Combining Immunotherapy and Anticancer Agents

The Right Path to Achieve Cancer Cure?

L. Apetoh; S. Ladoire; G. Coukos; F. Ghiringhelli


Ann Oncol. 2015;26(9):1813-1823. 

In This Article

The Emergence of Novel Immunotherapeutic Strategies Against Cancer

Dendritic Cells

Owing to their strong ability to initiate and control T-cell responses, dendritic cells (DCs) have long been regarded as attractive candidates for the design of immunotherapy strategies.[21] Initially, DC cancer vaccines consisted of ex vivo generated DCs that were loaded with tumor antigens. Although this vaccination strategy has successfully elicited anticancer immune responses in cancer patients, no or very few clinical benefits were noted (reviewed in[22]). This was, for instance, illustrated by the results of a phase III randomized clinical trial comparing the clinical efficacy of DC therapy to dacarbazine in metastatic melanoma patients. The objective response rate for DC therapy was 3.8%, explaining the early discontinuation of the study because of lack of efficacy.[23] Proposed reasons explaining the lack of efficacy of DC vaccines include the use of inappropriate DC maturation cocktail, thus compromising the ability of DC to induce anticancer responses. To circumvent these issues, different strategies are being implemented. Cytokine combinations to favor DC maturation and antigen presentation properties to design more effective DC-based immunization strategies are being tested (reviewed in[22,24]). A recent study demonstrated that the ability of a DC vaccine to increase, without any associated side effects, the breadth and diversity of melanoma neoantigen-specific T cells further enhances the potential of this approach.[25]

Adoptive T-cell Therapy

Adoptive T-cell therapy for cancer aims to eliminate cancer cells upon administration of T cells into tumor-bearing hosts. In 1955, Mitchison[26] initially demonstrated the feasibility of this approach in mice. In 1966, Southam et al. studied the anticancer activity of leucocytes from tumor-bearing patients against their respective tumor and raised the hypothesis that lymphocytes from cancer patients have a specific inhibitory effect against cancer cells. This provided impetus to exploit T cells from cancer patients to design effective adoptive transfer strategies. Subsequent studies focused on the isolation, expansion and reinfusion of tumor-infiltrating lymphocytes (TILs) into cancer patients. The studies pioneered by Rosenberg and colleagues relied on the culture of tumor-derived lymphocytes in the presence of high doses of IL-2 that were then transferred into tumor-bearing patients. Although encouraging results were noted, side effects impeded the large clinical implementation of this strategy. The safety and efficacy of TIL therapy was improved by implementing preconditioning regimens driving lymphodepletion. The use of a preconditioning regimen relying on cyclophosphamide and fludarabine was shown to eliminate the endogenous lymphocyte repertoire and favor growth and long-lasting persistence of the transferred TILs. This has led to improved responses rate up to 40%.[27] Research in T-cell biology has also improved TIL culture conditions, leading to shorter TIL expansion phase, thereby reducing the time from TIL collection to their reinfusion into cancer patients.

An alternative strategy to target tumor cells using T cells is the engineering of CAR T cells, which are endowed with a specific ability to recognize and kill cancer cells. CARs contain a fusion protein of light and heavy chains from an antibody, linked to the signaling machinery of the TCR. Such structure enables T-cell activation upon CAR recognition of its target. As CARs are not Major Histocompatibility complex (MHC)-restricted, they are insensitive to tumor-driven immunosuppression mediated through downregulation of MHC molecules. Another advantage of CAR T cells is the possibility of transducing genes that will enhance further T-cell functions upon activation or chemokine receptors that will favor T-cell homing. Although the initial trials in ovarian cancer and renal cell carcinoma using CAR T cells were disappointing because of toxicity and limited T-cell persistence in the tumor microenvironment,[28,29] the use of second-generation CAR T cells in leukemic patients has resulted in remarkable anticancer effects.[30,31] The success of CAR T-cell therapy in these diseases was associated with a high level of CAR T-cell proliferation following infusion into patients. The feasibility of this approach was further established in 30 patients suffering from relapsed or refractory acute lymphoblastic leukemia. CAR T-cell therapy targeting CD19 led to complete remission in 27 patients and sustained remission was achieved with a 6-month overall survival rate of 78%.[32] This notable efficacy has led to the United States Food and Drug Administration to designate the anti-CD19 CAR T-cell therapy as a 'breakthrough therapy'.


The efficacy of IL-2 as an anticancer agent has been investigated in multiple cancer types. It has been shown that high doses of IL-2 could be effective in eliciting anticancer responses in renal cell carcinoma and melanoma. Nevertheless, the overall response rates were low and the associated toxicities were severe (reviewed in[22]). IL-2 was further shown to drive the expansion of regulatory T cells which in turn suppress anticancer immune responses. This has prompted the test of additional cytokines for their anticancer potential upon in vivo administration. IL-15 was later identified as an interesting candidate molecule. The anticancer effects of IL-15 have been demonstrated in several preclinical tumor models. The underlying mechanisms have been subsequently identified. IL-15 has been shown to enhance NK cell effector functions. In addition, IL-15 was shown to support the proliferation and effector functions of CD8 T cells in the presence of regulatory T cells, suggesting that IL-15 could preserve the persistence and anticancer functions of T cells in the tumor microenvironment.[33] The ability of IL-15 to activate T-cell functions was further exploited in the context of T-cell therapy. Culture of TILs with IL-15 was indeed shown to improve the quality of CD8 T cells for adoptive therapy.[34] CAR T cells expressing the IL-15 gene featured greater expansion in vitro and reduced the cell death rate over control CAR T cells. Upon adoptive transfer in mice, IL-15 expressing CARs showed enhanced anticancer effects in vivo.[35] The clinical implementation of IL-15 began in 2009. The safety and efficacy of IL-15 is currently being tested in patients with lymphoma, melanoma, or renal cell carcinoma ([36] and NCT01572493, NCT01385423).

In addition to IL-15, IL-21 is also an immunomodulatory cytokine that is currently being tested for its anticancer activity in humans. Preclinical studies using IL-21-overexpressing tumors revealed that IL-21 prevented B16 melanoma and MCA205 carcinoma growth and increased mouse survival.[37] In addition, administration of IL-21 was found to control CD8 T-cell expansion and effector functions and to synergize with IL-15, leading to the rejection of large melanoma tumors in mice.[38] The ability of IL-21 to enhance T-cell functionality was shown in the context of adoptive transfer. CD8 T cells cultured with IL-21 enhanced their anticancer activity, leading to rejection of large tumors upon transfer.[39] Similarly, IL-21 enhances CAR T-cell anticancer functions for effective immunotherapy against B-cell malignancies.[40] We have also recently reported that naïve CD4 T cells differentiated into effector Th9 cells in the presence of TGF-β, IL-4 and IL-1β, secreted high levels of IL-21 and mediated IL-21-dependent anticancer effects against melanoma tumors upon adoptive transfer.[41] IL-21 mediated its anticancer activity through activation of NK and CD8 T cells which in turn controlled tumor progression through IFNγ. The clinical efficacy of IL-21 was investigated in phase I and II trials involving melanoma, renal cell carcinoma and metastatic colorectal cancer patients. In the phase II melanoma trial including 40 patients, the overall response rate to IL-21 was 22.5%, with 9 patients exhibiting partial responses and with 16 who had stable disease.[42]

Checkpoint Inhibitors

A balance between co-stimulatory and inhibitory signals regulates the amplitude and the quality of T-cell responses driven by TCR signaling. T cells require CD28-mediated co-stimulation (also known as signal 2) for the full acquisition of effector functions. However, excessive T-cell activation can result in the loss of self-tolerance, underscoring the importance of immune inhibitory pathways, or immune checkpoints, that regulate T-cell activity. The immunosuppressive tumor microenvironment directly affects the expression of immune checkpoint proteins, thereby favoring resistance to anti-tumor immune response. T cells are essential effectors for cancer immune surveillance, and inhibition of T-cell-dependent anti-tumor response can promote tumor progression.[43] Engagement of the CD28 homologue receptor cytotoxic T-lymphocyte-associated protein 4 (CTLA-4) on T cells by co-stimulatory molecules negatively regulates T-cell activation.[44] Leach et al.[45] have exploited this modulation of T-cell function therapeutically. They showed that administration of neutralizing CTLA-4 antibody into tumor-bearing mice resulted in tumor rejection.[45] In addition, mice that had rejected their tumors following anti-CTLA-4 treatment were protected against subsequent tumor rechallenge, indicating the establishment of immunological memory.[45] Additional mouse and human studies have validated these results and shown that CTLA-4 blockade triggers anticancer immune responses. Importantly, inhibition of CTLA-4 signaling not only enhances effector T-cell functions, but it also renders effector T cells insensitive to regulatory T-cell-driven suppression. Infusion of anti-CTLA-4 antibodies after vaccination with irradiated, autologous tumor cells secreting GM-CSF (GVAX)-induced anti-tumor immunity but no toxicity in metastatic melanoma patients.[46] The clinical efficacy of anti-CTLA-4 therapy was further confirmed in a phase III clinical trial where ipilimumab, a human mAb against CTLA-4, was shown to enhance the overall survival of metastatic melanoma patients.[47] The demonstrated anticancer activity of ipilimumab (Yervoy) led to its approval by the FDA for the treatment of metastatic melanoma.

Other key inhibitory checkpoints that are relevant in cancer immunotherapy include PD-1 and Tim-3. Expression of the PD-1 receptor is induced in T cells upon activation.[48] Tumor cells can drive T-cell dysfunction because of their expression of PD-1 receptor ligands, PD-L1 and PD-L2.[49–52] Iwai et al.[52] have shown that transgenic expression of PD-L1 in mastocytoma tumor cells prevented their elimination by CTL and enhanced their invasiveness in vivo. Thus, cancer tissues limit the host immune response through PD-1 ligands and their ligation to PD-1 on antigen-specific CD8 T cells, a phenomenon termed adaptive immune resistance. The molecular bases accounting for adaptive immune resistance remain elusive. However, it has been suggested that the therapeutic efficacy of PD-1 blockade is due to the restoration of CD8 T-cell effector function in the tumor microenvironment.[53] Preclinical models have demonstrated that blockade of PD-L1/PD-1 interactions could reinforce anticancer immune responses and promote tumor control.[51,52] In 2014, pembrolizumab and nivolumab, two anti-PD-1 antibodies, were approved by the FDA for the treatment of advanced melanoma patients (Table 1). Tim-3 is another T-cell inhibitory receptor that was initially identified on fully differentiated Th1 cells. The Tim-3 ligand, galectin-9, induces T-cell death.[64] In the tumor microenvironment, dysfunctional CD8 T cells could be identified by the co-expression of Tim-3 and PD-1. Importantly, the concomitant administration of neutralizing Tim-3 and PD-1 antibodies showed synergistic effects in preventing tumor outgrowth.[65] As Tim-3 and PD-1 expression are associated with tumor antigen-specific CD8+ T-cell dysfunction in melanoma patients and prevent the expansion of tumor antigen-specific CD8 T cells induced by vaccination,[66,67] evaluating the clinical efficacy of anti-Tim-3 antibodies in a clinical setting will be of high interest. Other therapies targeting immune checkpoints are currently in development such as agonist antibodies targeting molecules which activate T cells such as CD137 (BMS-663513),[68] OX40 (MEDI6383) NCT02221960, CD40 (CP870,893)[69] or GITR (TRAX518) NCT01239134 as well as drugs favoring DC activation such as LAG3-Fusion protein (IMP321).[70]

One of the challenging problems with the use of checkpoint inhibitors is the management of autoimmune side effects called immune-related adverse events (irAEs) (for detailed review see.[71,72] Mild to severe irAEs are observed with ipilimumab in about 60% of patients and in about 15% with anti-PD-1 drugs. irAEs include dermatologic, gastrointestinal, hepatic, endocrine, and other less common inflammatory events. IrAEs are believed to arise from general immunologic enhancement, and temporary immunosuppression with corticosteroids, tumor necrosis factor-alpha antagonists, mycophenolate mofetil or other agents can be an effective treatment in most cases. Interestingly, an association between irAEs and clinical outcome was observed for anti-CTLA-4 therapy.[73] The recent approval of anti-CTLA-4 and anti-PD1 mAbs in the clinic opens a new field in cancer immunotherapy. The discovery that anti-PD-1 mAb treatment could be effective in many types of cancers like melanoma, renal carcinoma, lung cancer, bladder cancer gastric cancer underscores the possible development of immune checkpoint inhibitors in many clinical contexts of solid tumors.

In addition to antibodies targeting checkpoint inhibitors, bispecific antibodies are being developed. These antibodies are artificial proteins that are composed of fragments of two different mAbs and consequently bind to two different types of antigens. This approach is used for cancer immunotherapy, where these proteins simultaneously bind to cytotoxic T cells using CD3 and a tumor cell target. Two drugs are currently available. Catumaxomab consists of one heavy chain and one light chain of an anti-EpCAM antibody and one heavy chain and one light chain of an anti-CD3 antibody as a consequence of which the chimeric protein can bind both EpCAM and CD3. In addition, the Fc-region can bind to an Fc receptor on accessory cells like other antibodies, which has led to calling the drug a trifunctional antibody. This structure allows both T cell and macrophage or DC activation to favor adaptive immune response and tumor cell lysis by immune effectors. This drug could be used to treat patients with ascites with EpCam + tumor cells. Blinatumomab is a bi-specific T-cell engager that combines two binding sites: a CD3 site for T cells and a CD19 site to target B cells. The drug works by linking these two cell types and activating the T cell to exert cytotoxic activity on the target cells. Blinatumomab could be used to target malignant B-cell lymphoma/leukemia and make blinatumomab a potential therapeutic option for pediatric and adult B-cell lymphoma or acute B-cell lymphoblastic leukemia.