During the past two decades, interleukin-12 (IL-12) has emerged as one of the most potent cytokines in mediating antitumor activity in a variety of preclinical models. Through pleiotropic effects on different immune cells that form the tumor microenvironment, IL-12 establishes a link between innate and adaptive immunity that involves different immune effector cells and cytokines depending on the type of tumor or the affected tissue. The robust antitumor response exerted by IL-12, however, has not yet been successfully translated into the clinics. The majority of clinical trials involving treatment with IL-12 failed to show sustained antitumor responses and were associated to toxic side effects. Here we discuss the therapeutic effects of IL-12 from preclinical to clinical studies, and will highlight promising strategies to take advantage of the antitumor activity of IL-12 while limiting adverse effects.
Interleukin-12 (IL-12) regulates inflammation by linking innate and adaptive immune responses. Most of the IL-12-induced effects are mediated by the secretion of interferon -γ.
IL-12 is a potent inducer of antitumor immunity in preclinical models.
The mechanisms by which IL-12 induces antitumor immune responses involve distinct effector cell types and cytokines depending on the tumor type and/or tumor location.
The delivery of IL-12 for therapeutic purposes focuses on novel methods to deliver this cytokine directly to the tumor site.
The robust antitumor response exerted by IL-12 in preclinical models has not yet been successfully translated into the clinics.
A better understanding of the multiple mechanisms by which IL-12 mediates tumor protection in different types of tumors or affected tissues.
Optimization of therapeutic schedules to locally deliver IL-12. Does it require a detailed knowledge of the molecular pathology of each individual tumor at a particular time?
Revisiting the use of IL-12 as an adjuvant in combinatorial treatments. Do we need to inhibit particular immunosuppressive mechanisms to improve clinical benefits?
How can we achieve durable, local, non-toxic antitumor responses with IL-12 in cancer patients? What is the best strategy to deliver this cytokine into the tumor microenvironment in a controlled manner?
The Biology of IL-12
Cytokines are among the chief players in controlling immune responses, and cytokine-based approaches for cancer therapy have been pursued in a number of ways. In that respect, the immunomodulatory cytokine IL-12, a key member of the IL-12 family of cytokines, emerged as a potent inducer of antitumor immunity. IL-12 was originally identified in 1989 as a natural killer (NK) cell-stimulatory factor with multiple biologic effects on peripheral blood lymphocytes.1 It is mainly produced by antigen-presenting cells (APCs) such as dendritic cells (DCs), monocytes, macrophages and B cells upon Toll-like receptor engagement.2 Thus, IL-12 is secreted as an early pro-inflammatory cytokine in response to infections.3 Additional amplifying signals such as interferon-γ (IFN-γ),4 IL-155 or cluster of differentiation (CD)40L–CD40 cell–cell interactions6 are necessary for the optimal production of biologically active IL-12. Conversely, IL-12 is negatively regulated through cytokines such as IL-10 and transforming growth factor-β1 (TGF-β1).7, 8 IL-12 is a heterodimer with a molecular weight of 70 kDa consisting of a heavy (p40) and a light (p35) chain subunit, which are covalently linked by disulfide bonds.9, 10, 11 While p40 is produced in abundance by phagocytic cells, p35 is ubiquitously and constitutively expressed only at low levels and is thought to require p40 co-expression for secretion of the biologically active cytokine.12
The sensing of IL-12 is mediated through the heterodimeric IL-12 receptor (IL-12R) composed of IL-12Rβ1 and IL-12Rβ2.13 Co-expression of both receptor subunits is required for the generation of high-affinity binding sites for IL-12. The IL-12R complex is found on NK cells, NK T and activated T cells14 but has also been detected on cell types of myeloid origin15 and tonsillar B cells.16 Naive T cells express IL-12Rβ1 but not IL-12Rβ2, which is critical for the signal transduction downstream of the receptor complex.17 Upon activation of T cells via the T-cell receptor, both IL-12 receptor chains are induced, which is additionally enhanced by IL-12 itself, IFN-γ, tumor necrosis factor-α (TNF-α) and anti-CD28 costimulation.18, 19 Successful triggering of the receptor activates the Janus kinase–STAT (signal transducer and activator of transcription) signaling pathway, predominantly leading to STAT4 phosphorylation, which mediates subsequent cellular responses.20, 21
IL-12 Sensing by Innate and Adaptive Lymphocytes
IL-12 has a key role in the regulation of inflammation by linking innate and adaptive immune responses. IL-12 release by microbe-sensing APCs results in subsequent activation and proliferation of NK and T cells and promotes their effector functions by inducing the transcription of cytokines and cytolytic factors such as perforin and granzyme B.22, 23, 24 Moreover, IL-12 polarizes T cells into a type 1 helper T (Th1) effector cell phenotype.25, 26, 27 Th1 polarization is further pronounced by IL-12 inhibiting the developmental program of type 2 helper T cells28 and interference with the differentiation of regulatory T cells (Tregs) and Th17 cells induced by TGF-β.29 Additionally, IL-12 programs effector T cells for optimal generation of effector memory T cells and T follicular helper cells.30, 31 Direct effects of IL-12 on APCs have also been reported. Even though the activation of IL-12R in these cells did not involve the canonical STAT pathway, it increased their ability to present poorly immunogenic tumor peptides.32, 33
A central mediator of IL-12-induced responses is IFN-γ, which is secreted upon IL-12 stimulation alone or with synergizing factors such as IL-2 and IL-18.34, 35 IFN-γ, in turn, acts on APCs to initiate or increase IL-12 secretion in a positive feedback loop.36 Apart from IFN-γ release, IL-12 triggers the secretion of a number of other factors, including TNF-α, granulocyte-macrophage colony-stimulating factor (GM-CSF) and IL-2.37
Mechanisms of Tumor Protection through IL-12
Whereas the role of IL-12 was primarily studied in the context of infection and autoimmunity, mice lacking the IL-12 specific subunit p35 developed significantly increased numbers of chemically induced papillomas38 and were more susceptible to N-methyl-N-nitrosourea-induced T-cell lymphomas39 compared to wild-type mice. In addition, IL-12p40 deficiency resulted in earlier appearance of 3-methylcholanthrene (MCA)-induced sarcomas, compared to wild-type mice.40 The observation that animals deficient for IL12Rβ2 developed spontaneous tumors at higher frequencies and showed enhanced growth of transplantable tumors confirmed the importance of IL-12 signaling in tumor protection.41 In humans, polymorphisms in the 3′-untranslated region of IL12A lead to decreased IL-12 production, which coincides with an increased susceptibility to develop glioblastomas.42 Despite the remarkable anticancer activity exerted by IL-12, other members of the IL-12 family play critical roles in the regulation of tumor development (see Box 1).
To determine the mechanisms by which IL-12 induces antitumor immune responses, cancer cells have been engineered to continuously release this cytokine. The overexpression of IL-12 in B16 melanoma, thricostatin-A (TSA) mammary adenocarcinoma and C26 colon carcinoma cells induced tumor suppression upon subcutaneous (s.c.) inoculation.43, 44, 45, 46 Whereas in B16 melanoma this effect was mediated by a subset of innate lymphoid cells (ILCs), the rejection of breast cancer TSA-IL-12 cells was dependent on CD8 cytotoxic T cells, secreting IFN-γ.43 However, tumor rejection in IL-12-transduced C26 colon carcinoma was partially dependent on GM-CSF-producing CD4 T cells or NK cells, but independent of IFN-γ.46 IL-12-responsive T cells were also ascribed a major role in rejection of tumors of the central nervous system.47, 48 These results indicate that differences in tumor cell type and tumor location determine the quality of the tumor immune response, involving distinct effector cell types and cytokines.
Although tumor-specific immune responses are pivotal for cancer control, changes in the vasculature status are also relevant for tumor development. Hence, overexpression of IL-12 in B16 melanoma tumors was shown to regulate the tumor vasculature, either by upregulating of adhesion molecules that may facilitate leukocyte recruitment44 or by inhibiting angiogenesis in an IFNγ-dependent manner.49, 50 The inhibitory effects of IL-12 on the tumor vessels have been associated with increased levels of the IFNγ-inducible chemokine (C-X-C motif) ligand (CXCL) 9 and CXCL10, and a decreased production of vascular endothelial growth factor (VEGF) and metalloproteinase-9.51, 52, 53, 54 In general, these studies highlight the importance of IL-12-induced IFN-γ in the control of tumor growth.
The main effects of IL-12 on the tumor microenvironment have been summarized in Figure 1.
Therapeutic Effects of IL-12 in Preclinical Models
The therapeutic potential of IL-12 has been extensively investigated in various preclinical models of cancer. Especially systemic intravenous (i.v.), intraperitoneal (i.p.), subcutaneous but also intratumoral (i.t.) delivery of recombinant IL-12 leads to reduced or delayed tumor growth and increased survival in transplantable, carcinogen-induced and spontaneous tumors arising in genetically modified mice.55, 56, 57 It is by now clear that the antitumor effectiveness of IL-12 is dose and context dependent.40, 55, 58 Delivery of IL-12 for therapeutic purposes has thus far been accomplished through direct infusion of the recombinant protein, by gene therapy using viral and non-viral vectors, electroporation, by IL-12-containing microspheres and nanoparticles or by the transfer of IL-12-overexpressing stromal and immune cell types (Figure 2).
Already in 1993, Brunda et al.59 reported an antitumor response of recombinant IL-12 in B16 melanoma, M5975 sarcoma and RENCA renal cell carcinoma when administered intraperitoneally. The antitumor effects of IL-12 were even observed when treatment was started at later stages after tumor inoculation and were found to be partially dependent on CD8 T cells. In a later study, however, both NK and NK T cells were reported to be the cell types responsible for IL-12-induced antitumor responses in a variety of transplantable tumor models including B16 melanoma.40 Interestingly, discrepancies between the relative roles of each cell population in suppressing tumor growth could be attributed to the dose and time of IL-12 administration.40 I.p. administration of IL-12 also induced effective antitumor immune responses against malignant glioma60 and 4T1 mammary carcinoma.61 Apart from direct effects on primary tumors, IL-12 was also able to eradicate lung metastasis when mammary tumors were surgically removed.61
The potent antitumor activity exerted by IL-12 made it an ideal candidate for combination with other therapy modalities directed to increase the immunogenicity of the tumor. In this respect, the combination of IL-12 with cytokines, chemotherapeutic agents, multipeptide vaccines and monoclonal antibodies potentiated the therapeutic activity of this cytokine in a variety of tumor models such as melanoma, bladder carcinoma and mammary carcinoma.62, 63, 64, 65, 66, 67 Even though the rationale for combining IL-12 with other cytokines was to achieve complementary and more durable immune-stimulating responses, most of these treatments resulted in high levels of systemic IFNγ production and therefore a potential degree of toxicity after translating them into the clinics.65 In combination with chemotherapy, an improved antitumor activity could only be observed in immunogenic tumors when IL-12 was administered early after chemotherapy, thus highlighting the importance of the timing for immune intervention in chemotherapy-induced antitumor responses.68 An example of a successful combinatorial treatment with IL-12 was reported in human epidermal growth factor receptor (HER)-2/neu transgenic mice, where the treatment with IL-12 together with tamoxifen or HER-2/neu multipeptide vaccines resulted in an effective prevention of tumor growth.66, 67 An enhanced tumor regression was also achieved upon co-administration of IL-12 and the anti-HER-2 antibody trastuzumab in colon adenocarcinoma.69
Even though systemic delivery of IL-12 showed great potential as an experimental anticancer agent, the instability and short half-life of this cytokine after bolus administration led to focus on novel methods to deliver it directly to the tumor site. Following this strategy, vom Berg et al.48 implanted osmotic minipumps to locally deliver IL-12 into the brain of GL-261 glioma-bearing mice. Notably, the combined treatment of IL-12 with systemic blockade of the co-inhibitory receptor cytotoxic T-lymphocyte antigen 4 (CTLA-4) eradicated even very advanced tumors at late disease stages in a T cell-dependent manner. On the basis of this evidence, the rationale of combining IL-12 with the targeting of regulatory pathways holds a great potential to overcome tumor-associated immune suppression.
Specific delivery of IL-12 to the tumor site was also achieved through gene therapy. Thus, delivery of IL-12 by electroporation or viral-based strategies reduced growth of established colon carcinoma, melanoma and brain tumors.70, 71, 72, 73, 74 In the case of breast cancer, the success of IL-12 gene therapy was dependent on the immunogenicity of the tumor. Whereas IL-12 led to regression of highly immunogenic TS/A tumors, the growth of 4T1 tumors, which are considered less immunogenic, was not affected.75 In the 4T1 model, however, the treatment resulted in a marked reduction of lung metastasis, an effect that was partially dependent on IFNγ-producing NK cells.75, 76 Despite most of these strategies provided a continuous release of IL-12 and IFNγ within the tumor, just a few of them reported that IL-12 was acting locally by showing low serum levels of these cytokines.71, 74 Nevertheless, synergistic antitumor responses in breast cancer have been achieved by combining local adenovirus-mediated gene transfer of IL-12 with T-cell chemoattractants (lymphotactin, CXCL10), costimulatory molecules (B7.1, glucocorticoid induced TNF receptor ligand, 4-1BB ligands), GM-CSF, radiotherapy or antiangiogenic therapy.77, 78, 79, 80, 81, 82, 83 Furthermore, the combination with chemotherapeutic or antiangiogenic agents in lung, skin and colorectal cancer, or with antiangiogenic agents in prostate cancer proved to be more efficient than viral-mediated IL-12 gene therapy alone in different types of tumors.84, 85, 86, 87, 88, 89 Even though local delivery of IL-12 by gene therapy resulted in a more sustained expression of IL-12 in comparison with the levels obtained by injecting the recombinant protein, the lack of selectivity and the occurrence of non-specific immune responses associated with the use of viral vectors remain a major concern when using this strategy to deliver IL-12.
In order to overcome these limitations, IL-12 gene was delivered to the tumor site embedded in biodegradable polymeric microspheres and nanoparticles. This method results in enhanced cellular uptake, tissue penetrability and escape from endolysosomal compartments. Tumor regression was observed in transplantable tumor models upon i.t. injection of biodegradable polylactic miscrospheres loaded with IL-12 alone90 or in combination with TNF-α, IL-18 or GM-CSF.91, 92, 93 An effective strategy for treating malignant glioma was described by Sonabend et al.,94 using modified polyethylenimine complexes as vehicles for IL-12 gene therapy. Of note, the synergy resulting from combining this treatment with chemotherapy resulted in 100% survival of treated mice.94
The transfer of cells engineered to produce IL-12 also proved to be successful in inducing long-term antitumor immunity. Early studies in this field reported the capacity of IL-12-secreting fibroblasts to delay tumor growth and eradicate established sarcomas.95, 96, 97 In the case of CT26 colon carcinoma, however, the inhibition of tumor growth by unpulsed IL-12-transduced DCs was significantly better than the one achieved by using IL-12-transduced fibroblasts or the IL-12 gene-encoding adenovirus itself.98 These approaches were followed by the development of therapeutic vaccines based on IL-12-expressing DCs that were additionally pulsed with tumor cell lysates or peptides. This method, used for colon carcinoma and melanoma, induced more potent tumor-specific T-cell responses than vaccination in the absence of IL-12.99, 100
Alternative ways of cell-mediated delivery of IL-12 have taken advantage of the tumor-homing capacity of mesenchymal stem cells101 or transfer of tumor-infiltrating lymphocytes. The latter approach has been used to deliver CD8+ T cells specific for melanoma antigens102, 103, 104 or engineered to express a chimeric antigen receptor (CAR) against CD19 in B-cell lymphomas.105 The administered IL-12 was found to activate myeloid cells by increasing the expression of Fas and cross-presentation, leading to the stimulation of tumor antigen-specific CD8 T cells and regression of established tumors102, 106 (Figure 1). More recently, the development of novel approaches that direct IL-12 activity to the tumor site focus on immunocytokines, for example, the fusion of the cytokine to an antibody that binds specifically to the tumor vasculature,107, 108 or to exposed deoxyribonucleic acid (DNA) in the necrotic core of a tumor.109 The targeting of necrotic areas within the tumor is of special interest due to the lack of perfusion of solid tumors and subsequent cell death.109 The use of antibody-targeted cytokines, however, needs to be carefully evaluated for each specific tumor context, since a high avidity and retention to the targeted tissue are essential for efficient therapeutic effects. In a combinatorial approach, a dual cytokine–antibody fusion protein that simultaneously targeted IL-12 and IL-2 to CD30+ lymphoma cells suppressed tumor growth more efficiently than by just targeting IL-12 or IL-2 alone.110
IL-12 to Treat Human Cancer
The potent antitumor effects of IL-12 in preclinical models justified the translation of this approach to a clinical setting. Unfortunately, systemic i.v. administration of recombinant IL-12 not only demonstrated poor efficacy but also caused severe adverse effects. Early studies evaluated the safety of i.v. or s.c. injected IL-12 in patients with metastatic renal carcinoma, melanoma, colon carcinoma, recurrent ovarian cancer, and neck and head carcinoma.111, 112, 113, 114, 115, 116, 117, 118 The goal was to administer IL-12 in a schedule that minimized common toxicities associated with cytokine therapy such as fever, fatigue, hematological changes or hyperglycemia. In general, the best way to administer IL-12 appeared to be in cycles consisting of either i.v. boluses for five consecutive days or s.c. injections for two consecutive weeks. Even though these trials established maximum tolerated doses for IL-12 for the different schedules, treatment response rates were not very promising, with only few cases of partial or complete responses (Table 1).111, 113, 114, 115, 118 Moreover, the systemic administration of IL-12 displayed schedule-dependent toxicity, which appeared to be reduced when a single test dose of IL-12 was administered i.v. 2 weeks before initiation of the scheduled daily treatment cycle.111, 115 The combination of IL-12 therapy with active vaccination against tumor-associated antigens119 or IFN-α120 did not further improve clinical responses in malignant melanoma or renal cell carcinoma. Even in patients with metastatic HER2+ breast cancer, where the combination of IL-12 with paclitaxel and trastuzumab resulted in a 52% rate of clinical benefit in a phase I trial, the combinatorial strategy was not further pursued.121 More encouraging results, however, were obtained when treating hematological cancers (Table 1). A phase I dose escalation trial with s.c. delivery of IL-12 twice a week for up to 24 weeks resulted in an overall response rate of 56% in cutaneous T-cell lymphoma patients.122 When the s.c. treatment was applied to patients with mycosis fungoide who had failed previous treatments, 43% of partial response to the treatment was observed.123 Promising results were also obtained for refractory non-Hodgkin’s B-cell lymphoma. Here, s.c. treatment with IL-12 led to partial or complete response in 21% of the patients, and almost 50% showed stable disease.124 This clinical response was further improved when the therapy was combined with the anti-CD20 antibody rituximab, achieving partial response in 25% and 42% of the patients, respectively.125 Even though the treatment with IL-12 in most of these patients was well tolerated, a repeated administration evoked increased levels of IL-10, which presumably neutralized the antitumor effects of IL-12.125, 126
I.t. delivery of IL-12 to cancer patients was pursued by gene therapy approaches (Table 1). In patients with malignant melanoma, i.t. delivery of a plasmid encoding IL-12 led to some beneficial clinical effect at the tumor site and even in non-treated lesions.127, 128, 129 The treatment seemed however less efficient when IL-12 was delivered encoded by a vector derived from a highly attenuated strain of the canarypox virus.130 Viral delivery of IL-12 was also used in patients with advanced digestive cancer, but only led to mild antitumor effects.131 Even though the treatment was well tolerated, adverse reactions were associated to vector injection.131 Hence, a conscious manipulation of the balance between antivirus and antitumor responses is of special importance when using oncolytic viruses for IL-12-based immunotherapy.132 In order to minimize toxicity, Rudman et al.133 delivered IL-12 to the tumor site by fusing it to the humanized antibody targeting the ED-B variant of fibronectin (AS1409). Despite the authors not confirming the targeting of this immunocytokine to the tumor, the treatment led to stable disease associated to moderate toxicity in 46% of patients with malignant melanoma.133 Low levels of toxicity were also achieved when delivering an IL-12 plasmid formulated with a synthetic lipopolymer (EGEN-001) to patients with ovarian cancer.134 The evaluation of EGEN-001 in a subsequent Phase II trial, however, only showed limited activity as well as increased toxicity in patients resistant to platinum.135
Taken together, systemic administration of IL-12 was tested extensively and failed, barring all future attempts in this direction. Local application, however, had a promising safety profile at similar or better antitumor efficacy (see all studies above) and warrants further investigation. Current efforts are evaluating some of the most successful approaches to deliver IL-12 intratumorally in different types of solid tumors. For instance, in situ electroporation of a plasmid coding for IL-12 is being tested into cancerous lesions of the skin such as malignant melanomas, cutaneous T-cell lymphomas and Merkel cell carcinoma (NCT01502293, NCT01440816, NCT01579318). Furthermore, IL-12-based immunocytokines are still in clinical development. First, the immunocytokine NHS-IL-12, consisting of two molecules of IL-12 fused to an anti-DNA human IgG1 antibody that presumably targets necrotic areas of the tumor,109 is being evaluated in a phase I trial for metastatic solid tumors (NCT01417546). Second, AS1409 is currently in clinical trials for renal cell carcinoma and malignant melanoma (NCT00625768). The biopolymer encoding for IL-12, EGEN-001, which only showed limited success when administered as monotherapy, is going to be tested in combination with chemotherapy in the context of ovarian cancer (NCT01489371) and colorectal carcinoma (NCT01300858). Finally, a novel two-component inducible gene expression system in which adenoviral IL-12 expression is controlled by an orally bioavailable small-molecule activator ligand is being currently tested. This approach is in clinical phase I trials for advanced melanoma (NCT01397708) and glioma (NCT02026271), as well as in phase II evaluation in metastatic breast cancer alone or in combination with chemotherapy (NCT01703754).
Even though some of the above-mentioned strategies to deliver IL-12 in cancer patients have shown promising preliminary results, until now IL-12-based immunotherapy has not achieved Food and Drug Administration (FDA) approval. In clear contrast, two other cytokines such as IL-2 and IFN-α were approved by the FDA as single agents for metastatic melanoma and renal cell carcinoma, and for the adjuvant treatment of high-risk melanoma, respectively.136 The success of high-dose IL-2, for instance, was associated to durable overall clinical responses, but the administration of the cytokine has to be carefully managed to avoid toxicities such as capillary leak syndrome.137 In the case of IFN-α, its use as an adjuvant therapy for stage III and IV melanoma still remains controversial. Even though two clinical trials launched in 1995 demonstrated longer relapse-free survival and overall survival rates when using this adjuvant in a high dose, the results obtained in subsequent trials were not so clear and failed to clarify the mechanism of action of this drug in the treatment of melanoma.138 Given the fact that tumor-targeted IL-12 acts primarily within the tumor microenvironment, as opposed to the activating systemic immune responses induced by IFN-α or IL-2, we believe that IL-12-based therapies will emerge primarily as combination therapies. We speculate that IL-12 alters the tumor microenvironment to enhance immunogenicity, which can then be further exploited through the use of check-point blockade or other strategies to enhance antitumor responses.
The potential of cytokines for cancer immunotherapy has been extensively investigated. In the case of IL-12, its potent antitumor properties were already observed more than 20 years ago upon systemic administration of the cytokine in various transplantable cancer models.59 Since then, several studies aimed to evaluate the use of IL-12 for therapeutical purposes by specifically delivering this cytokine within the tumor site. Even though several of these approaches resulted in impressive antitumor responses, the translation into the clinics was sobering. The reasons for that are still being discussed in the oncology field. On the one hand, the schedule optimization for therapeutic IL-12 delivery in clinical trials has proved to be challenging. Even though the most successful way to administer IL-12 appeared to be in cycles of twice weekly injections, repeated administration of the cytokine could contribute to increase the immunosuppressive properties of the tumor by the induction of IL-10.139, 140 On the other hand, the use of IL-12 as an adjuvant in combinatorial treatments requires a detailed knowledge of the molecular pathology of each individual tumor in order to achieve clinical benefits. In this respect, the combination of IL-12 with therapies that block the type of immunosuppressive activity characteristic of the different tumor models could be of potential use. Finally, durable, non-toxic anti-cancer responses with IL-12 will likely only be achieved with a controlled and tumor-targeted delivery of the cytokine. Several of these approaches (EGEN-001, the immunocytokine NHS-IL-12, the control of IL-12 expression by an orally activator ligand or the CAR-modified IL-12-expressing T cells) are already advancing in clinical trials. Clearly, as we only now start to understand the multiple mechanisms by which IL-12 mediates tumor protection in more detail, it is time to revisit the use of IL-12 in clinical studies. Blind systemic administration of IL-12 will not be pursued in the future, but tumor-targeted IL-12 delivery combined with radiation-, chemo- and immunotherapy, respectively, holds great promise for the future of cancer immunotherapy.
chimeric antigen receptor
chemokine (C-X-C motif) ligand 9
chemokine (C-X-C motif) ligand 10
cluster of differentiation
Food and Drug Administration
granulocyte-macrophage colony stimulating factor
human epidermal growth factor receptor 2
signal transducer and activator of transcription
T helper 17 cell
regulatory T cell
transforming growth factor-β1
type 1 helper T cell
vascular endothelial growth factor
Kobayashi M, Fitz L, Ryan M, Hewick RM, Clark SC, Chan S et al. Identification and purification of natural killer cell stimulatory factor (NKSF), a cytokine with multiple biologic effects on human lymphocytes. J Exp Med 1989; 170: 827–845.
Trinchieri G, Rengaraju M, D'Andrea A, Valiante NM, Kubin M, Aste M et al. Producer cells of interleukin 12. Parasitol Today 1993; 9: 97.
Medzhitov R . Toll-like receptors and innate immunity. Nat Rev Immunol 2001; 1: 135–145.
Ma X, Chow JM, Gri G, Carra G, Gerosa F, Wolf SF et al. The interleukin 12 p40 gene promoter is primed by interferon gamma in monocytic cells. J Expl Med 1996; 183: 147–157.
Kuwajima S, Sato T, Ishida K, Tada H, Tezuka H, Ohteki T . Interleukin 15-dependent crosstalk between conventional and plasmacytoid dendritic cells is essential for CpG-induced immune activation. Nat Immunol 2006; 7: 740–746.
Schulz O, Edwards AD, Schito M, Aliberti J, Manickasingham S, Sher A et al. CD40 triggering of heterodimeric IL-12 p70 production by dendritic cells in vivo requires a microbial priming signal. Immunity 2000; 13: 453–462.
D'Andrea A, Aste-Amezaga M, Valiante NM, Ma X, Kubin M, Trinchieri G . Interleukin 10 (IL-10) inhibits human lymphocyte interferon gamma-production by suppressing natural killer cell stimulatory factor/IL-12 synthesis in accessory cells. J Exp Med 1993; 178: 1041–1048.
Du C, Sriram S . Mechanism of inhibition of LPS-induced IL-12p40 production by IL-10 and TGF-beta in ANA-1 cells. J Leukoc Biol 1998; 64: 92–97.
Gately MK, Desai BB, Wolitzky AG, Quinn PM, Dwyer CM, Podlaski FJ et al. Regulation of human lymphocyte proliferation by a heterodimeric cytokine, IL-12 (cytotoxic lymphocyte maturation factor). J Immunol 1991; 147: 874–882.
Gubler U, Chua AO, Schoenhaut DS, Dwyer CM, McComas W, Motyka R et al. Coexpression of two distinct genes is required to generate secreted bioactive cytotoxic lymphocyte maturation factor. Proc Natl Acad Sci USA 1991; 88: 4143–4147.
Schoenhaut DS, Chua AO, Wolitzky AG, Quinn PM, Dwyer CM, McComas W et al. Cloning and expression of murine IL-12. J Immunol 1992; 148: 3433–3440.
Babik JM, Adams E, Tone Y, Fairchild PJ, Tone M, Waldmann H . Expression of murine IL-12 is regulated by translational control of the p35 subunit. JImmunol 1999; 162: 4069–4078.
Rogge L, Papi A, Presky DH, Biffi M, Minetti LJ, Miotto D et al. Antibodies to the IL-12 receptor beta 2 chain mark human Th1 but not Th2 cells in vitro and in vivo. J Immunol 1999; 162: 3926–3932.
Presky DH, Yang H, Minetti LJ, Chua AO, Nabavi N, Wu CY et al. A functional interleukin 12 receptor complex is composed of two beta-type cytokine receptor subunits. Proc Natl Acad Sci USA 1996; 93: 14002–14007.
Grohmann U, Belladonna ML, Bianchi R, Orabona C, Ayroldi E, Fioretti MC et al. IL-12 acts directly on DC to promote nuclear localization of NF-kappaB and primes DC for IL-12 production. Immunity 1998; 9: 315–323.
Airoldi I, Gri G, Marshall JD, Corcione A, Facchetti P, Guglielmino R et al. Expression and function of IL-12 and IL-18 receptors on human tonsillar B cells. J Immunol 2000; 165: 6880–6888.
Zou J, Presky DH, Wu CY, Gubler U . Differential associations between the cytoplasmic regions of the interleukin-12 receptor subunits beta1 and beta2 and JAK kinases. J Biol Chem 1997; 272: 6073–6077.
Afkarian M, Sedy JR, Yang J, Jacobson NG, Cereb N, Yang SY et al. T-bet is a STAT1-induced regulator of IL-12R expression in naive CD4+ T cells. Nat Immunol 2002; 3: 549–557.
Szabo SJ, Dighe AS, Gubler U, Murphy KM . Regulation of the interleukin (IL)-12R beta 2 subunit expression in developing T helper 1 (Th1) and Th2 cells. J Exp Med 1997; 185: 817–824.
Bacon CM, McVicar DW, Ortaldo JR, Rees RC, O'Shea JJ, Johnston JA . Interleukin 12 (IL-12) induces tyrosine phosphorylation of JAK2 and TYK2: differential use of Janus family tyrosine kinases by IL-2 and IL-12. J Exp Med 1995; 181: 399–404.
Bacon CM, Petricoin EF 3rd, Ortaldo JR, Rees RC, Larner AC, Johnston JA et al. Interleukin 12 induces tyrosine phosphorylation and activation of STAT4 in human lymphocytes. Proc Natl Acad Sci USA 1995; 92: 7307–7311.
Aste-Amezaga M, D'Andrea A, Kubin M, Trinchieri G . Cooperation of natural killer cell stimulatory factor/interleukin-12 with other stimuli in the induction of cytokines and cytotoxic cell-associated molecules in human T and NK cells. Cell Immunol 1994; 156: 480–492.
Perussia B, Chan SH, D'Andrea A, Tsuji K, Santoli D, Pospisil M et al. Natural killer (NK) cell stimulatory factor or IL-12 has differential effects on the proliferation of TCR-alpha beta+, TCR-gamma delta+ T lymphocytes, and NK cells. J Immunol 1992; 149: 3495–3502.
Salcedo TW, Azzoni L, Wolf SF, Perussia B . Modulation of perforin and granzyme messenger RNA expression in human natural killer cells. J Immunol 1993; 151: 2511–2520.
Hsieh CS, Macatonia SE, Tripp CS, Wolf SF, O'Garra A, Murphy KM . Development of TH1 CD4+ T cells through IL-12 produced by Listeria-induced macrophages. Science 1993; 260: 547–549.
Manetti R, Parronchi P, Giudizi MG, Piccinni MP, Maggi E, Trinchieri G et al. Natural killer cell stimulatory factor (interleukin 12 [IL-12]) induces T helper type 1 (Th1)-specific immune responses and inhibits the development of IL-4-producing Th cells. J Exp Med 1993; 177: 1199–1204.
Micallef MJ, Ohtsuki T, Kohno K, Tanabe F, Ushio S, Namba M et al. Interferon-gamma-inducing factor enhances T helper 1 cytokine production by stimulated human T cells: synergism with interleukin-12 for interferon-gamma production. Eur J Immunol 1996; 26: 1647–1651.
Djuretic IM, Levanon D, Negreanu V, Groner Y, Rao A, Ansel KM . Transcription factors T-bet and Runx3 cooperate to activate Ifng and silence Il4 in T helper type 1 cells. Nat Immunol 2007; 8: 145–153.
Prochazkova J, Pokorna K, Holan V . IL-12 inhibits the TGF-beta-dependent T cell developmental programs and skews the TGF-beta-induced differentiation into a Th1-like direction. Immunobiology 2012; 217: 74–82.
Chowdhury FZ, Ramos HJ, Davis LS, Forman J, Farrar JD . IL-12 selectively programs effector pathways that are stably expressed in human CD8+ effector memory T cells in vivo. Blood 2011; 118: 3890–3900.
Schmitt N, Bustamante J, Bourdery L, Bentebibel SE, Boisson-Dupuis S, Hamlin F et al. IL-12 receptor beta1 deficiency alters in vivo T follicular helper cell response in humans. Blood 2013; 121: 3375–3385.
Bianchi R, Grohmann U, Vacca C, Belladonna ML, Fioretti MC, Puccetti P . Autocrine IL-12 is involved in dendritic cell modulation via CD40 ligation. J Immunol 1999; 163: 2517–2521.
Grohmann U, Bianchi R, Ayroldi E, Belladonna ML, Surace D, Fioretti MC et al. A tumor-associated and self antigen peptide presented by dendritic cells may induce T cell anergy in vivo, but IL-12 can prevent or revert the anergic state. J Immunol 1997; 158: 3593–3602.
Chan SH, Perussia B, Gupta JW, Kobayashi M, Pospisil M, Young HA et al. Induction of interferon gamma production by natural killer cell stimulatory factor: characterization of the responder cells and synergy with other inducers. J Exp Med 1991; 173: 869–879.
Okamura H, Tsutsui H, Kashiwamura S, Yoshimoto T, Nakanishi K . Interleukin-18: a novel cytokine that augments both innate and acquired immunity. Adv Immunol 1998; 70: 281–312.
Grohmann U, Belladonna ML, Vacca C, Bianchi R, Fallarino F, Orabona C et al. Positive regulatory role of IL-12 in macrophages and modulation by IFN-gamma. J Immunol 2001; 167: 221–227.
Trinchieri G . Interleukin-12 and the regulation of innate resistance and adaptive immunity. Nat Rev Immunol 2003; 3: 133–146.
Meeran SM, Mantena SK, Meleth S, Elmets CA, Katiyar SK . Interleukin-12-deficient mice are at greater risk of UV radiation-induced skin tumors and malignant transformation of papillomas to carcinomas. Mol Cancer Ther 2006; 5: 825–832.
Liu J, Xiang Z, Ma X . Role of IFN regulatory factor-1 and IL-12 in immunological resistance to pathogenesis of N-methyl-N-nitrosourea-induced T lymphoma. Journal of immunology 2004; 173: 1184–1193.
Smyth MJ, Taniguchi M, Street SE . The anti-tumor activity of IL-12: mechanisms of innate immunity that are model and dose dependent. J Immunol 2000; 165: 2665–2670.
Airoldi I, Di Carlo E, Cocco C, Sorrentino C, Fais F, Cilli M et al. Lack of Il12rb2 signaling predisposes to spontaneous autoimmunity and malignancy. Blood 2005; 106: 3846–3853.
Zhao B, Meng LQ, Huang HN, Pan Y, Xu QQ . A novel functional polymorphism, 16974 A/C, in the interleukin-12-3' untranslated region is associated with risk of glioma. DNA Cell Biol 2009; 28: 335–341.
Cavallo F, Signorelli P, Giovarelli M, Musiani P, Modesti A, Brunda MJ et al. Antitumor efficacy of adenocarcinoma cells engineered to produce interleukin 12 (IL-12) or other cytokines compared with exogenous IL-12. J Natl Cancer Inst 1997; 89: 1049–1058.
Eisenring M, vom Berg J, Kristiansen G, Saller E, Becher B . IL-12 initiates tumor rejection via lymphoid tissue-inducer cells bearing the natural cytotoxicity receptor NKp46. Nat Immunol 2010; 11: 1030–1038.
Martinotti A, Stoppacciaro A, Vagliani M, Melani C, Spreafico F, Wysocka M et al. CD4 T cells inhibit in vivo the CD8-mediated immune response against murine colon carcinoma cells transduced with interleukin-12 genes. Eur J Immunol 1995; 25: 137–146.
Zilocchi C, Stoppacciaro A, Chiodoni C, Parenza M, Terrazzini N, Colombo MP . Interferon gamma-independent rejection of interleukin 12-transduced carcinoma cells requires CD4+ T cells and Granulocyte/Macrophage colony-stimulating factor. J Exp Med 1998; 188: 133–143.
Vetter M, Hofer MJ, Roth E, Pircher HP, Pagenstecher A . Intracerebral interleukin 12 induces glioma rejection in the brain predominantly by CD8+ T cells and independently of interferon-gamma. J Neuropathol Exp Neurol 2009; 68: 525–534.
Vom Berg J, Vrohlings M, Haller S, Haimovici A, Kulig P, Sledzinska A et al. Intratumoral IL-12 combined with CTLA-4 blockade elicits T cell-mediated glioma rejection. J Exp Med 2013; 210: 2803–2811.
Gerber SA, Moran JP, Frelinger JG, Frelinger JA, Fenton BM, Lord EM . Mechanism of IL-12 mediated alterations in tumour blood vessel morphology: analysis using whole-tissue mounts. Br J Cancer 2003; 88: 1453–1461.
Sorensen EW, Gerber SA, Frelinger JG, Lord EM . IL-12 suppresses vascular endothelial growth factor receptor 3 expression on tumor vessels by two distinct IFN-gamma-dependent mechanisms. Journal of Immunol 2010; 184: 1858–1866.
Dias S, Boyd R, Balkwill F . IL-12 regulates VEGF and MMPs in a murine breast cancer model. Int J Cancer 1998; 78: 361–365.
Kanegane C, Sgadari C, Kanegane H, Teruya-Feldstein J, Yao L, Gupta G et al. Contribution of the CXC chemokines IP-10 and Mig to the antitumor effects of IL-12. J Leukoc Biol 1998; 64: 384–392.
Mitola S, Strasly M, Prato M, Ghia P, Bussolino F . IL-12 regulates an endothelial cell-lymphocyte network: effect on metalloproteinase-9 production. J Immuno 2003; 171: 3725–3733.
Sgadari C, Angiolillo AL, Tosato G . Inhibition of angiogenesis by interleukin-12 is mediated by the interferon-inducible protein 10. Blood 1996; 87: 3877–3882.
Colombo MP, Trinchieri G . Interleukin-12 in anti-tumor immunity and immunotherapy. Cytokine Growth Factor Rev 2002; 13: 155–168.
Noguchi Y, Jungbluth A, Richards EC, Old LJ . Effect of interleukin 12 on tumor induction by 3-methylcholanthrene. Proc Natl Acad Sci USA 1996; 93: 11798–11801.
Vizler C, Rosato A, Calderazzo F, Quintieri L, Fruscella P, Wainstok de Calmanovici R et al. Therapeutic effect of interleukin 12 on mouse haemangiosarcomas is not associated with an increased anti-tumour cytotoxic T-lymphocyte activity. Br J Cancer 1998; 77: 656–662.
Boggio K, Di Carlo E, Rovero S, Cavallo F, Quaglino E, Lollini PL et al. Ability of systemic interleukin-12 to hamper progressive stages of mammary carcinogenesis in HER2/neu transgenic mice. Cancer Res 2000; 60: 359–364.
Brunda MJ, Luistro L, Warrier RR, Wright RB, Hubbard BR, Murphy M et al. Antitumor and antimetastatic activity of interleukin 12 against murine tumors. J Exp Med 1993; 178: 1223–1230.
Kishima H, Shimizu K, Miyao Y, Mabuchi E, Tamura K, Tamura M et al. Systemic interleukin 12 displays anti-tumour activity in the mouse central nervous system. Br J Cancer 1998; 78: 446–453.
Shi X, Liu J, Xiang Z, Mitsuhashi M, Wu RS, Ma X . Gene expression analysis in Interleukin-12-induced suppression of mouse mammary carcinoma. Int J Cancer 2004; 110: 570–578.
Norton JA, Li M, Lee NC, Tsung K . Inhibition of host signal transducer and activator of transcription factor 6 results in cure with cyclophosphamide and interleukin 12 immunotherapy. Ann Surg Oncol 2006; 13: 118–124.
Teicher BA, Ara G, Buxton D, Leonard J, Schaub RG . Optimal scheduling of interleukin 12 and chemotherapy in the murine MB-49 bladder carcinoma and B16 melanoma. Clin Cancer Res 1997; 3: 1661–1667.
Zagozdzon R, Giermasz A, Golab J, Stoklosa T, Jalili A, Jakobisiak M . The potentiated antileukemic effects of doxorubicin and interleukin-12 combination are not dependent on nitric oxide production. Cancer lett 1999; 147: 67–75.
Weiss JM, Subleski JJ, Wigginton JM, Wiltrout RH . Immunotherapy of cancer by IL-12-based cytokine combinations. Expert Opin Biol Ther 2007; 7: 1705–1721.
Nanni P, Nicoletti G, De Giovanni C, Landuzzi L, Di Carlo E, Iezzi M et al. Prevention of HER-2/neu transgenic mammary carcinoma by tamoxifen plus interleukin 12. Int J Cancer 2003; 105: 384–389.
Nanni P, Nicoletti G, De Giovanni C, Landuzzi L, Di Carlo E, Cavallo F et al. Combined allogeneic tumor cell vaccination and systemic interleukin 12 prevents mammary carcinogenesis in HER-2/neu transgenic mice. J Exp Med 2001; 194: 1195–1205.
Zhang L, Feng D, Yu LX, Tsung K, Norton JA . Preexisting antitumor immunity augments the antitumor effects of chemotherapy. Cancer Immunol Immunother 2013; 62: 1061–1071.
Jaime-Ramirez AC, Mundy-Bosse BL, Kondadasula S, Jones NB, Roda JM, Mani A et al. IL-12 enhances the antitumor actions of trastuzumab via NK cell IFN-gamma production. J Immunol 2011; 186: 3401–3409.
Faggioli F, Soldati S, Scanziani E, Cato EM, Adorni F, Vezzoni P et al. Effects of IL-12 gene therapy on spontaneous transgenic and transplanted breast tumors. Breast Cancer Res Treat 2008; 110: 223–226.
Lucas ML, Heller L, Coppola D, Heller R . IL-12 plasmid delivery by in vivo electroporation for the successful treatment of established subcutaneous B16.F10 melanoma. Molecular therapy: the journal of the American Society of Gene Therapy 2002; 5: 668–675.
Parker JN, Gillespie GY, Love CE, Randall S, Whitley RJ, Markert JM . Engineered herpes simplex virus expressing IL-12 in the treatment of experimental murine brain tumors. Proc Natl Acad Sci USA 2000; 97: 2208–2213.
Roche FP, Sheahan BJ, O'Mara SM, Atkins GJ . Semliki Forest virus-mediated gene therapy of the RG2 rat glioma. Neuropathol Appl Neurobiol 2010; 36: 648–660.
Bramson JL, Hitt M, Addison CL, Muller WJ, Gauldie J, Graham FL . Direct intratumoral injection of an adenovirus expressing interleukin-12 induces regression and long-lasting immunity that is associated with highly localized expression of interleukin-12. Hum Gene Ther 1996; 7: 1995–2002.
Rakhmilevich AL, Janssen K, Hao Z, Sondel PM, Yang NS . Interleukin-12 gene therapy of a weakly immunogenic mouse mammary carcinoma results in reduction of spontaneous lung metastases via a T-cell-independent mechanism. Cancer Gene Ther 2000; 7: 826–838.
Shi X, Cao S, Mitsuhashi M, Xiang Z, Ma X . Genome-wide analysis of molecular changes in IL-12-induced control of mammary carcinoma via IFN-gamma-independent mechanisms. J Immunol 2004; 172: 4111–4122.
Emtage PC, Wan Y, Hitt M, Graham FL, Muller WJ, Zlotnik A et al. Adenoviral vectors expressing lymphotactin and interleukin 2 or lymphotactin and interleukin 12 synergize to facilitate tumor regression in murine breast cancer models. Hum Gene Ther 1999; 10: 697–709.
Gyorffy S, Palmer K, Podor TJ, Hitt M, Gauldie J . Combined treatment of a murine breast cancer model with type 5 adenovirus vectors expressing murine angiostatin and IL-12: a role for combined anti-angiogenesis and immunotherapy. J Immunol 2001; 166: 6212–6217.
Lohr F, Hu K, Haroon Z, Samulski TV, Huang Q, Beaty J et al. Combination treatment of murine tumors by adenovirus-mediated local B7/IL12 immunotherapy and radiotherapy. Mol Ther 2000; 2: 195–203.
Palmer K, Hitt M, Emtage PC, Gyorffy S, Gauldie J . Combined CXC chemokine and interleukin-12 gene transfer enhances antitumor immunity. Gene Ther 2001; 8: 282–290.
Putzer BM, Hitt M, Muller WJ, Emtage P, Gauldie J, Graham FL . Interleukin 12 and B7-1 costimulatory molecule expressed by an adenovirus vector act synergistically to facilitate tumor regression. Pro Natl Acad Sci USA 1997; 94: 10889–10894.
Rakhmilevich AL, Hooper AT, Hicklin DJ, Sondel PM . Treatment of experimental breast cancer using interleukin-12 gene therapy combined with anti-vascular endothelial growth factor receptor-2 antibody. Mol Cancer Ther 2004; 3: 969–976.
Chang CJ, Chen YH, Huang KW, Cheng HW, Chan SF, Tai KF et al. Combined GM-CSF and IL-12 gene therapy synergistically suppresses the growth of orthotopic liver tumors. Hepatology 2007; 45: 746–754.
Cao L, Zeng Q, Xu C, Shi S, Zhang Z, Sun X . Enhanced antitumor response mediated by the codelivery of paclitaxel and adenoviral vector expressing IL-12. Mol Pharm 2013; 10: 1804–1814.
Malvicini M, Ingolotti M, Piccioni F, Garcia M, Bayo J, Atorrasagasti C et al. Reversal of gastrointestinal carcinoma-induced immunosuppression and induction of antitumoural immunity by a combination of cyclophosphamide and gene transfer of IL-12. Mol Oncol 2011; 5: 242–255.
Passer BJ, Cheema T, Wu S, Wu CL, Rabkin SD, Martuza RL . Combination of vinblastine and oncolytic herpes simplex virus vector expressing IL-12 therapy increases antitumor and antiangiogenic effects in prostate cancer models. Cancer Gene Ther 2013; 20: 17–24.
Sin JI, Park JB, Lee IH, Park D, Choi YS, Choe J et al. Intratumoral electroporation of IL-12 cDNA eradicates established melanomas by Trp2(180-188)-specific CD8+ CTLs in a perforin/granzyme-mediated and IFN-gamma-dependent manner: application of Trp2(180-188) peptides. Cancer Immunol Immunother 2012; 61: 1671–1682.
Torrero MN, Henk WG, Li S . Regression of high-grade malignancy in mice by bleomycin and interleukin-12 electrochemogenetherapy. Clin Cancer Res 2006; 12: 257–263.
Malvicini M, Rizzo M, Alaniz L, Pinero F, Garcia M, Atorrasagasti C et al. A novel synergistic combination of cyclophosphamide and gene transfer of interleukin-12 eradicates colorectal carcinoma in mice. Clin Cancer Res 2009; 15: 7256–7265.
Egilmez NK, Jong YS, Sabel MS, Jacob JS, Mathiowitz E, Bankert RB . In situ tumor vaccination with interleukin-12-encapsulated biodegradable microspheres: induction of tumor regression and potent antitumor immunity. Cancer Res 2000; 60: 3832–3837.
Hill HC, Conway TF Jr., Sabel MS, Jong YS, Mathiowitz E, Bankert RB et al. Cancer immunotherapy with interleukin 12 and granulocyte-macrophage colony-stimulating factor-encapsulated microspheres: coinduction of innate and adaptive antitumor immunity and cure of disseminated disease. Cancer Res 2002; 62: 7254–7263.
Sabel MS, Skitzki J, Stoolman L, Egilmez NK, Mathiowitz E, Bailey N et al. Intratumoral IL-12 and TNF-alpha-loaded microspheres lead to regression of breast cancer and systemic antitumor immunity. Ann Surg Oncol 2004; 11: 147–156.
Sabel MS, Su G, Griffith KA, Chang AE . Intratumoral delivery of encapsulated IL-12, IL-18 and TNF-alpha in a model of metastatic breast cancer. Breast Cancer Res Treat 2010; 122: 325–336.
Sonabend AM, Velicu S, Ulasov IV, Han Y, Tyler B, Brem H et al. A safety and efficacy study of local delivery of interleukin-12 transgene by PPC polymer in a model of experimental glioma. Anticancer Drugs 2008; 19: 133–142.
Govaerts AS, Guillaume T, Andre M, Bayat B, Feyens AM, Hawley TS et al. Retroviral-mediated transfer of genes encoding interleukin-2 and interleukin-12 into fibroblasts increases host antitumor responsiveness. Cancer Gene Ther 1999; 6: 447–455.
Tahara H, Zeh HJ 3rd, Storkus WJ, Pappo I, Watkins SC, Gubler U et al. Fibroblasts genetically engineered to secrete interleukin 12 can suppress tumor growth and induce antitumor immunity to a murine melanoma in vivo. Cancer Res 1994; 54: 182–189.
Zitvogel L, Lotze MT . Role of interleukin-12 (IL12) as an anti-tumour agent: experimental biology and clinical application. Res Immunol 1995; 146: 628–638.
Furumoto K, Arii S, Yamasaki S, Mizumoto M, Mori A, Inoue N et al. Spleen-derived dendritic cells engineered to enhance interleukin-12 production elicit therapeutic antitumor immune responses. Int J Cancer 2000; 87: 665–672.
He Y, Liu EM, Yang XQ, Li X, Liu W . [Effects of recombinant Balillus Calmette-Guerin secreting IL-12 vaccination on development of T cell subsets in neonatal BALB/c mice]. Xi Bao Yu Fen Zi Mian Yi Xue Za Zhi [Chin J Cell Mol Immunol] 2008; 24: 634–636.
Okada N, Iiyama S, Okada Y, Mizuguchi H, Hayakawa T, Nakagawa S et al. Immunological properties and vaccine efficacy of murine dendritic cells simultaneously expressing melanoma-associated antigen and interleukin-12. Cancer Gene Ther 2005; 12: 72–83.
Seo SH, Kim KS, Park SH, Suh YS, Kim SJ, Jeun SS et al. The effects of mesenchymal stem cells injected via different routes on modified IL-12-mediated antitumor activity. Gene Ther 2011; 18: 488–495.
Kerkar SP, Goldszmid RS, Muranski P, Chinnasamy D, Yu Z, Reger RN et al. IL-12 triggers a programmatic change in dysfunctional myeloid-derived cells within mouse tumors. J Clin Inves 2011; 121: 4746–4757.
Lisiero DN, Soto H, Liau LM, Prins RM . Enhanced sensitivity to IL-2 signaling regulates the clinical responsiveness of IL-12-primed CD8(+) T cells in a melanoma model. J Immunol 2011; 186: 5068–5077.
Zhang L, Kerkar SP, Yu Z, Zheng Z, Yang S, Restifo NP et al. Improving adoptive T cell therapy by targeting and controlling IL-12 expression to the tumor environment. Mol Ther 2011; 19: 751–759.
Pegram HJ, Lee JC, Hayman EG, Imperato GH, Tedder TF, Sadelain M et al. Tumor-targeted T cells modified to secrete IL-12 eradicate systemic tumors without need for prior conditioning. Blood 2012; 119: 4133–4141.
Kerkar SP, Leonardi AJ, van Panhuys N, Zhang L, Yu Z, Crompton JG et al. Collapse of the tumor stroma is triggered by IL-12 induction of Fas. Mol Ther 2013; 21: 1369–1377.
Halin C, Rondini S, Nilsson F, Berndt A, Kosmehl H, Zardi L et al. Enhancement of the antitumor activity of interleukin-12 by targeted delivery to neovasculature. Nat Biotechnol 2002; 20: 264–269.
Sommavilla R, Pasche N, Trachsel E, Giovannoni L, Roesli C, Villa A et al. Expression, engineering and characterization of the tumor-targeting heterodimeric immunocytokine F8-IL12. Protein Eng Des Sel 2010; 23: 653–661.
Fallon J, Tighe R, Kradjian G, Guzman W, Bernhardt A, Neuteboom B et al. The immunocytokine NHS-IL12 as a potential cancer therapeutic. Oncotarget 2014; 5: 1869–1884.
Jahn T, Zuther M, Friedrichs B, Heuser C, Guhlke S, Abken H et al. An IL12-IL2-antibody fusion protein targeting Hodgkin's lymphoma cells potentiates activation of NK and T cells for an anti-tumor attack. PLoS One 2012; 7: e44482.
Atkins MB, Robertson MJ, Gordon M, Lotze MT, DeCoste M, DuBois JS et al. Phase I evaluation of intravenous recombinant human interleukin 12 in patients with advanced malignancies. Clin Cancer Res 1997; 3: 409–417.
Bajetta E, Del Vecchio M, Mortarini R, Nadeau R, Rakhit A, Rimassa L et al. Pilot study of subcutaneous recombinant human interleukin 12 in metastatic melanoma. Clin Cancer Res 1998; 4: 75–85.
Hurteau JA, Blessing JA, DeCesare SL, Creasman WT . Evaluation of recombinant human interleukin-12 in patients with recurrent or refractory ovarian cancer: a gynecologic oncology group study. Gynecol Oncol 2001; 82: 7–10.
Lacy MQ, Jacobus S, Blood EA, Kay NE, Rajkumar SV, Greipp PR . Phase II study of interleukin-12 for treatment of plateau phase multiple myeloma (E1A96): a trial of the Eastern Cooperative Oncology Group. Leuk Res 2009; 33: 1485–1489.
Leonard JP, Sherman ML, Fisher GL, Buchanan LJ, Larsen G, Atkins MB et al. Effects of single-dose interleukin-12 exposure on interleukin-12-associated toxicity and interferon-gamma production. Blood 1997; 90: 2541–2548.
Motzer RJ, Rakhit A, Thompson JA, Nemunaitis J, Murphy BA, Ellerhorst J et al. Randomized multicenter phase II trial of subcutaneous recombinant human interleukin-12 versus interferon-alpha 2a for patients with advanced renal cell carcinoma. J Interferon Cytokine Res 2001; 21: 257–263.
van Herpen CM, Looman M, Zonneveld M, Scharenborg N, de Wilde PC, van de Locht L et al. Intratumoral administration of recombinant human interleukin 12 in head and neck squamous cell carcinoma patients elicits a T-helper 1 profile in the locoregional lymph nodes. Clin Cancer Res 2004; 10: 2626–2635.
Gollob JA, Mier JW, Veenstra K, McDermott DF, Clancy D, Clancy M et al. Phase I trial of twice-weekly intravenous interleukin 12 in patients with metastatic renal cell cancer or malignant melanoma: ability to maintain IFN-gamma induction is associated with clinical response. Clin Cancer Res 2000; 6: 1678–1692.
Cebon J, Jager E, Shackleton MJ, Gibbs P, Davis ID, Hopkins W et al. Two phase I studies of low dose recombinant human IL-12 with Melan-A and influenza peptides in subjects with advanced malignant melanoma. Cancer Immun 2003; 3: 7.
Alatrash G, Hutson TE, Molto L, Richmond A, Nemec C, Mekhail T et al. Clinical and immunologic effects of subcutaneously administered interleukin-12 and interferon alfa-2b: phase I trial of patients with metastatic renal cell carcinoma or malignant melanoma. J Clinl Oncol 2004; 22: 2891–2900.
Bekaii-Saab TS, Roda JM, Guenterberg KD, Ramaswamy B, Young DC, Ferketich AK et al. A phase I trial of paclitaxel and trastuzumab in combination with interleukin-12 in patients with HER2/neu-expressing malignancies. Mol Cancer Ther 2009; 8: 2983–2991.
Rook AH, Wood GS, Yoo EK, Elenitsas R, Kao DM, Sherman ML et al. Interleukin-12 therapy of cutaneous T-cell lymphoma induces lesion regression and cytotoxic T-cell responses. Blood 1999; 94: 902–908.
Duvic M, Sherman ML, Wood GS, Kuzel TM, Olsen E, Foss F et al. A phase II open-label study of recombinant human interleukin-12 in patients with stage IA, IB, or IIA mycosis fungoides. J Am Acad Dermatol 2006; 55: 807–813.
Younes A, Pro B, Robertson MJ, Flinn IW, Romaguera JE, Hagemeister F et al. Phase II clinical trial of interleukin-12 in patients with relapsed and refractory non-Hodgkin's lymphoma and Hodgkin's disease. Clin Cancer Res 2004; 10: 5432–5438.
Ansell SM, Witzig TE, Kurtin PJ, Sloan JA, Jelinek DF, Howell KG et al. Phase 1 study of interleukin-12 in combination with rituximab in patients with B-cell non-Hodgkin lymphoma. Blood 2002; 99: 67–74.
Portielje JE, Kruit WH, Schuler M, Beck J, Lamers CH, Stoter G et al. Phase I study of subcutaneously administered recombinant human interleukin 12 in patients with advanced renal cell cancer. Clin Cancer Res 1999; 5: 3983–3989.
Daud AI, DeConti RC, Andrews S, Urbas P, Riker AI, Sondak VK et al. Phase I trial of interleukin-12 plasmid electroporation in patients with metastatic melanoma. J Clin Oncol 2008; 26: 5896–5903.
Heinzerling L, Burg G, Dummer R, Maier T, Oberholzer PA, Schultz J et al. Intratumoral injection of DNA encoding human interleukin 12 into patients with metastatic melanoma: clinical efficacy. Hum Gene Ther 2005; 16: 35–48.
Mahvi DM, Henry MB, Albertini MR, Weber S, Meredith K, Schalch H et al. Intratumoral injection of IL-12 plasmid DNA—results of a phase I/IB clinical trial. Cancer Gene Ther 2007; 14: 717–723.
Triozzi PL, Allen KO, Carlisle RR, Craig M, LoBuglio AF, Conry RM . Phase I study of the intratumoral administration of recombinant canarypox viruses expressing B7.1 and interleukin 12 in patients with metastatic melanoma. Clin Cancer Res 2005; 11: 4168–4175.
Sangro B, Mazzolini G, Ruiz J, Herraiz M, Quiroga J, Herrero I et al. Phase I trial of intratumoral injection of an adenovirus encoding interleukin-12 for advanced digestive tumors. J Clin Oncol 2004; 22: 1389–1397.
Molinier-Frenkel V, Gahery-Segard H, Mehtali M, Le Boulaire C, Ribault S, Boulanger P et al. Immune response to recombinant adenovirus in humans: capsid components from viral input are targets for vector-specific cytotoxic T lymphocytes. J Virol 2000; 74: 7678–7682.
Rudman SM, Jameson MB, McKeage MJ, Savage P, Jodrell DI, Harries M et al. A phase 1 study of AS1409, a novel antibody-cytokine fusion protein, in patients with malignant melanoma or renal cell carcinoma. Clin Cancer Res 2011; 17: 1998–2005.
Anwer K, Barnes MN, Fewell J, Lewis DH, Alvarez RD . Phase-I clinical trial of IL-12 plasmid/lipopolymer complexes for the treatment of recurrent ovarian cancer. Gene therapy 2010; 17: 360–369.
Alvarez RD, Sill MW, Davidson SA, Muller CY, Bender DP, Debernardo RL et al. A phase II trial of intraperitoneal EGEN-001, an IL-12 plasmid formulated with PEG-PEI-cholesterol lipopolymer in the treatment of persistent or recurrent epithelial ovarian, fallopian tube or primary peritoneal cancer: A Gynecologic Oncology Group study. Gynecol Oncol 2014 4 133: 433–438.
Lee S, Margolin K . Cytokines in cancer immunotherapy. Cancers 2011; 3: 3856–3893.
Schwartz RN, Stover L, Dutcher J . Managing toxicities of high-dose interleukin-2. Oncology 2002; 16 (11 Suppl 13): 11–20.
Sabel MS, Sondak VK . Pros and cons of adjuvant interferon in the treatment of melanoma. Oncologist 2003; 8: 451–458.
Meyaard L, Hovenkamp E, Otto SA, Miedema F . IL-12-induced IL-10 production by human T cells as a negative feedback for IL-12-induced immune responses. J Immunol 1996; 156: 2776–2782.
Portielje JE, Lamers CH, Kruit WH, Sparreboom A, Bolhuis RL, Stoter G et al. Repeated administrations of interleukin (IL)-12 are associated with persistently elevated plasma levels of IL-10 and declining IFN-gamma, tumor necrosis factor-alpha, IL-6, and IL-8 responses. Clin Cancer Res 2003; 9: 76–83.
Vignali DA, Kuchroo VK . IL-12 family cytokines: immunological playmakers. Nat Immunol 2012 Aug; 13: 722–728.
Langowski JL, Zhang X, Wu L, Mattson JD, Chen T, Smith K et al. IL-23 promotes tumour incidence and growth. Nature 2006; 442: 461–465.
Teng MW, von Scheidt B, Duret H, Towne JE, Smyth MJ . Anti-IL-23 monoclonal antibody synergizes in combination with targeted therapies or IL-2 to suppress tumor growth and metastases. Cancer Res 2011; 71: 2077–2086.
Grivennikov SI, Wang K, Mucida D, Stewart CA, Schnabl B, Jauch D et al. Adenoma-linked barrier defects and microbial products drive IL-23/IL-17-mediated tumour growth. Nature 2012; 491: 254–258.
Teng MW, Andrews DM, McLaughlin N, von Scheidt B, Ngiow SF, Moller A et al. IL-23 suppresses innate immune response independently of IL-17A during carcinogenesis and metastasis. Pro Natl Acad Sci USA 2010; 107: 8328–8333.
Hu J, Yuan X, Belladonna ML, Ong JM, Wachsmann-Hogiu S, Farkas DL et al. Induction of potent antitumor immunity by intratumoral injection of interleukin 23-transduced dendritic cells. Cancer Res 2006; 66: 8887–8896.
Kaiga T, Sato M, Kaneda H, Iwakura Y, Takayama T, Tahara H . Systemic administration of IL-23 induces potent antitumor immunity primarily mediated through Th1-type response in association with the endogenously expressed IL-12. J Immunol 2007; 178: 7571–7580.
Reay J, Gambotto A, Robbins PD . The antitumor effects of adenoviral-mediated, intratumoral delivery of interleukin 23 require endogenous IL-12. Cancer Gene Ther 2012; 19: 135–143.
Murugaiyan G, Saha B . IL-27 in tumor immunity and immunotherapy. Trends Mol Med 2013; 19: 108–116.
Cocco C, Giuliani N, Di Carlo E, Ognio E, Storti P, Abeltino M et al. Interleukin-27 acts as multifunctional antitumor agent in multiple myeloma. Clin Cancer Res 2010; 16: 4188–4197.
Cocco C, Pistoia V, Airoldi I . Anti-leukemic properties of IL-12, IL-23 and IL-27: differences and similarities in the control of pediatric B acute lymphoblastic leukemia. Crit Rev Oncol Hematol 2012; 83: 310–318.
Shimizu M, Shimamura M, Owaki T, Asakawa M, Fujita K, Kudo M et al. Antiangiogenic and antitumor activities of IL-27. J Immunol 2006; 176: 7317–7324.
Yoshimoto T, Morishima N, Mizoguchi I, Shimizu M, Nagai H, Oniki S et al. Antiproliferative activity of IL-27 on melanoma. J Immunol 2008; 180: 6527–6535.
Zorzoli A, Di Carlo E, Cocco C, Ognio E, Ribatti D, Ferretti E et al. Interleukin-27 inhibits the growth of pediatric acute myeloid leukemia in NOD/SCID/Il2rg-/- mice. Clin Cancer Res 2012; 18: 1630–1640.
Oniki S, Nagai H, Horikawa T, Furukawa J, Belladonna ML, Yoshimoto T et al. Interleukin-23 and interleukin-27 exert quite different antitumor and vaccine effects on poorly immunogenic melanoma. Cancer Res 2006; 66: 6395–6404.
Salcedo R, Stauffer JK, Lincoln E, Back TC, Hixon JA, Hahn C et al. IL-27 mediates complete regression of orthotopic primary and metastatic murine neuroblastoma tumors: role for CD8+ T cells. J Immunol 2004; 173: 7170–7182.
Pflanz S, Timans JC, Cheung J, Rosales R, Kanzler H, Gilbert J et al. IL-27, a heterodimeric cytokine composed of EBI3 and p28 protein, induces proliferation of naive CD4+ T cells. Immunity 2002; 16: 779–790.
Collison LW, Workman CJ, Kuo TT, Boyd K, Wang Y, Vignali KM et al. The inhibitory cytokine IL-35 contributes to regulatory T-cell function. Nature 2007; 450: 566–569.
Wang Z, Liu JQ, Liu Z, Shen R, Zhang G, Xu J et al. Tumor-derived IL-35 promotes tumor growth by enhancing myeloid cell accumulation and angiogenesis. J Immunol 2013; 190: 2415–2423.
Edited by H-U Simon
About this article
Cite this article
Tugues, S., Burkhard, S., Ohs, I. et al. New insights into IL-12-mediated tumor suppression. Cell Death Differ 22, 237–246 (2015). https://doi.org/10.1038/cdd.2014.134
Masking the immunotoxicity of interleukin-12 by fusing it with a domain of its receptor via a tumour-protease-cleavable linker
Nature Biomedical Engineering (2022)
Cancer Immunology, Immunotherapy (2022)
Cellular Oncology (2022)
Breast Cancer Research and Treatment (2022)
Drug Delivery and Translational Research (2022)