CD4(+) and CD8(+) T cells in the fight against cancer
(2).
T cells play an important role in in vivo rejection of human melanoma. Human
melanoma antigens recognized by autologous T cells were identified. These
antigens are classified as tissue (melanocyte)-specific proteins, cancer-testis
antigens (proteins expressed in normal testis and various cancers),
tumor-specific peptides derived from mutations in tumor cells, and others. A
variety of mechanisms generating T cell epitopes on tumor cells were discovered.
Various clinical observations, including tumor regression observed in adoptive
transfer of gp100-reactive T cells suggest that these identified melanoma
peptides may function as tumor rejection antigens. Immunodominant common
epitopes that could expand melanoma-reactive cytotoxic T lymphocytes (CTLs) in
vitro were found in the MART-1 and gp100 antigens. New immunization
protocols--including immunization with peptides, recombinant viruses, plasmid
DNAs, and dendritic cells pulsed with peptides as well as adoptive transfer of
in vitro-generated CTLs by stimulation with antigenic peptides--were developed
(phase I clinical trials have been performed in the Surgery Branch of the
National Cancer Institute, Bethesda, MD, U.S.A.). Immunization with the gp100(209(210M))
peptide that was modified to have high HLA-A2 binding affinity, along with
incomplete Freund's adjuvant and interleukin (IL)-2, resulted in a 42% response
rate in patients with melanoma. These immunotherapies need further improvement
due to the mechanisms of tumor escape from T cell responses (3).
Most major advances in human cancer immunology and immunotherapy have come from
studies in melanoma. We are beginning to understand the immune repertoire of T
cells and antibodies that are active against melanoma, with recent glimpses of
the CD4(+) T cell repertoire. The view of what the immune system can see is
extending to mutations and parts of the genome that are normally invisible (4).
Pancreatic cancer is the fifth leading cause of cancer deaths in the
The immune repertoire contains T cells and B cells that can recognize
autologous cancer cells. This repertoire is directed against self, and in some
cases altered self (mutations). Priming immune responses against self antigens
can be difficult. Strategies are presented using altered self to elicit
immunity against self in poorly immunogenic tumor models. Mechanisms underlying
immunity to self antigens on cancer cells show that the immune system can use
diverse strategies for cancer immunity, in both the immunization and the
effector phases. CD4+ T cells are typically, but not always, required for
immunization. The effector phase of tumor immunity can involve cytotoxic T
cells, macrophages with activating Fc receptors, and/or killer domain
molecules. This diversity in the effector phase is observed even when
immunizing with conserved paralogs. A consequence of tumor immunity is
potentially autoimmunity, which may be undesirable. Autoimmunity uses similar
mechanisms as tumor immunity, but tumor immunity and autoimmunity can uncouple.
These studies open up strategies for active immunization against cancer (6).
Cancer Antigens
The spectrum of human antigens allows a monitoring of various pathological
processes such as autoimmune disorders and tumorigenesis. Serological analysis
of cDNA expression libraries (SEREX) is now used to search for new
cancer-associated antigens, which are potential diagnostic markers or targets
for immunotherapy of cancer (7).
The
immune response can effectively hamper the progression of preclinical stages of
tumor growth. Medicine in the postgenomic era offers an increasing possibility
of detecting healthy individuals at risk of developing cancer who could benefit
from tumor-preventive vaccines. The identification of novel tumor antigens that
fulfill two conditions will be crucial for the development of cancer
immunoprevention. First, an ideal antigen should have a crucial pathogenetic
role in tumor growth to avoid the selection of antigen-loss variants. Second,
the antigen should be recognizable by the immune system even in MHC-loss
variants and should therefore be recognized both by antibodies and T cells.
Identifying such antigens will also provide new targets for cancer
immunotherapy (8).
Cancer/testis (CT) antigens are a category of tumor antigens with normal
expression restricted to male germ cells in the testis but not in adult somatic
tissues. In some cases, CT antigens are also expressed in ovary and in
trophoblast. In malignancy, this gene regulation is disrupted, resulting in CT
antigen expression in a proportion of tumors of various types. Since their
initial identification by T-cell epitope cloning, the list of CT antigens has
been greatly expanded through serological expression cloning (SEREX) and
differential mRNA expression analysis, and approximately 20 CT antigens or
antigen families have been identified to date. Characteristics commonly shared
by CT antigens, aside from the highly tissue-restricted expression profile,
include existence as multigene families, frequent mapping to chromosome X,
heterogeneous protein expression in cancer, likely correlation with tumor
progression, induction of expression by hypomethylation and/or histone
acetylation, and immunogenicity in cancer patients. Spontaneous humoral and
cell-mediated immune responses have been demonstrated against several CT
antigens, including NY-ESO-1, MAGE-A, and SSX antigens. Since CT antigens are
immunogenic and highly restricted to tumors, their discovery has led directly
to the development of antigen-specific cancer vaccines, and clinical trials
with MAGE-A and NY-ESO-1 are in progress (9).
Our understanding of how immune responses are generated and regulated drives
the design of possible immunotherapies for cancer patients. Cancer vaccines
that are able to induce tumor-specific immune responses in cancer patients are
not always followed by tumor rejection. Two possible reasons that might explain
this dichotomy of cancer immunology. First, the immune response generated,
although detectable, may not be quantitatively sufficient to reject the tumor.
Second, the tumor microenvironment may modulate tumor cell susceptibility to
the systemic immune response induced by the immunization (10).
Cytolytic T lymphocytes (CTL) play a major role in the recognition and
destruction of tumor cells by the immune system. Some of these antigens,
including those encoded by the MAGE genes, are absent on all normal cells, and
therefore constitute ideal targets for cancer vaccines aimed at increasing the
activity of anti-tumor lymphocytes. Such vaccines are currently tested in
clinical trials with melanoma patients. These antigens consist of small
peptides that are presented by HLA molecules and that result from the
degradation of intracellular proteins. This degradation is performed by an
intracellular proteolytic complex called the proteasome. Dendritic cells, which
in the lymph node are responsible for antigen presentation to the lymphocytes
in order to initiate the immune response, are inefficient to produce some
peptides because they contain a different proteasome called
"immunoproteasome" (11).
One of the most significant advances in the field of modern tumor immunology is
the identification of genes encoding tumor-rejection antigens that are
recognized by human leukocyte antigen (HLA) class I-restricted and
tumor-specific cytotoxic T lymphocytes (CTLs). Several peptides encoded by
these genes are now under clinical trial as cancer vaccines, and major tumor
regression has been observed in some melanoma patients. These results indicate
that identification of the peptides capable of inducing CTLs may provide a new
modality of cancer therapy. Itoh et al. (12) investigated tumor-rejection
antigens from epithelial cancers, and reported 7 genes encoding tumor-rejection
antigens and peptides available for specific immunotherapy of HLA-A26 or -A24
patients with epithelial cancers. Furthermore, they identified more than 10 genes encoding
tumor-rejection antigens and peptides available for specific immunotherapy of
HLA-A2 patients with epithelial cancers. Therefore these new antigens and
peptides could be applicable to the treatment of numerous epithelial cancer
patients.
Cytotoxic T-cell responses to shared tumor antigens have been characterized for
several tumor types, and the MHC-associated peptides that comprise these
antigens have been defined at a molecular level. These provide new tools to
determine whether immune responses can be generated with these tumor antigens,
and there are data to suggest that such immune responses can be generated.
However, it is also clear that tumor cells can evade immune responses directed
against some shared antigens, by downregulating expression of MHC or of the
antigenic protein(s), as well as by more active methods such as secretion of
immunosuppressive cytokines. Awareness of these mechanisms of immune escape
will help to direct development of the next generation of tumor vaccines.
Targeting unique antigens and modulating the cytokine environment likely will
be critical to comprehensive vaccine systems in the future (13).
The adoptive transfer of tumor-infiltrating lymphocytes along with interleukin
2 into autologous patients resulted in the objective regression of tumor in
about 30% of patients with melanoma, indicating that these T cells play a role
in tumor rejection. To understand the molecular basis of the T cell-cancer cell
interaction Wang (14) and others started to search for tumor antigens expressed
on cancer cells recognized by T cells. This led to the identification of
several major histocompatibility complex (MHC) class I restricted tumor
antigens. These tumor antigens have been classified into several categories:
tissue-specific differentiation antigens, tumor-specific shared antigens, and
tumor-specific unique antigens. Because CD4+ T cells play a central role in
orchestrating the host immune response against cancer, infectious diseases, and
autoimmune diseases, a novel genetic approach has recently been developed to
identify these MHC class II restricted tumor antigens. The identification of
both MHC class I and II restricted tumor antigens provides new opportunities
for the development of therapeutic strategies against cancer.
In order to enhance cell mediated cytotoxicity, bispecific antibodies (BsAbs),
molecules combining two or more antibodies with different antigenic
specificities, have been developed as new agents for immunotherapy. Kudo et al.
(15) recent studies revealed that simultaneous administration of two kinds of
BsAbs (anti-tumor x anti-CD3 plus anti-tumor x anti-CD28) together with
lymphokine activated killer cells with a T cell phenotype (T-LAK cells)
inhibited growth of human xenotransplanted tumors in severe combined
immunodeficient (SCID) mice, while single BsAb was without effect. Three kinds
of BsAbs (anti-tumor x anti-CD3, anti-tumor x anti-CD28, anti-tumor x anti-CD2)
showed the highest cytotoxicity against tumor cells when given simultaneously
with T-LAK cells or peripheral blood mononuclear cells in vitro and in vivo.
BsAbs can be preserved for immediate application, while cytotoxic T lymphocytes
(CTLs) must be made-to-order, and are time-consuming to prepare. Tumor
associated antigens, such as MAGE antigens, SART antigens, MUC1 antigen, c-erbB
2 antigen or cancer/testis antigens can be served to target antigens for BsAb
production. By conjugation with antibodies to effector cells (anti-CD3,
anti-CD28, anti-CD16, anti-CD64, anti-CD89 or anti-CD2), many kinds of BsAbs
can be produced to cover most types of cancers from different organs. Therefore
this strategy might be ubiquitously applicable to most malignancies.
Melanogenesis-related proteins play important roles in melanin synthesis and
antigenicity of melanomas. Identification of highly expressed
melanoma-associated antigens (MAA) that are immunogenic in humans will provide
potential targets for cancer vaccines. Melanogenesis-related proteins have been
shown to be MAA. Autoantibody responses to these MAA have been shown to react
with melanoma cells and melanocytes, and suggested to play a role in
controlling melanoma progression. To assess antibody responses to potential
melanoma/melanocyte autoantigens, the open-reading frame sequences of
tyrosinase, tyrosinase-related protein (TRP)-1, TRP-2, and melanoma-associated
glycoprotein antigen family (gp100/pmel17) genes were cloned and expressed as
recombinant proteins in E. coli (16). Purified recombinant antigens were
employed to detect antibodies in sera of melanoma patients and normal healthy
donors. By affinity enzyme-linked immunosorbent assay and western blotting, all
recombinant antigens were shown to be antigenic. The main subclass of antibody
response to these antigens was IgG. Most importantly this study demonstrated
anti-TRP-2 and anti-gp100/pmel17 IgG responses in melanoma patients. Only one
of 23 normal donors had an antibody response to the antigens tested.
MAA-specific IgG antibodies in sera were assessed in melanoma patients (n = 23)
pre- and post-polyvalent melanoma cell vaccine treatment. Polyvalent melanoma
cell vaccine treatment enhanced anti-MAA antibody responses; however, only
anti-TRP-2 and anti-gp100/pmel17 antibody response was enhanced. These studies
suggest that four melanogenesis-related proteins are autoimmunogenic and can be
used as potential targets for active-specific immunotherapy.
The adoptive transfer of cytotoxic T lymphocytes (CTLs) derived from tumor-infiltrating
lymphocytes (TIL) along with interleukin 2 (IL-2) into autologous patients with
cancer resulted in the objective regression of tumor, indicating that these
CTLs recognized cancer rejection antigens on tumor cells. To understand the
molecular basis of T cell-mediated antitumor immunity, several groups started
to search for such tumor antigens in melanoma as well as in other types of
cancers. A number of tumor antigens were isolated by the use of cDNA expression
systems and biochemical approaches. These tumor antigens could be classified
into several categories: tissue-specific differentiation antigens,
tumor-specific shared antigens, and tumor-specific unique antigens. However,
the majority of tumor antigens identified to date are nonmutated, self
proteins. This raises important questions regarding the mechanism of antitumor
activity and autoimmune disease. The identification of human tumor rejection
antigens provides new opportunities for the development of therapeutic
strategies against cancer (17).
Cancer vaccines
Multiple novel immunotherapy strategies have reached the stage of testing in
clinical trials that were accelerated by recent advances in the
characterization of tumor antigens and by a more precise knowledge of the regulation
of cell-mediated immune responses. The key steps in the generation of an immune
response to cancer cells include loading of tumor antigens onto
antigen-presenting cells in vitro or in vivo, presenting antigen in the
appropriate immune stimulatory environment, activating cytotoxic lymphocytes,
and blocking autoregulatory control mechanisms. This knowledge has opened the
door to antigen-specific immunization for cancer using tumor-derived proteins
or RNA, or synthetically generated peptide epitopes, RNA, or DNA. The critical
step of antigen presentation has been facilitated by the coadministration of
powerful immunologic adjuvants, the provision of costimulatory molecules and
immune stimulatory cytokines, and the ability to culture dendritic cells. Advances
in the understanding of the nature of tumor antigens and their optimal
presentation, and in the regulatory mechanisms that govern the immune system,
have provided multiple novel immunotherapy intervention strategies that are
being tested in clinical trials (18).
The critical role of antigen-specific T cells in cancer immunotherapy has been
amply demonstrated in many model systems. Though success of clinical trials
still remains far behind expectation, the continuous improvement in our
understanding of the biology of the immune response will provide the basis of
optimized cancer vaccines and allow for new modalities of cancer treatment. The
future will mainly be concerned with allogeneic bone marrow cell
transplantation after non-myeloablative conditioning, because this approach
could provide a major breakthrough in cancer immunotherapy (19). Concerning
active vaccination protocols the following aspects will be addressed: i) the
targets of immunotherapeutic approaches; ii) the response elements needed for
raising a therapeutically successful immune reaction; iii) ways to achieve an
optimal confrontation of the immune system with the tumor and iv) supportive
regimen of immunomodulation. Many questions remain to be answered in the field
of allogeneic bone marrow transplantation after non-myeloablative conditioning
to optimize the therapeutic setting for this likely very powerful tool of
cancer therapy.
Active immunotherapy using dendritic cells (DCs) to deliver tumor antigens has
generated considerable excitement among oncologists worldwide. Although most
tumor antigens used in immunotherapeutic approaches are tumor-associated,
often, little is known about the underlying biology of the target. Antigen
expression is a prerequisite for tumor formation or maintenance by the use of
'obligate' tumor antigens. The prototype for this class of antigens is the p53
tumor antigen, which is mutated in > 50% of human malignancies. The direct
involvement of p53 in the malignant transformation of tumors makes it an
attractive target for immunotherapy. p53-Reactive antibodies have been found in
patients with various types of cancer, demonstrating that the human immune
system can recognize and respond to tumor-associated p53. Extensive preclinical
experimentation has now validated the translation of p53-expressing DCs into a
clinical setting. Clinical trials are ongoing to evaluate the safety and
antitumor responses elicited by DCs transduced with adenoviral-p53 in cancer
patients (20).
Tumor vaccination strategies have been increased over the past years. This
increase began with the identification of tumor antigens recognized by the
immune system. Better understanding of the immune system and increasing
knowledge about the antigen presentation process and the role of dendritic
cells have opened new therapeutic possibilities. DNA vaccines, already
successfully used against viral antigens and covering a broad repertoire of
epitopes, might also be of advantage in tumor immunotherapy. Design and
selection of vectors are of considerable importance for the vaccination. There
are three major types of DNA-based recombinant cancer vaccines: DNA from tumor
antigens can be used 1) to modify dendritic cells, 2) as 'naked' DNA-vaccine or
3) to construct recombinant viral vaccines (21).
It is now clear that many human tumor antigens can be recognised by the immune
system. These tumor antigens can be classified into several groups including
cancer-testis, differentiation, tissue specific, over-expressed, and
viral-associated antigens. In many cases, there is a known molecular basis of
carcinogenesis which provides the explanation for the differentiated expression
of these antigens in tumors compared with normal cells. Improved understanding
of the biology of the immune response, particularly of immune recognition and
activation of T-cells, allow better design of vaccines. Pre-clinical
comparative studies allow evaluation of optimal vaccine strategies which can
then be delivered to the clinic. Currently, a range of cancer vaccines are
being tested including those using tumor cells, proteins, peptides, viral
vectors, DNA or dendritic cells. Ultimately, this research should give rise to
an entirely new modality of cancer treatments (22).
The identification of antigens on tumor cells has led to significant
contributions to the field of immunotherapy. One of the most active areas under
investigation in cancer immunotherapy is the development of vaccines against
melanoma antigens. Induction of immunity against tumor antigens can follow
multiple routes using different mechanisms. Crucial to the development of
active immunization and other immunotherapies is the discovery and
understanding of the molecular identity of antigens and the mechanisms involved
in tumor immunity, as well as escape from immunity (23).
Antigenic differences between normal and malignant cells form the basis of
clinical immunotherapy protocols. Because the antigenic phenotype varies widely
among different cells within the same tumor mass, immunization with a vaccine
that stimulates immunity to a broad array of tumor antigens expressed by the
entire population of malignant cells is likely to be more efficacious than
immunization with a vaccine for a single antigen. One strategy is to prepare a
vaccine by transfer of DNA from the patient's tumor into a highly immunogenic
cell line. Weak tumor antigens, characteristic of malignant cells, become
strongly antigenic if they are expressed by immunogenic cells. In animal models
of melanoma and breast cancer, immunization with a DNA-based vaccine is
sufficient to deter tumor growth and to prolong the lives of tumor-bearing mice
(24).
Berd (25) has devised a novel approach to active immunotherapy based on
modification of autologous cancer cells with the hapten, dinitrophenyl (DNP).
The treatment program consists of multiple intradermal injections of
DNP-modified autologous tumor cells mixed with BCG. Administration of
DNP-vaccine to patients with metastatic melanoma induces a unique reaction - the
development of inflammation in metastatic masses. Histologically, this consists
of infiltration of T lymphocytes, most of which are CD8+. These T cells usually
produce gamma interferon in situ. Moreover, they represent expansion of T cell
clones with novel T cell receptor structures. Occasionally, administration of
DNP-vaccine results in partial or complete regression of measurable metastases.
The most common site of regression has been small lung metastases.
Administration of DNP-vaccine to patients in the post-surgical adjuvant setting
produces a more striking clinical effect. Berd et al. have treated 214 patients
with clinically evident stage III melanoma who had undergone lymphadenectomy.
With a median follow-up time of 4.4 years (1.8-10.4 years) the 5-year overall
survival (OS) rate is 47% (one nodal site = 51%, two nodal sites = 33%). These
results appear to be comparable to those obtained with high dose interferon.
More recent studies suggest that this therapeutic approach is also applicable
to ovarian cancer. There appear to be no insurmountable impediments to applying
this approach to much larger numbers of patients or to developing it as a
standard cancer treatment.
Certain anti-idiotypic antibodies that bind to the antigen-combining sites of
antibodies can effectively mimic the three-dimensional structures and functions
of the external antigens and can be used as surrogate antigens for active
specific immunotherapy. Extensive studies in animal models have demonstrated
the efficacy of these vaccines for triggering the immune system to induce
specific and protective immunity against bacterial, viral and parasitic
infections as well as tumors. Several monoclonal anti-idiotype antibodies that
mimic distinct human tumor-associated antigens have been developed and
characterized. Encouraging results have been obtained in recent clinical trials
using these anti-idiotype antibodies as vaccines (26).
Immunization with anti-idiotype (Id) antibodies represents a novel new approach
to active immunotherapy. Extensive studies in animal tumor models have
demonstrated the efficacy of anti-Id vaccines in preventing tumor growth and
curing mice with established tumor. Bhattacharya-Chatterjee
et al. (27) have developed and characterized several murine monoclonal
anti-Id antibodies (Ab2) which mimic distinct human tumor-associated antigens
(TAA) and can be used as surrogate antigens for triggering active anti-tumor
immunity in cancer patients.
Immunization with dendritic cells loaded with tumor antigens could represent a
powerful method of inducing antitumor immunity. Studies from several
laboratories have shown that immunization with dendritic cells pulsed with
specific antigens prime cytotoxic T-cells and engender tumor immunity. The
majority of cancer patients who lack an identified tumor antigen and/or cannot
provide sufficient tumor tissue for antigen preparation are excluded from
treatment with cancer vaccines based on using either specific tumor antigens or
mixtures of tumor-derived antigens in the form of peptides or proteins isolated
from tumor cells. Vaccination with the mRNA content of tumor cells would extend
the scope of vaccination to this group of patients as well because RNA can be
amplified from very few cancer cells (28).
The adoptive transfer of tumor-infiltrating lymphocytes (TIL) along with
interleukin (IL)-2 into autologous patients with cancer resulted in the
objective regression of tumor, indicating that T cells play an important role
in tumor regression. In the last few years, efforts have been made towards
understanding the molecular basis of T-cell-mediated antitumor immunity and
elucidating the molecular nature of tumor antigens recognized by T cells. Tumor
antigens identified thus far could be classified into several categories:
tissue-specific differentiation antigens, tumor-specific shared antigens and
tumor-specific unique antigens. CD4+ T cells play a central role in
orchestrating the host immune response against cancer, infectious diseases and
autoimmune diseases. The identification of tumor rejection antigens provides
new opportunities for the development of therapeutic strategies against cancer
(29).
Human tumors express a number of protein antigens that can be recognized by T
cells, thus providing potential targets for cancer immunotherapy. Dendritic
cells (DCs) are rare leukocytes that are uniquely potent in their ability to
present antigens to T cells, and this property has prompted their recent
application to therapeutic cancer vaccines. Isolated DCs loaded with tumor
antigen ex vivo and administered as a cellular vaccine have been found to
induce protective and therapeutic anti-tumor immunity in experimental animals.
In pilot clinical trials of DC vaccination for patients with non-Hodgkin's lymphoma
and melanoma, induction of anti-tumor immune responses and tumor regressions
have been observed. Additional trials of DC vaccination for a variety of human
cancers are under way, and methods for targeting tumor antigens to DCs in vivo
are also being explored. Exploitation of the antigen-presenting properties of
DCs thus offers promise for the development of effective cancer immunotherapies
(30).
Recently, cancer immunotherapy has emerged as a therapeutic option for the
management of cancer patients. This is based on the fact that our immune
system, once activated, is capable of developing specific immunity against
neoplastic but not normal cells. Increasing evidence suggests that
cell-mediated immunity, particularly T-cell-mediated immunity, is important for
the control of tumor cells. Several experimental vaccine strategies have been
developed to enhance cell-mediated immunity against tumors. Some of these tumor
vaccines have generated promising results in murine tumor systems. In addition,
several phase I/II clinical trials using these vaccine strategies have shown
extremely encouraging results in patients (31).
Animal studies have shown that vaccination with genetically modified tumor
cells or with dendritic cells (DC) pulsed with tumor antigens are potent
strategies to elicit protective immunity in tumor-bearing animals, more potent
than "conventional" strategies that have been tested in clinical
settings with limited success. While both vaccination strategies are forms of
cell therapy requiring complex and costly ex vivo manipulations of the
patient's cells, current protocols using dendritic cells are considerably
simpler and would be more widely available. Vaccination with defined tumor
antigens presented by DC has obvious appeal. However, in view of the expected
emergence of antigen-loss variants as well as natural immunovariation,
effective vaccine formulations must contain mixtures of commonly, if not
universally, expressed tumor antigens. When, or even if, such common tumor
antigens will be identified cannot be, predicted, however. Thus, for the
foreseeable future, vaccination with total-tumor-derived material as source of
tumor antigens may be preferable to using defined tumor antigens. Vaccination
with undefined tumor-derived antigens will be limited, however, by the
availability of sufficient tumor tissue for antigen preparation. Because the
mRNA content of single cells can be amplified, tumor mRNA, or corresponding
cDNA libraries, offer an unlimited source of tumor antigens. DC transfected
with tumor RNA were shown to engender potent antitumor immunity in animal
studies. Thus, immunotherapy using autologous DC loaded with unfractionated
tumor-derived antigens in the form of RNA emerges as a potentially powerful and
broadly useful vaccination strategy for cancer patients (32).
Cancer immunotherapy
Adoptive immunotherapy--the isolation of antigen-specific cells, their ex vivo
expansion and activation, and subsequent autologous administration--is a
promising approach to inducing antitumor immune responses. The molecular
identification of tumor antigens and the ability to monitor the persistence and
transport of transferred cells has provided new insights into the mechanisms of
tumor immunotherapy. Recent studies have shown the effectiveness of
cell-transfer therapies for the treatment of patients with selected metastatic
cancers. These studies provide a blueprint for the wider application of
adoptive-cell-transfer therapy, and emphasize the requirement for in vivo
persistence of the cells for therapeutic efficacy (33).
There is clear evidence that certain forms of immunotherapy can be successful
against certain cancers. However, it would appear that cancerous cells of
various origin are exceptionally adept at subverting the immune response.
Consequently, it is probable that the most efficacious therapy will be one in
which multiple responses of the immune system are activated. There is currently
an embarrassment of riches with regard to multiple vaccine strategies in the
clinic, although no single method seems to hold the solution (34).
Despite advances in chemotherapy and surgical techniques, patients with cancer
often develop local recurrence or metastatic spread. Recent advances in
molecular biology, coupled with new insights in tumor immunology, have led to
the design of novel antitumor vaccines. Poxviruses are a large family of DNA
viruses that provide an effective and safe vector system for vaccine
development. The poxvirus strategy has been successfully documented in animal
models, and has been used to express both tumor-associated antigens and immune
stimulatory molecules (35).
Prostate cancer is the most common malignancy in American men. Metastatic
prostate cancer is incurable, with the currently best treatment, androgen
ablation, being only palliative. Therefore, there is a need to develop new,
more effective therapies against this disease. Multiple immunotherapeutic
strategies are being explored for the treatment of prostate cancer, with the
hope that such treatment will be more effective and have fewer side effects
than current treatment options. Several immunotherapy strategies have been
shown to be effective against prostate tumors in animal models, and many of
these strategies are beginning to be tested in clinical trials for their
efficacy against human prostate cancer. It is likely that effective treatment
of prostate cancer will require the use of both immunotherapeutic and
traditional approaches in multimodality treatments. In addition, for
immunotherapy to be effective against prostate cancer, ways to overcome immune
evasion and immunosuppression by the tumor cells will need to be developed
(36).
Despite the identification of tumor antigens and their subsequent generation in
subunit form for use as cancer vaccines, whole tumor cells remain a potent
vehicle for generating anti-tumor immunity. This is because tumor cells express
an array of target antigens for the immune system to react against, avoiding
problems associated with major histocompatibility complex (MHC)-restricted
epitope identification for individual patients. Furthermore, whole cells are
relatively simple to propagate and are potentially efficient at contributing to
the process of T cell priming. However, whole cells can also possess properties
that allow for immune evasion, and so the question remains of how to enhance
the immune response against tumor cells so that they are rejected. Scenarios
where whole tumor cells may be utilised in immunotherapy include autologous
tumor cell vaccines generated from resected primary tumor, allogeneic
(MHC-disparate) cross-reactive tumor cell line vaccines, and immunotherapy of
tumors in situ. Since tumor cells are considered poorly immunogenic, mainly
because they express self-antigens in a non-stimulatory context, the
environment of the tumor cells may have to be modified to become stimulatory by
using immunological adjuvants. Recent studies have re-evaluated the relative
roles of direct and cross-priming in generating anti-tumor immunity and have
highlighted the need to circumvent immune evasion (37).
The Wilms tumor gene WT1 is expressed in leukemias and various kinds of solid
tumors, including lung and breast cancer, and exerts an oncogenic function in
these malignancies, suggesting that WT1 protein is a novel, overexpressed tumor
antigen. The WT1 protein, in fact, is an attractive tumor rejection antigen in
animal models. Stimulation in vitro of peripheral blood mononuclear cells with
HLA-A*2402--and HLA-A*0201--restricted 9-mer WT1 peptides elicits WT1-specific
cytotoxic T-lymphocytes (CTLs), and the CTLs kill endogenously WT1-expressing
leukemia or solid tumor cells. Furthermore, WT1 immunoglobulin M (IgM) and IgG
antibodies are detected in patients with hematopoietic malignancies such as
acute myeloid leukemia, chronic myeloid leukemia, and myelodysplastic
syndromes, indicating that WT1 protein overexpressed by leukemia cells is
indeed immunogenic. Taken together, these results demonstrate that WT1 protein
is a promising tumor antigen for cancer immunotherapy against leukemias and
various kinds of solid tumors, including lung and breast cancer (38).
In the last few years, a great deal of efforts have been directed towards
understanding the molecular basis of T cell-mediated anti-tumor immunity and
elucidating the molecular nature of tumor antigens recognized by T cells.
Identification of a number of major histocompatibility complex (MHC) class
I-restricted melanoma antigens has led to clinical trials aimed at developing
effective cancer vaccines. These studies showed some evidence of therapeutic
effect on the treatment of cancer, but the exclusive use of CD8+ T cells may
not be effective in eradicating tumor. This rekindles interest in the role of
CD4+ T cells in antitumor immunity, which play a central role in orchestrating
the host immune response against cancer. Thus, Wang et al. (39) have attempted to identify MHC class
II-restricted tumor antigens recognized by tumor-specific CD4+ T cells. The
identification of tumor rejection antigens provides new opportunities for the
development of therapeutic strategies against cancer.
Interleukin (IL)-2 and IL-15 are two cytokine growth factors that regulate
lymphocyte function and homeostasis. Early clinical interest in the use of IL-2
in the immunotherapy of renal cell carcinoma and malignant melanoma
demonstrated the first efficacy for cytokine monotherapy in the treatment of
neoplastic disease. Advances in our understanding of the cellular and molecular
biology of IL-2 and its receptor complex have provided rationale to better
utilize IL-2 to expand and activate immune effectors in patients with cancer.
Exciting new developments in monoclonal antibodies recognizing tumor targets
and tumor vaccines have provided new avenues to combine with IL-2 therapy in
cancer patients. IL-15, initially thought to mediate similar biological effects
as IL-2, has been shown to have unique properties in basic and pre-clinical
studies that may be of benefit in the immunotherapy of cancer (40).
Several recent developments have hallmarked progress in tumor immunology and
immunotherapy. The use of interleukin-2 (IL-2) in cancer patients demonstrated
that an immunological manipulation was capable of mediating the regression of
established growing cancers in humans. The identification of the genes encoding
cancer antigens and the development of means for effectively immunizing
patients against these antigens has opened important new avenues of exploration
for the development of effective active and cell-transfer immunotherapies for
patients with cancer (41).
A wide range of strategies in cancer immunotherapy has been developed in the
last decade, some of which are currently being used in clinical settings. The
development of these immunotherapeutical strategies has been facilitated by the
generation of relevant transgenic animal models. Since the different strategies
in experimental immunotherapy of cancer each aim to activate different immune
system components, a variety of transgenic animals have been generated either
expressing tumor associated, HLA, oncogenic or immune effector cell molecule
proteins (42).
Immunotherapy is in its infancy for many diseases, whether they are neoplastic
or autoimmune. The major issues for cancer immunotherapy today involve the
definition of molecular targets and the generation of effector mechanisms to
attack the targets of interest. Soft tissue sarcomas provide a unique
opportunity to examine the immune response against defined antigens. Many types
of sarcomas contain tumor-specific chromosomal translocations encoding fusion
proteins, which are attractive targets for immunotherapy. Our understanding of
the immune system is also coming into clearer focus with the discovery of
dendritic cells as powerful natural adjuvants and the teasing out of mechanisms
leading to immunity versus tolerance as examples. It is hoped that the
intersection of cellular immunology and sarcoma molecular biology will lead to
new modalities of therapy for this group of patients with this heterogeneous
group of diseases (43).
Studies of the administration of interleukin-2 to patients with metastatic
melanoma or kidney cancer have shown that immunological manipulations can
mediate the durable regression of metastatic cancer. The molecular
identification of cancer antigens has opened new possibilities for the
development of effective immunotherapies for patients with cancer. Clinical
studies using immunization with peptides derived from cancer antigens have
shown that high levels of lymphocytes with anti-tumor activity can be raised in
cancer-bearing patients. Highly avid anti-tumor lymphocytes can be isolated
from immunized patients and grown in vitro for use in cell-transfer therapies.
Current studies are aimed at understanding the mechanisms that enable the
cancer to escape from immune attack.
The idea that there
might be an immune response to cancer has been around for many years.
Immunotherapy has a long history, but is only rarely considered as the
treatment of choice. Immunotherapy has encountered a number of intrinsic
difficulties in cancer, such as the antigenic resemblance between the tumor and
normal cells, the rapid kinetic proliferation of tumor cells and their reduced
immunogenicity. There are various types of immunotherapy. Aspecific
immunotherapy augments the body' s immune response without targeting specific
tumoral antigens. In adoptive immunotherapy, cells are administered with
antitumoral reactivity to mediate neoplasm regression. Specific active
immunotherapy is based on the principle that neoplasm cells contain immunogenic
sites against which an antitumoral immune response can be induced in an attempt
to stimulate the immune system to target specific tumoral antigens. Vaccines
against cancer cells are based on a more precise identification of the tumoral
antigen components. Passive immunotherapy was limited by the difficulty of
obtaining high titering and specificity in early attempts using polyclonal
antisera; monoclonal antibodies are currently used alone or in association with
radioactive substances and cytotoxic agents. Enormous progress has been made
this century in the use of immunotherapy for cancer treatment. It seems likely
that the next century will see its increased afficacy, making it one of the
possible therapeutic options (45).
Despite major advances in our understanding of adaptive immunity and dendritic
cells, consistent and durable responses to cancer vaccines remain elusive and
active immunotherapy is still not an established treatment modality. The key to
developing an effective anti-tumor response is understanding why, initially,
the immune system is unable to detect transformed cells and is subsequently
tolerant of tumor growth and metastasis. Ineffective antigen presentation
limits the adaptive immune response; however, we are now learning that the
host's innate immune system may first fail to recognize the tumor as posing a
danger. Recent descriptions of stress-induced ligands on tumor cells recognized
by innate effector cells, new subsets of T cells that regulate tumor tolerance
and the development of spontaneous tumors in mice that lack immune effector
molecules, beckon a reflection on our current perspectives on the interaction
of transformed cells with the immune system and offer new hope of stimulating
therapeutic immunity to cancer (46).
Immunotherapy approaches to fight cancer are based on the principle of mounting
an immune response against a self-antigen expressed by the tumor cells. In
order to reduce potential autoimmunity side-effects, the antigens used should
be as tumor-specific as possible. A complementary approach to experimental
tumor antigen discovery is to screen the human genome in silico, particularly
the databases of "Expressed Sequence Tags" (ESTs), in search of tumor-specific
and tumor-associated antigens. The public databases currently provide a massive
amount of ESTs from several hundreds of cDNA tissue libraries, including
tumoral tissues from various types. Vinals et al. (47) described a novel method
of EST database screening that allows new potential tumor-associated genes to
be efficiently selected. The resulting list of candidates is enriched in known
genes, described as being expressed in tumor cells.
The concept of immunotherapy of cancer is more than a century old, but only
recently have molecularly defined therapeutic approaches been developed. The
identification of tumor antigens can now be accelerated by methods allowing the
amplification of gene products selectively or preferentially transcribed in the
tumor. However, determining the potential immunogenicity of such gene products
remains a demanding task, since major histocompatibility complex (MHC)
restriction of T cells implies that for any newly defined antigen,
immunogenicity will have to be defined for any individual MHC haplotype.
Tumor-derived peptides eluted from MHC molecules of tumor tissue are also a
promising source of antigen. Tumor antigens are mostly of weak immunogenicity,
because the vast majority are tumor-associated differentiation antigens already
'seen' by the patient's immune system. Effective therapeutic vaccination will
thus require adjuvant support, possibly by new approaches to immunomodulation
such as bispecific antibodies or antibody-cytokine fusion proteins.
Tumor-specific antigens, which could be a more potent target for immunotherapy,
mostly arise by point mutations and have the disadvantage of being not only
tumor-specific, but also individual-specific. Therapeutic vaccination will
probably focus on defined antigens offered as protein, peptide or nucleic acid.
Irrespective of the form in which the antigen is applied, emphasis will be
given to the activation of dendritic cells as professional antigen presenters.
Dendritic cells may be loaded in vitro with antigen, or, alternatively, initiation
of an immune response may be approached in vivo by vaccination with RNA or DNA,
given as such or packed into attenuated bacteria. The importance of activation
of T helper cells has only recently been taken into account in cancer
vaccination. Activation of cytotoxic T cells is facilitated by the provision of
T helper cell-derived cytokines. T helper cell-dependent recruitment of
elements of non-adaptive defence, such as leucocytes, natural killer cells and
monocytes, is of particular importance when the tumor has lost MHC class I
expression. Barriers to successful therapeutic vaccination include: (i) the
escape mechanisms developed by tumor cells in response to immune attack; (ii)
tolerance or anergy of the evoked immune response; (iii) the theoretical
possibility of provoking an autoimmune reaction by vaccination against
tumor-associated antigens; and (iv) the advanced age of many patients, implying
reduced responsiveness of the senescent immune system (48).
Generating an antitumor immune response in tumor-bearing host has been impaired
by several characteristics of both patient and tumor cells. Amongst those
requirements is the necessity of generating immune effectors that are specific
to tumor cells. The last two decades have seen the description of many so
called tumor "specific" antigens. Indeed, strictly specific tumor
antigens are scarce. Most antigens are tumor-associated antigens, also shared
by normal tissues. Telomerase and its activity have recently been recognized as
a major marker of tumoral growth in more than 80% of cancers. Some telomerase
subunits might be ideal, if not universal, targets to an antitumor immune
response in patients with cancer. Many other major parameters remain to be
understood and to be mastered (49).
The survival of patients with cancer has improved steadily but incrementally
over the last century, with the advent of effective anticancer treatments such
as chemotherapy and radiotherapy. However, the majority of patients with
metastatic disease will not be cured by these measures and will eventually die
of their disease. New and more effective methods of treating these patients are
required urgently. The immune system is a potent force for rejecting
transplanted organs or microbial pathogens, but effective spontaneous
immunologically induced cancer remissions are very rare. In recent years, much
has been discovered about the mechanisms by which the immune system recognizes
and responds to cancers. The specific antigens involved have now been defined
in many cases. Improved adjuvants are available. Means by which cancer cells
overcome immunological attack can be exploited and overcome. Most importantly,
the immunological control mechanisms responsible for initiating and maintaining
an effective immune response are now much better understood. It is now possible
to manipulate immunological effector cells or antigen-presenting cells ex vivo
in order to induce an effective antitumour response. At the same time, it is
possible to recruit other aspects of the immune system, both specific (e.g.
antibody responses) and innate (natural killer cells and granulocytes) (50).
Immunotherapy of cancer is entering into a new phase of active investigation
both at the pre-clinical and clinical level. This is due to the exciting
developments in basic immunology and tumor biology that have allowed a
tremendous increase in our understanding of mechanisms of interactions between
the immune system and tumor cells. Clinical approaches are diverse but can now
be based on strong scientific rationales. The analysis of the available
clinical results suggests that, despite some disappointments, there is room for
optimism that both active immunotherapy (vaccination) and adoptive
immunotherapy may soon become part of the therapeutic arsenal to combat cancer
in a more efficient way (51).
Advancements in the understanding of cellular immunity within the last decade,
along with the characterization of tumor antigens, have led to
immunotherapeutic approaches for cancer therapy. This mode of treatment is
expected to provide more tumor-specific activity, thereby being less toxic to
normal cells than standard modalities. Clinical trials are underway throughout
the world to determine whether immunotherapy is a practical and viable
alternative to conventional cancer therapies. Unlike radiotherapy and
chemotherapy, wherein tumor regression is the standard for determining efficacy
of the regimens, immunotherapy has to be evaluated by the examination of
several immunological parameters within patients (52).
The identification
of tumor-associated antigens recognized by cellular or humoral effectors of the
immune system has opened new perspectives for cancer therapy. Different groups
of cancer-associated antigens have been described as targets for cytotoxic T
lymphocytes (CTLs) in vitro and in vivo: 1) cancer-testis (CT) antigens, which
are expressed in different tumors and normal testis; 2) melanocyte
differentiation antigens; 3) point mutations of normal genes; 4) antigens that
are overexpressed in malignant tissues; and 5) viral antigens. Clinical studies
with peptides derived from these antigens have been initiated to induce
specific CTL responses in vivo. Immunological and clinical parameters for the
assessment of peptide-specific reactions have been defined, i.e., delayed-type
hypersensitivity (DTH), CTL, autoimmmune, and tumor regression responses.
Preliminary results demonstrate that tumor-associated peptides alone elicit
specific DTH and CTL responses leading to tumor regression after intradermal
injection. Granulocyte-macrophage colony-stimulating factor (GM-CSF) was proven
effective in enhancing peptide-specific immune reactions by amplification of
dermal peptide-presenting dendritic cells. Long-lasting complete tumor
regressions have been observed after induction of peptide-specific CTLs.
However, in single cases with disease progression after an initial tumor
response, either a loss of the respective tumor antigen targeted by CTLs or of
the presenting major histocompatibility complex (MHC) class I allele was
detected as a mechanism of immune escape under immunization. Based on these
observations, cytokines to enhance antigen and MHC class I expression in vivo
are being evaluated to prevent immunoselection. Recently, a strategy utilizing
spontaneous antibody responses to tumor-associated antigens (SEREX) has led to
the identification of a new CT antigen, NY-ESO-1, which is regarded as one of
the most immunogenic antigens known today inducing spontaneous immune responses
in 50% of patients with NY-ESO-1-expressing cancers. Clinical studies involving
antigenic constructs that induce both antibody and CTL responses will show
whether these are more effective for immunotherapy of cancer (53).
Tumors express proteins not commonly found in normal cells, or over-express
certain proteins. These may in some cases serve as target antigens for
immunological attack. It is therefore essential to improve our understanding of
the nature of these target epitopes and the cells which recognize them, in
order to develop immunotherapy as a realistic treatment for cancer (54).
Advances in molecular biology have enabled specific antigens present on
colorectal cells to be characterized, against which immune responses may be
generated. This, in combination with our inability to significantly alter
survival from this condition, has resurrected an interest in immunotherapy as a
potential treatment option. A number of approaches currently constitute
immunotherapeutic options for colorectal cancer. A number of treatment
modalities are already in phase III studies, although clearly not all will
fulfill their initial promise. Surgeons need to be aware of the advances in
this rapidly expanding field, and keep an open mind as to their efficacy (55).
The goal of harnessing the immune system to recognize tumor as
"nonself" is not new. Now, thanks to new knowledge and new
techniques, however, modalities that seek to activate the host immune system
are becoming increasingly feasible as treatments for advanced malignancies (56).
The major impact of recent scientific advances, such as the discovery of genes
and gene products, has been to facilitate development of immunotherapies based
on the specific stimulation of immune reactions against characterized tumor
antigens (57).
Over
the last decade, there has been a considerable increase in understanding of
immune responses against cancers, the antigenic structures on tumor cells
recognised by the immune system, and the development of more effective
vaccines. There is, however, very limited understanding of why the immune
system most often fails to control tumor growth and progression. In some
patients, it is difficult to demonstrate immune responses to their tumors, and
it may be assumed that this reflects poor recognition of tumor antigens,
induction of anergy in lymphocytes, or suppression of immune responses by
tumor-derived factors. In other patients, tumor progression appears to occur
despite the presence of antibody or cell-mediated responses. This may indicate
selection of tumor cells that have lost tumor antigens or HLA antigens by
immune responses against the tumor. Tumor cells may also become resistant to
mediators of apoptosis, such as Fas ligand and tumor necrosis factor-related
apoptosis-inducing ligand used by lymphocytes to kill tumor cells. It is
suggested that development of effective immunotherapy will need to include
strategies that take into account these limitations of immune responses and
classification of tumors according to the treatment approach most likely to
succeed (58).
Heat shock proteins (Hsps), ubiquitous in nature, act as chaperones for
peptides and other proteins. They have been implicated in loading immunogenic
peptides onto major histocompatibility complex molecules for presentation to T cells.
When isolated from tumor cells, Hsps are complexed with a wide array of
peptides, some of which serve as tumor-specific antigens. Animal studies have
demonstrated that heat shock protein--peptide complexes (HSPPCs) from tumor
cells can act as vaccines to prevent or treat tumors. Potent and specific tumor
antigens have long been the holy grail in cancer immunotherapy; HSPPCs from
tumor cells could become a safe and reliable source of tumor-specific antigens
for clinical application (59).
Immunotherapy of mice with preexisting cancers with heat shock protein
preparations derived from autologous cancer resulted in retarded progression of
the primary cancer, a reduced metastatic load, and prolongation of life-span.
Treatment with heat shock protein preparations derived from cancers other than
the autologous cancer did not provide significant protection. Spontaneous
cancers (lung cancer and melanoma), chemically induced cancers (fibrosarcoma
and colon carcinoma), and an ultraviolet radiation-induced spindle cell
carcinoma were tested, and the results support the efficacy of autologous
cancer-derived heat shock protein-peptide complexes in immunotherapy of cancers
without the need to identify specific tumor antigenic epitopes (60).
Dendritic
cells
The identification of tumor specific antigens has provided important advance in
tumor immunology. It is now established that specific cytotoxic T lymphocytes
(CTL) and natural killer cells infiltrate tumor tissues and are effector cells
able to control tumor growth. However, such a natural antitumor immunity has
limited effects in cancer patients. Failure of host defenses against tumor is
consecutive to several mechanisms which are becoming targets to design new
immunotherapeutic approaches. CTL are critical components of the immune
response to human tumors and induction of strong CTL responses is the goal of
most current vaccine strategies. Effectiveness of cytokine therapy, cancer
vaccines and injection of cells improving cellular immunity have been established
in tumor grafted murine models. Clinical trials are underway. Today, interest
is particularly focused on cell therapy: injected cells are either "ready
to use" effector cells (lymphocytes) or antigen presenting cells able to
induce a protective immune reaction in vivo (dendritic cells). The challenge
ahead lie in the careful optimization of the most promising strategies in
clinical situation (61).
The response of hepatocellular carcinoma (HCC) to therapy is often
disappointing and new modalities of treatment are clearly needed. Active
immunotherapy based on the injection of autologous dendritic cells (DC)
co-cultured ex vivo with tumor antigens has been used in pilot studies in
various malignancies such as melanoma and lymphoma with encouraging results. In
the study of Ladhams et al. (62), the preparation and exposure of patient DC to
autologous HCC antigens and re-injection in an attempt to elicit antitumor
immune responses were described. Therapy was given to two patients, one with
hepatitis C and one with hepatitis B, who had large, multiple HCC and for whom
no other therapy was available. No significant side-effects were observed. The
clinical course was unchanged in one patient, who died a few months later. The
other patient, whose initial prognosis was considered poor, is still alive and
well more than 3 years later with evidence of slowing of tumor growth based on
organ imaging.
The characterization of tumor-associated antigens recognized by human T
lymphocytes in a major histocompatibility complex (MHC)-restricted fashion has
opened new possibilities for immunotherapeutic approaches to the treatment of
human cancers. Dendritic cells (DC) are professional antigen presenting cells
that are well suited to activate T cells toward various antigens, such as
tumor-associated antigens, due to their potent costimulatory activity. The
availability of large numbers of DC, generated either from hematopoietic
progenitor cells or monocytes in vitro or isolated from peripheral blood, has
profoundly changed pre-clinical research as well as the clinical evaluation of
these cells. Accordingly, appropriately pulsed or transfected DC may be used
for vaccination in the field of infectious diseases or tumor immunotherapy to
induce antigen-specific T cell responses. These observations led to pilot
clinical trials of DC vaccination for patients with cancer in order to
investigate the feasibility, safety, as well as the immunologic and clinical
effects of this approach. Initial clinical studies of human DC vaccines are
generating encouraging preliminary results demonstrating induction of
tumor-specific immune responses and tumor regression. Nevertheless, much work
is still needed to address several variables that are critical for optimizing
this approach and to determine the role of DC-based vaccines in tumor
immunotherapy (63).
Dendritic cells are among the most efficient antigen-presenting cells of our
immune system and they play a crucial role in immunity reactions such as the
activation of T and B cells and the induction or maintenance of tolerance. New
culture methods allow us to generate dendritic cells in sufficient numbers for
further studies and for the preparation of antigen-loaded dendritic cells for
clinical application in cancer patients. In animal studies immunization with
antigen-loaded dendritic cells offered protection from growth of injected tumor
cells. In experimental clinical studies in cancer patients with e.g. metastatic
renal carcinoma, melanoma and B cell lymphoma some lasting remissions were observed
after administration of antigen-loaded dendritic cells. Side effects were
minor. Unanswered questions on tumor vaccines with antigen-loaded dendritic
cells concern specific matters, such as optimal culture methods and antigen
loading, and general matters, such as dose, frequency, duration and route of
administration. Also, no method is currently available by which the in vivo
immune response can be measured accurately (64).
Research over the last two years has explored a number of possible approaches
to applying dendritic cell immunotherapy to the treatment of human cancers. The
chosen strategy in clinical situations will vary for individual patients and
will depend on the type of tumor, availability of tissue samples and potential
source of dendritic cells. The isolation of fractionated tumor peptide from
individual patients' tumors for use with autologous stem cell-derived dendritic
cells may provide, in at least some cases, an effective and practical approach
to cancer immunotherapy. This approach will provide a 'closed' system of tumor
immunotherapy with all components (dendritic cells, antigen and cytotoxic T
lymphocytes) being provided by the patient and applied in a tailor-made fashion
to individual patients as an adjuvant to current anti-tumor therapies (65).
Cancer antibodies
The specificity of antibodies has been harnessed to target cancer cells and the
first therapeutic antibodies for use in oncology are now finding application in
the clinic. Studies are currently under way to develop new and improved
antibodies. Recent developments have been made in the identification of novel
targets, including the use of genomic and proteomic technologies. Several
methods are also being developed to enhance antibody efficacy (66).
Bi-specific antibodies (BsAbs) combine immune cell activation with tumor cell
recognition as a result of which tumor cells are killed by pre-defined effector
cells (67).
Antibody-based therapy of human cancers has led to several remarkable outcomes,
particularly in the therapy of breast cancer and lymphoma. Many solid tumors
have proven less responsive, due in part to difficulties in the tumor-selective
delivery of antibodies and potential cytolytic effectors. However, antibodies
that directly perturb signaling mechanisms in cells derived from epithelial
malignancies have shown benefit; examples include antibodies directed against
the extracellular domains of HER2/neu and epidermal growth factor receptor. A
long-term goal of immunotherapy has been to induce anti-tumor inflammatory
responses that can directly cause tumor regression or induce adaptive responses
against tumor-related antigens (68).
Specific targeting
of tumor cells may be achieved by using monoclonal antibodies to tumor
antigens. Edrecolomab is a mouse-derived monoclonal IGg2A antibody directed
against the human tumor-associated CO17-1A (or Ep-CAM) antigen, and is the
first monoclonal antibody approved for cancer therapy. Encouraging results of
several clinical trials were recently reported using edrecolomab for adjuvant
therapy after surgery of Duke's C colorectal cancer. Side effects and toxicity
profiles compare favorably to conventional regimens of radio- or chemotherapy.
Future challenges lie in further improvement of these novel therapeutics,
hopefully generating benefit for a larger number of cancer patients (69).
Gangliosides on tumor cells have been suggested as potential target antigens
for specific immunotherapy in various types of cancer including small cell lung
cancer (SCLC). Brezicka et al. (70) have compared the expression of three
gangliosides that have been described as tumor-associated antigens, FucGM1, GM2
and GD3 in SCLC tissue specimens collected at autopsy, using a double-layer
immunofluorescence staining method and specific monoclonal antibodies (Mabs)
directed against these ganliosides. They found expression of FucGM1, GD3 and
GM2 in 70% (n=20), 60% (n=15) and 40% of the tumor cells in all lesions from
the same patient (five of eight cases). These results indicate that FucGM1 is a
relevant ganglioside antigen in SCLC, and suggest that specific immunotherapy
involving more than one ganglioside antigen in SCLC should at least include
FucGM1 and GD3.
There is now a considerable body of information documenting the autoimmune
consequences of antibodies induced by growing malignancies, or by passively
administered and actively induced antibodies, in cancer patients against
antigens shared by normal and malignant tissues. This provides a rich source of
information addressing the consequences of autoantibodies against a range of
antigens. Antibodies against cell-surface or intracellular antigens in the
central nervous system (CNS) or on epithelial surfaces of normal tissues do not
generally result in autoimmunity, but the same types and titers of antibodies
against cell surface antigens in the subepidermal skin, peripheral nerves,
blood, or vascular sites such as the spleen and bone marrow readily induce
autoimmunity. The blood brain barrier of the CNS and apical antigen expression
and the basement membrane in epithelial tissues, may protect these sites from
antibody induced damage. Cancer cells, however, are protected by neither
unidirectional antigen expression nor basement membranes. Vaccine induced
antibodies against a variety of cancer cell surface antigens have been
associated with prevention of tumor recurrence in preclinical models and in
vaccinated cancer patients, in the absence of demonstrable autoimmunity. This
forms the basis for a series of ongoing Phase III trials with single or
polyvalent antigen cancer vaccines designed for optimal antibody induction
(71).
Immunotherapy of cancer is still mainly an experimental treatment. Some
monoclonal antibodies have been approved for adjuvant therapy of cancer in
patients, but active immunization strategies have not yet matured to this
stage. The fact that vaccination against viral diseases is effective has primed
high expectations for successful vaccination against cancer as well. Indeed, in
some animal models, therapeutic results could be obtained against short-term
established tumors, which paved the way for clinical trials. However, the first
results with active immunization in cancer patients were disappointing and this
led to a careful examination of current protocols and the search for more
effective approaches. Evaluation of the available data suggests that cancer
patients may not be comparable in their immune response to cancer vaccines with
healthy persons. Furthermore, the tumor seems to be able to develop several
immune-escape mechanisms, which either inactivate the specific immune cells or
prevent activation of potential effector mechanisms against the tumor (72).
Genetic immunotherapy
The establishment of cancer in a host involves at least two major events: the
escape of tumor cells from normal growth control and their escape from
immunological recognition. Because of this nature of their development, cancer
cells seem to be predominatly poorly immunogenic. In contrast to the previous
idea that cancer cells express no recognizable antigens, recent progress in the
identification and characterization of tumor antigens, as well as the expansion
of knowledge on the cellular and molecular mechanisms of antigen recognition by
the immune system, have raised the possibility of using immunotherapy to treat
certain tumors. Information on these mechanisms has been obtained in three
crucial areas: 1) the role of cytokines in the regulation of the immune
response, 2) the molecular characterization of tumor antigens in both mouse and
human tumors, and 3) the molecular mechanisms of T cell activation and antigen
presentation. Such information has provided new insight into tumor immunology
and immunotherapy. Furthermore, recombinant DNA technology allows for
modification of the genome of mammalian cells for therapeutic purposes in
several diseases. Several novel strategies have been developed to derive
genetically modified tumor cells and use them as cellular vaccines to induce antitumor
immunity in animal tumor models. This combined modality of genetically modified
tumor cells and immunotherapy has been termed immunogene therapy of tumors.
Crucial to this approach has been the ability to transfer into normal or
neoplastic cells genes known to increase the immunogenicity of cells, which
subsequently can be used to augment immune reactions in tumor-bearing mice or
cancer patients. While there has been success in inducing antitumor immunity in
some tumor models, there are difficulties and limitations in the application of
these gene-modified tumor cells for the treatment of preexisting tumors (73).
Genetic immunization refers to treatment strategies where gene transfer methods
are used to generate immune responses against cancer. Our growing knowledge of
the mechanisms regulating the initiation and maintenance of cytotoxic immune
responses has provided the rationale for the design of several genetic
immunization strategies. Tumor cells have been gene-modified to express immune
stimulatory genes and are then administered as tumor vaccines, in an attempt to
overcome tumor cell ignorance by the immune system. With the description of
well-characterized tumor antigens, multiple strategies have been proposed
mainly aimed at optimal tumor antigen presentation by antigen-presenting cells
(APC). Among APC, the dendritic cells have been recognized as the most powerful
cells in this class, and have become the target for introducing tumor antigen
genes to initiate antitumor immune responses. The detailed knowledge of how the
immune system can be activated to specifically recognize tumor antigens, and
the mechanisms involved in the control of this immune response, provide the
basis for modern genetic immunization strategies for cancer treatment (74).
T lymphocytes play a crucial role in the host's immune response to cancer.
Although there is ample evidence for the presence of tumor-associated antigens
on a variety of tumors, they are seemingly unable to elicit an adequate
antitumor immune response. Modern cancer immunotherapies are therefore designed
to induce or enhance T cell reactivity against tumor antigens. Vaccines
consisting of tumor cells transduced with cytokine genes in order to enhance
their immunogenicity have been intensely investigated in the past decade and
are currently being tested in clinical trials. With the development of novel
gene transfer technologies it has now become possible to transfer cytokine
genes directly into tumors in vivo. The identification of genes encoding
tumor-associated antigens and their peptide products which are recognized by
cytotoxic T lymphocytes in the context of major histocompatibility complex
class I molecules has allowed development of DNA-based vaccines against defined
tumor antigens. Recombinant viral vectors expressing model tumor antigens have
shown promising results in experimental models. This has led to clinical trials
with replication-defective adenoviruses encoding melanoma-associated antigens
for the treatment of patients with melanoma. An attractive alternative concept
is the use of plasmid DNA, which can elicit both humoral and cellular immune
responses following injection into muscle or skin. New insights into the
molecular biology of antigen processing and presentation have revealed the
importance of dendritic cells for the induction of primary antigen-specific T
cell responses. Considerable clinical interest has arisen to employ dendritic
cells as a vehicle to induce tumor antigen-specific immunity. Advances in
culture techniques have allowed the generation of large numbers of
immunostimulatory dendritic cells in vitro from precursor populations derived
from blood or bone marrow. Experimental immunotherapies which now transfer genes
encoding tumor-associated antigens or cytokines directly into professional
antigen-presenting cells such as dendritic cells are under evaluation in
pre-clinical studies at many centers. Gene therapy strategies, such as in vivo
cytokine gene transfer directly into tumors as well as the introduction of
genes encoding tumor-associated antigens into antigen-presenting cells hold
considerable promise for the treatment of patients with cancer (75).
Adjuvant immunotherapy
In the course of a century, tumor immunology has revealed a picture of a very
complex immune system involving the recognition and eradication of
malignancies. Many tumors evade the immune system, and understanding of tumor
escape mechanisms is the key to a successful immunotherapy for cancer. A wide
array of tumor immunotherapy modalities have been developed, many of which have
reached the phase of clinical trials, with some satisfactory results (76).
Although surgery remains the mainstay for the treatment of most solid tumors,
investigators are seeking complementary therapies to eradicate microscopic
disease, which causes tumor relapse even after an apparently complete surgical
excision. Although adjuvant chemotherapy has achieved some significant results,
the control of minimal residual disease is still a challenge for clinicians.
Among novel therapeutic approaches, immunotherapy holds promise. This
anticancer strategy aims at triggering a highly specific endogenous killing
machine against tumor cells. Recent progress in tumor immunology has improved
our understanding of host-immune system interactions. In particular, new
technologies have fostered the identification of potentially immunogenic tumor
antigens that can be used as suitable targets for immune effector cells. After
observing immunotherapy-mediated clinical responses in patients with metastatic
disease, investigators have started evaluating this anticancer modality in the
adjuvant setting (77).
Cancer
urotherapy
Urotherapy
is suggested as a kind of immunotherapy for cancer patients. Unlike the clonal
immunotherapy the urine of the cancer patients contain the many tumor antigens
which constitute the tumor. Oral auto-urotherapy will provide the intestinal
lymphatic system the tumor antigens against which they may produce antibodies
due to non-self recognition. These antibodies may be transpierced through the
blood stream and attack the tumor and its cells (78).
Bibliography
1. Shu S, Plautz GE, Krauss JC, Chang AE. Tumor immunology. JAMA. 1997 Dec 10;278(22):1972-81.
2. Cerundolo V. T cells work together to fight cancer. Curr Biol. 1999 Sep
23;9(18):R695-7.
3. Kawakami Y. New cancer therapy by immunomanipulation:
development of immunotherapy for human melanoma as a model system. Cornea. 2000 May;19(3
Suppl):S2-6.
4. Houghton AN, Gold JS, Blachere NE.
Immunity against cancer: lessons learned from
melanoma. Curr Opin Immunol. 2001 Apr;13(2):134-40.
5. Lieberman SM, Horig H, Kaufman HL.
Innovative treatments for pancreatic cancer. Surg Clin North Am. 2001
Jun;81(3):715-39.
6. Turk MJ, Wolchok JD, Guevara-Patino
JA, Goldberg SM, Houghton AN. Multiple
pathways to tumor immunity and concomitant autoimmunity. Immunol Rev. 2002
Oct;188:122-35.
7. Koroleva EP, Lagar'kova MA,
Khlgatian SV, Shebzukhov IuV, Meshcheriakov AA, lichinitser MR, Nedospasov SA,
Kuprash DV. Serological study of a
repertoire of human cancer antigens and autoantigens. Mol Biol (Mosk). 2004
Mar-Apr;38(2):233-8.
8. Lollini PL, Forni G. Cancer immunoprevention: tracking down persistent
tumor antigens.
Trends Immunol. 2003 Feb;24(2):62-6.
9. Scanlan MJ, Gure AO, Jungbluth AA,
Old LJ,
10. Perez-Diez A, Marincola FM. Immunotherapy against antigenic tumors: a game with
a lot of players.
Cell Mol Life Sci. 2002 Feb;59(2):230-40.
11. van den Eynde B. Identification of cancer antigens of relevance for
specific cancer immunotherapy. Bull Mem Acad R Med Belg. 2001;156(10-12):548-55.
12. Itoh K, Yamana H, Shichijo S,
Yamada A. Human tumor-rejection
antigens and peptides from genes to clinical research.
13. Slingluff CL. Targeting unique tumor antigens and modulating the
cytokine environment may improve immunotherapy for tumors with immune escape
mechanisms. Cancer
Immunol Immunother. 1999 Oct;48(7):371-3.
14. Wang RF. Human tumor antigens: implications for cancer
vaccine development.
J Mol Med. 1999 Sep;77(9):640-55.
15. Kudo T, Suzuki M, Katayose Y, Shinoda
M, Sakurai N, Kodama H, Ichiyama M, Takemura S, Yoshida H, Saeki H, Saijyo S,
Takahashi J, Tominaga T, Matsuno S. Specific
targeting immunotherapy of cancer with bispecific antibodies. Tohoku J Exp Med. 1999
Aug;188(4):275-88.
16. Huang SK, Okamoto T, Morton DL,
Hoon DS. Antibody responses to
melanoma/melanocyte autoantigens in melanoma patients. J Invest Dermatol. 1998
Oct;111(4):662-7.
17. Wang RF. Tumor antigens discovery: perspectives for cancer
therapy. Mol Med.
1997 Nov;3(11):716-31.
18. Ribas A, Butterfield LH, Glaspy JA,
Economou JS. Current developments in
cancer vaccines and cellular immunotherapy. J Clin Oncol. 2003 Jun 15;21(12):2415-32.
19. Zoller M. Immunotherapy of cancer by active vaccination: does
allogeneic bone marrow transplantation after non-myeloablative conditioning
provide a new option?
Technol Cancer Res Treat. 2003 Jun;2(3):237-60.
20. Chada S, Mhashilkar A, Roth JA,
Gabrilovich D. Development of vaccines
against self-antigens: the p53 paradigm. Curr Opin Drug Discov Devel. 2003 Mar;6(2):169-73.
21. Pecher G. DNA-based tumor vaccines. Onkologie. 2002
Dec;25(6):528-32.
22. Dermime S, Armstrong A, Hawkins RE,
Stern PL. Cancer vaccines and
immunotherapy. Br
Med Bull. 2002;62:149-62.
23. Perales MA, Wolchok JD. Melanoma vaccines. Cancer Invest. 2002;20(7-8):1012-26.
24. Cohen EP. DNA-based vaccines for the treatment of cancer--an
experimental model.
Trends Mol Med. 2001 Apr;7(4):175-9.
25. Berd D. Autologous, hapten-modified vaccine as a treatment
for human cancers.
Vaccine. 2001 Mar 21;19(17-19):2565-70.
26. Bhattacharya-Chatterjee M,
Chatterjee SK, Foon KA. The
anti-idiotype vaccines for immunotherapy. Curr Opin Mol Ther. 2001 Feb;3(1):63-9.
27. Bhattacharya-Chatterjee M,
Chatterjee SK, Foon KA. Anti-idiotype
vaccine against cancer. Immunol Lett. 2000 Sep 15;74(1):51-8.
28. Mitchell DA, Nair SK. RNA transfected dendritic cells as cancer vaccines. Curr Opin Mol Ther. 2000
Apr;2(2):176-81.
29. Wang RF, Rosenberg SA. Human tumor antigens for cancer vaccine development. Immunol Rev. 1999
Aug;170:85-100.
30. Timmerman JM, Levy R. Dendritic cell vaccines for cancer immunotherapy. Annu Rev Med. 1999;50:507-29.
31. Chen CH, Wu TC. Experimental vaccine strategies for cancer
immunotherapy. J
Biomed Sci. 1998 Jul-Aug;5(4):231-52.
32. Gilboa E,
33. Dudley ME, Rosenberg SA. Adoptive-cell-transfer therapy for the treatment of
patients with cancer.
Nat Rev Cancer. 2003 Sep;3(9):666-75.
34. Whelan M, Whelan J, Russell N,
Dalgleish A. Cancer immunotherapy: an
embarrassment of riches? Drug Discov Today. 2003 Mar 15;8(6):253-8.
35. Kwak H, Horig H, Kaufman HL.
Poxviruses as vectors for cancer
immunotherapy. Curr
Opin Drug Discov Devel. 2003 Mar;6(2):161-8.
36. Markiewicz MA, Kast WM. Advances in immunotherapy for prostate cancer. Adv Cancer Res. 2003;87:159-94.
37. Ward S, Casey D, Labarthe MC,
Whelan M, Dalgleish A, Pandha H, Todryk S. Immunotherapeutic potential of whole tumour cells. Cancer Immunol Immunother. 2002
Sep;51(7):351-7.
38. Sugiyama H. Cancer immunotherapy targeting WT1 protein. Int J Hematol. 2002
Aug;76(2):127-32.
39. Wang RF, Zeng G, Johnston SF, Voo
K, Ying H. T cell-mediated immune
responses in melanoma: implications for immunotherapy. Crit Rev
Oncol Hematol. 2002 Jul;43(1):1-11.
40. Fehniger TA, Cooper MA, Caligiuri
MA. Interleukin-2 and
interleukin-15: immunotherapy for cancer. Cytokine Growth Factor Rev. 2002 Apr;13(2):169-83.
41. Rosenberg SA. Progress in the development of immunotherapy for the
treatment of patients with cancer. J Intern Med. 2001 Dec;250(6):462-75.
42. McLaughlin PM, Kroesen BJ, Harmsen
MC, de Leij LF. Cancer
immunotherapy: insights from transgenic animal models. Crit Rev Oncol Hematol. 2001
Oct;40(1):53-76.
43. Maki RG. Soft tissue sarcoma as a model disease to examine
cancer immunotherapy.
Curr Opin Oncol. 2001 Jul;13(4):270-4.
44. Rosenberg SA. Progress in human tumour immunology and
immunotherapy.
Nature. 2001 May 17;411(6835):380-4.
45. Bertolaccini L, Olivero G. Cancer immunotherapy. A future therapeutical choice? Minerva Chir. 2001
Apr;56(2):183-91.
46. Smyth MJ, Godfrey DI, Trapani JA.
A fresh look at tumor immunosurveillance and
immunotherapy. Nat
Immunol. 2001 Apr;2(4):293-9.
47. Vinals C, Gaulis S, Coche T.
Using in silico transcriptomics to search for
tumor-associated antigens for immunotherapy. Vaccine. 2001 Mar 21;19(17-19):2607-14.
48. Matzku S, Zoller M. Specific immunotherapy of cancer in elderly
patients. Drugs
Aging. 2001;18(9):639-64.
49. Rousseau R, Soria JC. Telomerase, a universal target in immunotherapy
strategies against tumor? Bull Cancer. 2000 Dec;87(12):895-901.
50. Davis ID. An overview of cancer immunotherapy. Immunol Cell Biol. 2000
Jun;78(3):179-95.
51. Bremers AJ, Parmiani G. Immunology and immunotherapy of human cancer:
present concepts and clinical developments. Crit Rev Oncol Hematol. 2000 Apr;34(1):1-25.
52. Shankar G, Salgaller ML. Immune monitoring of cancer patients undergoing
experimental immunotherapy. Curr Opin Mol Ther. 2000 Feb;2(1):66-73.
53. Knuth A, Jager D, Jager E. Cancer immunotherapy in clinical oncology. Cancer Chemother Pharmacol.
2000;46 Suppl:S46-51.
54. Pawelec G, Engel A, Adibzadeh M.
Prerequisites for the immunotherapy of cancer. Cancer Immunol Immunother. 1999
Jul;48(4):214-7.
55. Maxwell-Armstrong CA, Durrant LG,
Scholefield JH. Immunotherapy
for colorectal cancer. Am J Surg. 1999 Apr;177(4):344-8.
56. Lum LG. T cell-based immunotherapy for cancer: a virtual
reality? CA Cancer
J Clin. 1999 Mar-Apr;49(2):74-100, 65.
57. Rosenberg SA. A new era of cancer immunotherapy: converting theory
to performance. CA
Cancer J Clin. 1999 Mar-Apr;49(2):70-3, 65.
58. Hersey P. Impediments to successful immunotherapy. Pharmacol Ther. 1999 Feb;81(2):111-9.
59. Przepiorka D, Srivastava PK.
Heat shock protein--peptide complexes as
immunotherapy for human cancer. Mol Med Today. 1998 Nov;4(11):478-84.
60. Tamura Y, Peng P, Liu K, Daou M,
Srivastava PK. Immunotherapy of tumors
with autologous tumor-derived heat shock protein preparations. Science. 1997 Oct
3;278(5335):117-20.
61. Catros-Quemener V, Bouet F, Genetet
N. Antitumor immunity and cellular cancer
therapies. Med Sci
(Paris). 2003 Jan;19(1):43-53.
62. Ladhams A, Schmidt C, Sing G,
Butterworth L, Fielding G, Tesar P, Strong R, Leggett B, Powell L, Maddern G,
Ellem K, Cooksley G. Treatment of
non-resectable hepatocellular carcinoma with autologous tumor-pulsed dendritic
cells. J
Gastroenterol Hepatol. 2002 Aug;17(8):889-96.
63. Meidenbauer N, Andreesen R,
Mackensen A. Dendritic cells for
specific cancer immunotherapy. Biol Chem. 2001 Apr;382(4):507-20.
64. Punt CJ, de Vries IJ, Mulders PF,
Adema GJ, Figdor CG. Immunology in
medical practice. XXV. Use of dendritic cells in the immunotherapy of cancer. Ned Tijdschr Geneeskd. 1999 Nov
27;143(48):2408-14.
65. Hermans IF, Moroni-Rawson P,
Ronchese F, Ritchie DS. The emerging
role of the dendritic cell in novel cancer therapies. N Z Med J. 1998 Apr
10;111(1063):111-3.
66. Trikha M, Yan L, Nakada MT. Monoclonal antibodies as therapeutics in oncology. Curr Opin Biotechnol. 2002
Dec;13(6):609-14.
67. Withoff S, Helfrich W, de Leij LF,
Molema G. Bi-specific antibody
therapy for the treatment of cancer. Curr Opin Mol Ther. 2001 Feb;3(1):53-62.
68. Weiner LM, Adams GP. New approaches to antibody therapy. Oncogene. 2000 Dec
11;19(53):6144-51.
69. Abicht A, Lochmuller H. Technology evaluation: edrecolomab, Centocor Inc. Curr Opin Mol Ther. 2000
Oct;2(5):593-600.
70. Brezicka T, Bergman B, Olling S,
Fredman P. Reactivity of monoclonal
antibodies with ganglioside antigens in human small cell lung cancer tissues. Lung Cancer. 2000
Apr;28(1):29-36.
71. Livingston PO, Ragupathi G,
Musselli C. Autoimmune and antitumor
consequences of antibodies against antigens shared by normal and malignant
tissues. J
Clin Immunol. 2000 Mar;20(2):85-93.
72. Velders MP, Schreiber H, Kast WM.
Active immunization against cancer cells:
impediments and advances. Semin Oncol. 1998 Dec;25(6):697-706.
73. Yoshizawa H, Kagamu H, Gejyo F.
Cancer immunogene therapy. Arch Immunol Ther Exp (Warsz).
2001;49(5):337-43.
74. Ribas A, Butterfield LH, Economou
JS. Genetic immunotherapy for
cancer. Oncologist.
2000;5(2):87-98.
75. Tuting T, Storkus WJ, Lotze MT.
Gene-based strategies for the immunotherapy of
cancer. J Mol Med.
1997 Jul;75(7):478-91.
76. Bremers AJ, Kuppen PJ, Parmiani G.
Tumour immunotherapy: the adjuvant treatment
of the 21st century?
Eur J Surg Oncol. 2000 Jun;26(4):418-24.
77. Mocellin S, Rossi CR, Lise M,
Marincola FM. Adjuvant immunotherapy
for solid tumors: from promise to clinical application. Cancer Immunol Immunother. 2002
Dec;51(11-12):583-95.
78. Eldor
J. Urotherapy
for patients with cancer. Med Hypotheses. 1997
Apr;48(4):309-15.