Perspective

Targeted Therapies to Improve Tumor Immunotherapy

Jonathan Begley1,2 and Antoni Ribas2,3,4

Abstract Durable tumor regression and potential cures of metastatic solid cancers can be achieved by a
variety of cellular immunotherapy strategies, including cytokine therapy, dendritic cell ^ based
vaccines, and immune-activating antibodies, when used in so-called immune-sensitive cancers
such as melanoma and renal cell carcinoma. However, these immunotherapy strategies have very
low tumor response rates, usually in the order of 5% to 10% of treated patients.We propose that
the antitumor activity of adequately stimulated tumor antigen ^ specific T cells is limited by local
factors within the tumor milieu and that pharmacologic modulation of this milieu may overcome
tumor resistance to immunotherapy. By understanding the mechanisms of cancer cell immune
escape, it may be possible to design rational combinatorial approaches of novel therapies able
to target immunosuppressive or antiapoptotic molecules in an attempt to reverse resistance to
immune system control. We term this mode of treatment ‘‘immunosensitization.’’ Ideal candidates
for immunosensitizing drugs would be targeted drugs that block key oncogenic mechanisms
in cancer cells resulting in a proapoptotic cancer cell milieu and at the same time do not nega-
tively interfere with critical lymphocyte functions.

One of the hallmarks of cancer responses to cellular immu-
notherapy is that they are extremely long lived, frequently
measured in years. However, objective responses (as opposed to
the more common claim of mixed responses or disease
stabilization with immunotherapy) are rare, usually noted in
a minority of patients, in the range of 5% to 10% (1, 2). The
improved understanding of the oncogenic events in cancer
and the development of multiple new highly targeted drugs
that block or neutralize oncogenic factors open the door for
potential novel combinatorial approaches with immunother-
apy. In this article, we postulate that targeted agents against
key immunosuppressive, oncogenic, or antiapoptotic factors
in cancer cells can sensitize cancers to immunotherapy. We
restrict the definition of tumor immunotherapy to mean to
activate cellular cytotoxic antitumor immune responses, mostly
mediated by CD8+ CTLs and natural killer (NK) cells. This is
opposed to therapy mediated by antibodies such as rituximab,
trastuzumab, or bevacizumab, which target surface or secreted
proteins in cancer cells but are not designed to induce adaptive
immune responses to the cancer. We suggest that optimal
combinations of these drugs and cellular immunotherapy
will result in improved antitumor responses as targeted drugs
Authors’ Affiliations: 1Monash University, Victoria, Melbourne, Australia; and
Departments of 2Surgery and 3Medicine, University of California at Los Angeles;
and 4Jonsson Comprehensive Cancer Center, Los Angeles, California
Received 11/2/07; revised 1/7/08; accepted 1/23/08.
Grant support: Harry J. Lloyd Charitable Trust and Melanoma Research
Foundation.
The costs of publication of this article were defrayed in part by the payment of page
charges. This article must therefore be hereby marked advertisement in accordance
with 18 U.S.C. Section 1734 solely to indicate this fact.
Requests for reprints: Antoni Ribas, Division of Hematology-Oncology,
University of California at Los Angeles Medical Center, 11-934 Factor Building,
10833 Le Conte Avenue, Los Angeles, CA 90095-1782. Phone: 310-206-3928;
Fax: 310-206-0914; E-mail: aribas@ mednet.ucla.edu.
F 2008 American Association for Cancer Research.
doi:10.1158/1078-0432.CCR-07-4804
fight immune resistance and allow the durable responses seen
with immunotherapy to take hold.
Limiting Steps for Effective Tumor
Immunotherapy
The multiple advances in understanding the immunobiology
of T-cell responses to cancer have not translated into wide-
spread clinical utility (1). Cytokine-based therapies, dendritic cell
(DC)-based vaccines, and treatment with immune-activating
antibodies, particularly with blocking antibodies to the CTL-
associated protein 4 (CTLA4), have a ceiling of tumor responses
in a subset of so-called immune-sensitive tumors (mainly
melanoma and renal cell carcinoma) in the range of 5% to 10%
of treated patients (1, 2). However, enthusiasm for tumor
immunotherapy is maintained by the realization that most
tumor responses are durable, leading to rare chances of cure in
patients with widely metastatic solid tumors. That is the basis
for the Food and Drug Administration licensing of the use of
high-dose interleukin (IL)-2 in metastatic melanoma and renal
cell carcinoma (3), and consistent with the findings emerging
from ongoing clinical studies developing DC-based vaccines
(2, 4) and anti-CTLA4 antibodies (5, 6). Therefore, although we
have the proof of concept about the effectiveness of tumor
immunotherapy, major improvements are needed in many key
aspects of this type of therapy.
There are many factors that influence tumor response to
immunotherapy. Neoplastic cells survive and proliferate in an
often hostile environment. Under constant selection pres-
sure, cells with a survival advantage replicate, thus ensuring
the tumor ‘‘evolves’’ to suit its environment. Tumor cells that
develop mechanisms to evade the immune system allow the
cancer to become immunoresistant in a process known as
cancer immunoediting (7). Within this context, the ability
of immune system cells to attack cancers is limited. Figure 1
depicts a hypothetical model of tumor immunotherapy

www.aacrjournals.org 4385 Clin Cancer Res 2008;14(14) July 15, 2008

Perspective
highlighting the key steps in generating an effective antitumor
immune response. At each step, the immune system or the
tumor may limit or prevent the effective elimination of cancer
cells by T cells. Obviously, this is a simplified model given the
complexity of the immune response and the multiple cell
subsets involved. However, we focus on the key interactions
leading to an effective adaptive immune response to cancer to
help describe what we believe are the key limitations to current
immunotherapy.
T-cell activation by antigen-presenting cells. The first step in
generating an effective antitumor immune response leads to the
licensing of CD8+ CTLs, which, through the T-cell receptor,
specifically recognize their cognate antigen presented by tumor
cells through MHC class I molecules. Most tumor antigens are
self-antigens (i.e., antigen normally expressed on healthy cells
and shared by cancer cells). T cells recognizing these self-
antigens would cause autoimmunity; self-reactive T cells are
subjected to negative selection in the thymus and are only
detectable in very low levels in the periphery. In addition,
tumor antigens are frequently down-regulated by cancer cells
and do not themselves lead to activation of tumor antigen –
specific CTL (8). For these antigens to be recognized, they need
to be presented by DC to CD4+ T helper cells or cross-presented
(a process in which exogenous antigen is presented in MHC
class I) by DC to CD8+ CTL (9) in an interaction modulated by
costimulatory and coinhibitory molecules (10). The develop-
ment of methods to generate DC ex vivo allowed their testing as
a powerful means to activate the immune system to cancer self-
antigens (11). The clinical results to date are suboptimal (1)
and highlight the need to address other variables involved
in immune-mediated tumor regression other than the use of
powerful vaccines.
Costimulatory and coinhibitory molecules. Antigen presenta-
tion by MHC recognized by T-cell receptors on T lymphocytes is
not enough to activate an adaptive immune response; it
Clin Cancer Res 2008;14(14) July 15, 2008
Fig. 1. Modelof CTL response to melanoma.
There are several steps required for the
induction of an effective antitumor immune
response: (1) antigen-presenting cells need
to activate CD8+ tumor-specific effector cells
overcoming the limitations of targeting
tumor self-antigens; (2) CD8+ cells need to
overcome homeostatic mechanisms that
limit immune activation, and then they need
to expand and traffic to tumor targets; (3)
CD8+ cells have to recognize antigen
presented by tumor cells and (4) overcome
tumor-induced local negative immune
regulators; and (5) the tumor cells need to be
sensitive to the lytic effects of the immune
effector cells, delivered either through
perforin/granzyme or by death receptor
ligation (Fas,TRAIL, andTNF). GzB,
granzyme B;Treg,Tregulatory cell;VEGF,
vascular endothelial growth factor; PgE2,
prostaglandin E2;TGF-h, transforming
growth factor-h;TAP, transporter associated
with antigen processing; PD1, programmed
death1.
critically requires the delivery of costimulatory signals provided
by B7 molecules (CD80 and CD86) recognized by their
positive receptor CD28 on T cells. The coinhibitory molecule
CLTA4 (CD152) has a pivotal role in dampening immune
responses to self-antigens by competing with CD28 (12).
Blocking antibodies to CTLA4 are in clinical development and
have shown durable immune-mediated response rates in the
order 5% to 20% in patients with metastatic melanoma (5, 6,
13, 14). Several other costimulatory and coinhibitory targets are
amenable to intervention, and specific antibodies against
CD40, CD80, CD134 (OX40), CD137 (41BB), programmed
death 1, and others are being tested (10). Therefore, several
off-the-shelf potent immunostimulating agents are entering
clinical testing for cancer, making it feasible the future study of
combinatorial approaches.
T-cell expansion and tumor targeting. After CD8+ CTLs are
activated against cells expressing tumor antigens, they need to
overcome homeostatic mechanisms that limit immune activa-
tion and expansion. They then traffic to tumors. The immune
system has multiple breaks limiting this activation process,
including the expression of CTLA4 by activated T cells leading
to interference in establishing lasting interactions between
activated T cells and target cells expressing their cognate
antigen (15).
Recognition of cancer cells. Activated tumor antigen – specific
CD8+ cells that have traveled to the tumor have to then recog-
nize their cognate antigen presented by tumor cells to speci-
fically deliver their cytotoxic signals. The tumor has developed
elaborated mechanisms to escape the immune system. Low-
level expression of antigen, MHC molecules (HLA in humans),
h2-microglobulin required for MHC assembly, or transporter
associated with antigen processing molecules required to
provide peptide epitopes for MHC stability on the cell surface,
may result in CD8+ CTL that cannot find their cognate antigen
on cancer cells (16).
4386 www.aacrjournals.org

Immunosuppressive tumor milieu. Local immunosuppressive
factors within the tumor milieu include secreted molecules,
such as transforming growth factor-h, IL-10, prostaglandin
E2, and vascular endothelial growth factor (VEGF). These
factors are produced either by the tumor cells themselves or by
surrounding host stromal cells on tumor cell signaling. The
suppressive tumor environment is further maintained by
immunosuppressive cells, such as T regulatory cells, and
indolamine 2,3-dioxygenase (IDO)-producing plasmacytoid
DC (17, 18). These are professional immune suppressor cells
that are frequently found inside tumors and provide means for
tumor escape from activated lymphocytes.
Escape from immune effector cell killing of target cells. Im-
mune effector cells (either CD8+ CTL or NK cells) kill their
targets by four defined mechanisms: the release of perforin and
granzymes (see ref. 19 for review) and the engagement of the
Fas, tumor necrosis factor (TNF)-related apoptosis-inducing
ligand (TRAIL), and TNF death receptors (see ref. 20 for
review). A simplified schematic of this process is shown
in Fig. 2. On entry into target cells facilitated by perforin,
granzyme B directly cleaves Bid (a proapoptotic Bcl-2 family
member) resulting in activation of caspase-3 and other
downstream effector caspases, leading to apoptotic cell death.
Engagement of the surface Fas, TRAIL, or TNF death receptors
by their secreted or cell surface – anchored ligands presented on
immune effector cells leads to the activation of caspase-8 within
the target cell and activation of the extrinsic pathway of
apoptosis. Caspase-8 also activates the intrinsic (also called
‘‘mitochondrial’’) apoptosis pathway (see ref. 21 for review),
which depends heavily on the balance between proapoptotic
and antiapoptotic molecules, particularly of the Bcl-2 family.
Intracellular effectors of cytotoxicity and apoptotic cell
death. The fate of a cell reflects the dynamic interplay between
apoptosis and survival; the Bcl-2 family is critical in maintain-
ing this balance. The family is classified into three groups: the
‘‘antiapoptotic,’’ ‘‘proapoptotic,’’ and ‘‘BH3-only’’ proteins (see
ref. 22 for further review). The antiapoptotic Bcl-2 family
members include Bcl-2, Bcl-XL, Bcl-w, Bcl-2A1, Mcl1, and Boo,
which promote cell survival by antagonizing apoptotic signals.
Overexpression of these molecules protects cells from apoptosis
and mediates cell survival (23). The proapoptotic Bcl-2 family
members include Bax, Bak, Box, and Bcl-XS. Overexpression of
these molecules has been shown to promote apoptosis (23).
These molecules are directly responsible for initiating apoptosis
in the mitochondrial pathway and are considered important
sensors of cellular stress (such as DNA damage). The BH3-only
subfamily is proapoptotic but is classified separately due to its
mechanism of action. It includes Bad, Bik, Bid, Hrk, Bim, Noxa,
Puma, and Bmf. The primary role of these molecules is the
activation of the proapoptotic family members.
Cancer resistance to apoptosis. Given the critical role of
proapoptotic and antiapoptotic proteins, it is not surprising
that altered expression of these proteins in cancerous cells often
allows them to resist apoptosis. Fas and TRAIL receptors are
down-regulated or undetectable in many cancers, obviously
leading to resistance to death receptor – induced apoptosis in
most cases (24). The nuclear factor-nB is a transcription factor
with constitutive expression in many cancers (25) and is
responsible for the transcription of many antiapoptotic
genes, including the FLICE inhibitory protein (FLIP). FLIP is
expressed at high levels in many cancers, where it competes
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Immunosensitization with Targeted Therapies
with caspase-8 for binding to the death-inducing signaling
complex. Expression of FLIP in cancer cells renders them insen-
sitive to Fas ligand or TRAIL. Bax or Bak expression is signifi-
cantly reduced in many cancers and confers worse prognosis
(26). Low expression of Apaf-1 (a proapoptotic molecule),
probably through methylation, is associated with chemore-
sistance (27). Equally, down-regulation of the death signal-
ing molecule Fas-associated protein with death domain and
caspase-8 has been shown in several cancers (28, 29). As
expected, up-regulation of inhibitors of apoptosis (antiapop-
totic molecules that inhibit the effector caspases) is overex-
pressed in tumor cells (30, 31).
Therapeutics with Immunopotentiating Profile
With an understanding of the potential role of the cellular
immune system in detecting and killing cancer cells, and
insight into how cancer cells avoid death from the immune
system, pharmacologic interventions may be designed to
sensitize cancer cells to immune attack. Table 1 provides the
features that we feel would make up an ideal immunosensitiz-
ing drug. It should potentiate the immune effector cell
recognition of cancer cells and shift the balance in the cancer
cell toward a proapoptotic milieu. Of particular importance,
potential immunosensitizing drugs must not be cytotoxic
against nor inhibit critical functions of immune cells.
Decreasing tumor-induced immune suppression. Secreted
proteins released by tumors that lead to immune escape can
be targets to monoclonal antibody blockade. For example, the
anti – VEGF antibody bevacizumab, in addition to its effect of
tumor vasculature, may decrease VEGF – induced inhibition
of DC and T-cell function (32). Monoclonal antibodies to
transforming growth factor-h and IL-10 are also potential
means to reverse immune escape induced by tumor-released
molecules. In addition, anti-inflammatory drugs such as aspirin
and other nonsteroidal anti-inflammatory drugs, including the
specific cyclooxygenase-2 inhibitor celecoxib, could be means to
inhibit cyclooxygenase-2 that leads to prostaglandin E2 produc-
tion (33).
Depletion of immunosuppressive cells. T regulatory cells,
myeloid suppressor cells, and IDO-positive plasmacytoid DC
have emerged as major immunosuppressive cells reported to be
implicated in escape of tumors from immune control (34, 35).
Depletion of CD4/CD25/FoxP3-positive natural T regulatory
cells can be achieved in mice using anti-CD25 antibodies,
which leads to improvement in responses to immunostimulat-
ing approaches such as CTLA4 blockade (36). There is much
interest in developing means to deplete human T regulatory
cells, although no compelling approach has yet been devel-
oped. The available antibodies to human CD25 approved by
the Food and Drug Administration are immunosuppressive as
opposed to immunostimulating, and there are mixed reports
on the ability of a CD25-targeted IL-2-diphtheria toxin fusion
protein (denileukin diftitox) to deplete T regulatory cell func-
tion (37). The function of the immunosuppressive enzyme IDO
can be inhibited by 1-methyl-tryptophan (17), and clinical
trials testing its effects in patients with cancer are planned. This
reagent would have the potential of synergizing with immu-
notherapy by inhibiting immune suppression by IDO.
Increase activating ligands for immune effector cells. Drugs
that mediate epigenetic mechanisms, such as the demethylating
Clin Cancer Res 2008;14(14) July 15, 2008
Perspective
agents and the histone deacetylase inhibitors, have multiple
effects on gene transcription. They tend to promote a pro-
apoptotic phenotype, can up-regulate MHC and tumor antigen
expression, induce ‘‘stress’’ molecules recognizable by NK-
activating receptors such as NKG2D, and increase expression of
surface death receptors (38). The ability of this classes of novel
agents to increase the surface expression of antigen-presenting
MHC molecules (HLA molecules in humans) and at the same
time increase expression of tumor antigens, such as the MAGE
family of cancer-testis antigens, may allow these agents to over-
come a major mechanism of tumor escape to cellular
immunotherapy (16).
Decreased inhibitory ligands for immune effector cells. Pep-
tides derived from proteasomal degradation are required for
assembly of MHC molecules in the endoplasmic reticulum
Table 1. Ideal attributes of targeted
immunosensitizing drugs
Effects on tumor cells
Specifically target a key oncogenic pathway
Increase death receptor expression
Increase proapoptotic molecules
Inhibit antiapoptotic molecules
Increase expression of tumor antigen or
NK-activating receptors
Effects on immune system cells
No cytotoxic effects on immune system cells
No interference with T-cell receptor, NK receptor,
or costimulatory signal transduction pathways
Clin Cancer Res 2008;14(14) July 15, 2008 4388
Fig. 2. Apoptotic pathways within the normal cell.
Cytotoxic immune cells can induce target cell death by
engaging death receptors or releasing perforin and
granzymes (Grn). This schematic shows a simplification
of the signaling activated by this process. DISC, death-
inducing signaling complex; FADD, Fas-associated
protein with death domain; IAP, inhibitor of apoptosis.
(39). This would suggest that ubiquitin ligase inhibitors and
proteasome inhibitors may decrease the pool of peptides able
to support assembly of surface MHC class I molecules.
Decreased expression of surface MHC class I allows cancer
cells to escape from CD8+ T-cell control, but at the same time,
they become more sensitive to NK cell recognition and killing.
NK cells are effector cells of the innate immune system, which
sense the loss of surface MHC molecules (frequent after viral
infections and in cancer cells) or the presence of foreign MHC
molecules (as occurs after organ transplants), which trigger
these potent cytotoxic cells. This possibility has been tested in
an experimental model of melanoma, suggesting that the
proteasome inhibitor bortezomib increased TNF-a – mediated
killing by immune cells (40).
Direct modulation of apoptotic pathways. Death receptors
are on the outer surface of cells and therefore accessible to
monoclonal antibodies. TRAIL receptors seem to be preferen-
tially expressed by cancer cells (and activated lymphocytes),
whereas Fas and TNF-a receptors are expressed more
promiscuously. A soluble TRAIL ligand construct and TRAIL
receptor-activating antibodies are in clinical development for
the treatment of cancer. These approaches may be viewed as
bypassing the need for immune effector cells but would be
unlikely to lead to sustained tumor regression without chronic
dosing. It is possible that such modulation of TRAIL receptors
may synergize with immunotherapy and other targeted therapy
approaches. Histone deacetylase inhibitors have been shown
to up-regulate the TRAIL death receptor DR5, leading to
sensitization of cancer cells to TRAIL-induced death (41, 42),
and therefore would be logical drugs to use in combination with
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both TRAIL-engaging reagents and immune effector cells.
Furthermore, the demethylating agent decitabine has been
shown to sensitize choroidal melanoma cell lines to the cyto-
toxic effects of IFNs by inducing the expression of proapoptotic
and growth-inhibitory genes (43).
The antiapoptotic members of the Bcl-2 family are potential
targets for immunosensitizing drugs. Bcl-2 is up-regulated in
many cancers (44), and inactivating its function may result in a
proapoptotic cancer cell milieu facilitating the cytotoxic effect
of immune cells. Small-molecule inhibitors of Bcl-2 family
members (45) have obvious pharmacologic advantages over
Bcl-2 antisense oligonucleotides (46) and may be logical
compounds to be used in combination with immunotherapy.
Other proteins in the apoptotic pathways also represent
potential targets for immunosensitizing agents. An inhibitor
of apoptosis inhibitor sensitized cells to TRAIL (47), making it a
reasonable drug to combine with immunotherapy. Apoptotic
pathways may also be targeted by altering transcription of
regulators of apoptosis. Inhibition of nuclear factor-nB in
melanoma sensitizes cells to TRAIL-induced (but not Fas or
TNF-a) apoptosis (48). An additional approach would be the
use of the proteasome inhibitor bortezomib, which is known to
inhibit nuclear factor-nB and down-regulate the antiapoptotic
molecule FLIP (49).
Alteration of Oncogenic Pathways Leading to
Immune Sensitization
Many interventions that damage cancer cells lead to a
proapoptotic milieu. Carefully sequenced treatment with
cytotoxic drugs or radiation therapy leading to DNA damage
stimulates DNA stress pathways, which can lead to immune
sensitization (50, 51). In cells that have functional p53, DNA
damage is sensed by p53, resulting in increased transcription of
death receptors (52). Although the adverse effects of standard
chemotherapy agents and radiation therapy, which are non-
specific genotoxins, on immune system cells make combina-
tions of chemotherapy and immunotherapy a rather poor
match, when adequately sequenced there may be situations
when cytotoxic therapy can be efficiently delivered to improve
on the antitumor activity of immunotherapy (51, 53).
Novel targeted therapies that disrupt dominant oncogenic
signals in cancer cells may result in increased sensitivity to
immune responses. There is a lot of interest in finding ways to
combine tyrosine kinase inhibitors, such as sorafenib and
sunitinib, with immunostimulating cytokines in the treatment
of renal cell carcinoma. A careful assessment of the effects on the
tumor, tumor microenvironment, and the immune system will be
required to define optimal combinations. Several clinical trials are
ongoing where these multitargeted tyrosine kinase inhibitors
are tested in combination with immunostimulating agents, such
as IL-2, IL-21, anti-CTLA4 antibodies, and poxvirus vaccines,
in patients with metastatic renal cell carcinoma and melanoma.
Inhibition of constitutively activated signal transduction
pathways in cancer cells also has the potential to decrease
tumor escape from immunotherapy. For example, loss of the
tumor suppressor PTEN in glioblastoma cells leads to
immune escape in part by increased translation of the
immune suppressor surface receptor B7-H1 (CD274, also
known as programmed death ligand 1). Treatment of these
cells with an Akt inhibitor (which blocked the Akt tyrosine
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Immunosensitization with Targeted Therapies
kinase constitutively activated in PTEN-deficient cells) re-
versed immune escape of glioblastoma cells (54). However,
in addition to its key role in oncogenesis, the phosphatidy-
linositol 3-kinase-Akt-mammalian target of rapamycin signal-
ing pathway is key to many lymphocyte functions. If targeted
drugs blocking this signal transduction pathway are to be
used as immune sensitizers, they will need to be carefully
scheduled or will need to show a high therapeutic window to
have the required effect on cancer cells without suppressing
lymphocyte functioning.
The signal transducer and activator of transcription (STAT)
proteins transmit cytoplasmic signals from polypeptide cyto-
kines and growth factors that have receptors with intrinsic or
associated tyrosine kinase activity. They have become promis-
ing molecular targets for cancer because they are persistently
activated in many cancers due to oncogene signaling through
STATs, especially STAT-3 and STAT-5 (55). These two STAT pro-
teins have key roles in regulating cell proliferation, resistance to
apoptosis, and sustained angiogenesis (55). In addition, STAT-3
has an important role in the immune system. Inhibiting STAT-3
in immune system cells increased DC, T-cell, and NK cell
function (56). Small-molecule inhibitors are being developed
to block STAT-3 and would therefore have the potential of
simultaneously attacking cancer cells and increasing immune
system function (56).
Issues About the Testing of Targeted Therapy-
Immunotherapy Combinatorial Approaches
Despite the promise, there are several practical limitations to
the combined testing of new agents in preclinical or clinical
development and tumor immunotherapy approaches. Preclin-
ical testing of a potentially immunosensitizing pharmacologic
agent and immunotherapy is best done in fully immunocom-
petent animal models. Ex vivo cell culture systems cannot fully
recapitulate a functional immune system and may provide
biased results, where potential adverse effects of the drug on
immune system cells may not be detected. However, implant-
able murine tumors may not be an adequate model for human
cancer because the oncogenic events that drive the cancers and
the relation between the cancer cells and the tumor milieu may
not be the same as in spontaneously arising tumors. A notable
example is the frequently used B16 murine melanoma, which
does not seem to have a constitutively active mitogen-activated
protein kinase pathway, a marked difference with human
melanomas where mutually exclusive activating mutations on
Nras of Braf are prevalent. The use of human tumor xenografts
requires implantation into severe immunodeficient mice,
obviously devoid of an immune system. Regeneration of a
human immune system in these mice is technically challenging,
and current approaches may not provide a fully functional
immune response to test immune sensitization approaches.
Another important limitation to the testing of the immune
sensitization combinatorial approach is access to adequate
compounds. It is widely known that intellectual property issues
limit the availability of emerging new drugs, and the testing of
combination regimens is frequently explicitly prohibited by the
manufacturers of the novel agents. Because the combination is
only relevant if there is a fully functioning immune system, the
concept frequently needs to be tested in a living animal. The
doses required for treatment in animal models are much higher
Clin Cancer Res 2008;14(14) July 15, 2008
Perspective

than the amount of agent that would be sufficient for in vitro which there is mechanistic understanding of how they
testing, so the immune sensitization combinatorial approach is potentiate each approach will be likely to be tested first. In
unlikely to be tested unless the manufacturer provides enough particular, pharmacologic means to affect tumor escape from
agent and explicit permission for combined agent testing. cellular immunotherapy would be a high priority for clinical
Alternatives such as using a published chemical structure to testing.
synthesize the agent de novo are likely to be limited by the high
costs of producing large amounts for testing in animal models.
Therefore, this approach usually requires a material transfer Conclusions
agreement with the manufacturer of the agent, with its resulting
Improvements in our understanding of the antiapoptotic
delay and limitations for combination testing. Testing in the
and immune escape mechanisms activated by oncogenic
clinic is also subjected to practical limitations. The same issues
events, and the ability to specifically alter them with targeted
related to the intellectual property of a combination described
drugs, will result in novel approaches allowing rational
for the preclinical testing apply to the clinic. Even when
combinations of drugs with immunostimulating effects. These
permission is obtained to test a new combination for which
combinations would bring together the elegance of targeted
there is strong scientific rationale, the combined testing may be
therapy for cancer with the ability of the immune system to
delayed until the safety, toxicity, and antitumor activity of each
lead to long-term regressions in some patients with metastatic
agent has been thoroughly tested independently.
cancers.
Despite all of these limitations, it is likely that in the next
several years the number of articles describing combinations of
novel targeted agents with established or new immunotherapy Disclosure of Potential Conflicts of Interest
approaches will continue to increase. This will be greatly
A. Ribas has received research funding and honoraria from Pfizer Inc.
facilitated once the novel agents have proven their benefit in
certain cancers and become standard of care therapies, as
exemplified by the ongoing clinical testing of multitargeted
Acknowledgments
tyrosine kinase inhibitors and cytokines. At some point, there
may be a need for prioritization of the combinations. It is clear We thank Glenn Begley for reviewing the manuscript and Benjamin Bonavida,
that combinations successfully tested in animal models for Hermes Garban, and James S. Economou for their valuable discussions.

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