Tumor regression induced by intratumor administration of adenovirus vector expressing CD40 ligand and naive dendritic cells

We have previously shown that in vivo genetic modification of tumors with an adenovirus (Ad) vector engineered to express CD40 ligand (AdmCD40L) induces tumor-specific CTLs and suppresses tumor growth in vivo. In the present study, we investigate the hypothesis that this treatment can be made more efficient with 10(2)-fold less Ad vector by also administering bone marrow-generated dendritic cells (DCs) to the tumor. Using AdmCD40L and the number of DCs that alone had no effect on tumor growth, the data show that the growth of CT26 (colon adenocarcinoma; H-2d) and B16 (melanoma; H-2b) murine s.c. tumors is significantly suppressed by direct administration of DCs into s.c. established tumors that had been pretreated with AdmCD40L 2 days previously. The antitumor effect produced by the combination therapy AdmCD40L + DCs correlated with in vivo priming of tumor-specific CTLs. The antitumor cell-mediated immunity was transferable to naive mice by spleen cells from AdmCD40L + DC-treated animals. The interactions between CD40L and CD40 within treated tumors were critical because tumor suppression was abrogated by coadministration to the tumors of neutralizing monoclonal antibody against CD40L along with the DCs. Finally, in vivo depletion and knockout mice experiments demonstrated that tumor regression produced by this combination therapy depends on CD8+ T cells, but not on CD4+ T cells. These findings should be useful in designing strategies for use of DCs and AdmCD40L in cancer immunotherapy.     

Antigen-specific cancer immunotherapy using a GM-CSF secreting allogeneic tumor cell-based vaccine

Granulocyte-macrophage colony-stimulating factor (GM-CSF)-transduced autologous tumor cell-based vaccines are currently one of the major forms of cancer vaccines. However, the preparation of GM-CSF-transduced autologous tumor vaccines is time-consuming and technically challenging. In addition, the host antigen presenting cells, rather than the tumor vaccine cells themselves, present tumor-specific antigens and prime the host T cells. Therefore, we tested the efficacy of antigen-specific allogeneic tumor vaccines. We used human papillomavirus 16 (HPV-16) E7 protein as a model tumor antigen, which is associated with the development of most cervical carcinoma. B16, a C57BL/6 (H-2(b)) derived melanoma cell line, was genetically engineered to produce GM-CSF alone (B16GM), HPV-16 E7 alone (B16E7), or both (B16GME7). These vaccine cells were injected into BALB/c (H-2(d)) mice (10(6) cells/mouse). Two weeks later, mice were challenged with 10(5) live HPV-16 E7(+) BL-1 (H-2(d)) tumor cells and monitored for tumor progression twice weekly. To determine the effective cell population in the antitumor immunity elicited by B16GME7, we carried out in vivo antibody depletion experiments using CD4 and CD8 specific antibodies. In addition, as a measure of the immune responses produced by B16GME7, we performed an in vitro cytotoxic T lymphocyte assay using a standard chromium release method. We found that all of the mice vaccinated with B16GME7 remained tumor free 49 days post-BL-1 challenge. In contrast, mice vaccinated with B16GM and B16E7 did not show any tumor protection against a similar dose of BL-1 cells. Furthermore, the antitumor immunity produced by B16GME7 was dependent on both CD4 and CD8 T cells. In addition, E7-specific cytotoxic T lymphocyte activity could be readily demonstrated in mice immunized with B16GME7. These results suggest that allogeneic tumor cells transduced with GM-CSF and the tumor antigen, HPV-16 E7, cannot only generate an E7-specific cytotoxic T lymphocytes response in vitro, but can also elicit a potent antitumor immune response against an E7 expressing tumor in vivo.     

Coexpression of CD40L and CD70 by semiallogenic tumor cells induces anti-tumor immunity

The immune system is potentially qualified to detect and eliminate tumor cells, but various mechanisms developed by tumor cells allow tumor escape. Strategies selected to promote antitumor responses have included genetic modifications of tumor cells to induce expression of costimulatory molecules. Moreover, alloantigens can also act as strong enhancers of the immune response. In this work, we have associated the expression of two costimulatory members of the TNF superfamily, CD40L and CD70 along with an allogenic MHC Class I (H-2K(d)) molecule expression on melanoma cells (B16F10, H-2(b)) to favor the antitumor immune response. B16F10 tumor growth slows significantly when CD40L and CD70 are coexpressed by tumor cells and the association with the allogenic molecule (H-2K(d)) enhances this effect. Growth kinetics of mock and CD40L-CD70-H-2K(d)-expressing B16F10 tumors in immunocompetent versus nu/nu and beige mice suggested that CD8(+) T lymphocytes and NK cells were involved in this antitumor immunity. A delay in mock tumor growth was observed when CD40L-CD70-H-2K(d)-expressing B16F10 cells and mock tumor cells were injected simultaneously and contralaterally. It was also shown that in vivo immunization of immunocompetent mice with CD40L-CD70-H-2K(d) B16F10 tumor cells improved the generation of cytotoxic lymphocytes against the wild-type melanoma cells expressing the syngenic MHC Class I molecule H-2K(b) (B16K1). These observations lay a path for new immunotherapeutic trials using semiallogenic fibroblasts expressing costimulatory molecules and tumor-associated antigens.     

Intratumoral administration of immature dendritic cells following the adenovirus vector encoding CD40 ligand elicits significant regression of established myeloma

Our previous study showed that J558 myeloma cells engineered CD40L lost their tumorigenicity in syngeneic mice, and the inoculation of J558/CD40L tumor cells further led to the protective immunity against wild tumors. In the present study, we investigated whether the vaccine can exert more efficient antitumor immunity by combination with adenovirus mediated CD40L gene therapy and immature dendritic cells (iDCs). The results demonstrated that intratumoral administration of iDCs 2 days after AdVCD40L injection, not only significantly suppressed the tumor growth, but also eradiated the established tumors in 40% of the mice. The potent antitumor effect produced by the combination therapy correlated with high expression of MHC, costimulatory and Fas molecules on J558 cells, which was derived from CD40L transgene expression. In addition, transgene CD40L expression could dramatically induce J558 cell apoptosis. Effectively capturing apoptotic bodies by iDCs in vivo could induce DC maturation, prime tumor-specific CTLs and tend to Th1-type immune response. Finally, in vivo depletion experimentation suggested both CD4+ and CD8+ T cells were involved in mediating the antitumor immune responses of combined treatment of AdVCD40L and iDCs, with CD8+ T cells being the major effector. These findings could be beneficial for designing strategies of DCs vaccine and CD40L for anticancer immunotherapy.     

IL-10 elicits IFNγ-dependent tumor immune surveillance

Tumor immune surveillance and cancer immunotherapies are thought to depend on the intratumoral infiltration of activated CD8(+) T?cells. Intratumoral CD8(+) T?cells are rare and lack activity. IL-10 is thought to contribute to the underlying immune suppressive microenvironment. Defying those expectations we demonstrate that IL-10 induces several essential mechanisms for effective antitumor immune surveillance: infiltration and activation of intratumoral tumor-specific cytotoxic CD8(+) T?cells, expression of the Th1 cytokine interferon-γ (IFNγ) and granzymes in CD8(+) T?cells, and intratumoral antigen presentation molecules. Consequently, tumor immune surveillance is weakened in mice deficient for IL-10 whereas transgenic overexpression of IL-10 protects mice from carcinogenesis. Treatment with pegylated IL-10 restores tumor-specific intratumoral CD8(+) T?cell function and controls tumor growth.     

Antitumor immunity against bladder cancer induced by ex vivo expression of CD40 ligand gene using retrovirus vector

The interaction between CD40 ligand (CD40L) and CD40 on antigen-presenting cells is essential for the initiation of antigen-specific T-cell responses. In order to clarify whether the expression of CD40L in tumor cells might be useful as a systemic therapy against bladder cancer, we investigated the antitumor immunity induced by CD40L in the mouse bladder cancer cell line MBT2. MBT2 was transduced by the retroviral vector expressing CD40L (MBT2-CD40L). Mouse bone marrow-derived dendritic cells cocultured with MBT2-CD40L cells produced eight times more IL-12 than those cocultured with parental MBT2 cells. In animal studies, subcutaneously inoculated MBT2-CD40L cells were rejected promptly. The vaccination of MBT2-CD40L cells induced antitumor immunity against parental tumors at a distant site. However, the antitumor effect of MBT2-CD40L inoculation was insufficient against pre-existing tumors. In the vaccination model, antibody ablation studies revealed that CD4(+) T cells were required for antitumor immunity, and tumor-specific cytotoxicity of sera was demonstrated. These data demonstrated that the antitumor immunity induced by CD40L was effective in the vaccination model and suggested that immunogene therapy using CD40L may be a new strategy of systemic therapy against bladder cancer.     

Signaling through OX40 enhances antitumor immunity

The existence of tumor-specific T cells, as well as their ability to be primed in cancer patients, confirms that the immune response can be deployed to combat cancer. However, there are obstacles that must be overcome to convert the ineffective immune response commonly found in the tumor environment to one that leads to sustained destruction of tumor. Members of the tumor necrosis factor (TNF) superfamily direct diverse immune functions. OX40 and its ligand, OX40L, are key TNF members that augment T-cell expansion, cytokine production, and survival. OX40 signaling also controls regulatory T-cell differentiation and suppressive function. Studies over the past decade have demonstrated that OX40 agonists enhance antitumor immunity in preclinical models using immunogenic tumors; however, treatment of poorly immunogenic tumors has been less successful. Combining strategies that prime tumor-specific T cells together with OX40 signaling could generate and maintain a therapeutic antitumor immune response.     

重组鼠CD40配体的Ankara痘病毒疫苗对鼠淋巴瘤的免疫治疗作用的研究

目的研究Ankara痘病毒和mCD40L的重组体TBC-MVA-mCD40L对小鼠淋巴瘤的免疫治疗作用。方法 应用流式细胞术检测经体外或者体内转染mCD40L的A20瘤细胞的表面共刺激分子CD80和CD86的表达。构建了BALB/c小鼠的B细胞淋巴瘤模型并分组接受瘤内注射TBC-MVA-mCD40L联合系统化疗的治疗模式。另外, 应用与A20细胞孵育后的TBC-MVA-mCD40L对鼠进行免疫, 观察其对鼠淋巴瘤的预防作用。通过观察小鼠接种的肿瘤的大小和生长情况来判断治疗效果。结果 转染CD40L后流式检测A20细胞表面共刺激分子CD80和CD86的表达增强。TBC-MVA-mCD40L免疫治疗联合化疗能成功地使肿瘤消退并使荷瘤鼠获得长期生存。TBC-MVA-mCD40L免疫预防接种能使小鼠获得长期的抗肿瘤免疫。结论 对B淋巴瘤细胞的CD40L基因修饰策略能有效增强机体的对抗B细胞淋巴瘤的系统免疫, 为将来B细胞淋巴瘤患者肿瘤内注射TBC-MVA-mCD40的免疫治疗和预防方法提供依据, 为该免疫治疗方法的临床应用奠定基础。     

Systemic antitumor effect of intratumoral injection of dendritic cells in combination with local photodynamic therapy.

PURPOSE: Photodynamic therapy (PDT), which is used clinically for the palliative treatment of cancer, induces local tumor cell death but has no effect on tumors in untreated sites. The purpose of this study was to determine if local PDT followed by intratumoral injection of na?ve dendritic cells (IT-DC) induces systemic antitumor immunity that can inhibit the growth of untreated as well as PDT + IT-DC-treated tumors. EXPERIMENTAL DESIGN: BALB/c or C57Bl/6 mice were injected s.c. with CT26 colorectal carcinoma cells and B16 melanoma cells, respectively, and following 10 to 12 days of tumor growth, the tumors were treated with PDT alone or PDT followed by IT-DC or IT-PBS. In other studies, tumors were established simultaneously in both lower flanks or in one flank and in the lungs, but only one flank was treated. RESULTS: Whereas neither PDT nor IT-DC alone was effective, PDT + IT-DC eradicated both CT26 and B16 tumors in a significant proportion of animals and prolonged the survival of mice of which the tumors were not cured. The spleens of mice treated with PDT + IT-DC contained tumor-specific cytotoxic and IFN-gamma-secreting T cells whereas the spleens of control groups did not. Moreover, adoptive transfer of splenocytes from successfully treated CT26 tumor-free mice protected na?ve animals from a subsequent challenge with CT26, and this was mediated mainly by CD8 T cells. Most importantly, PDT plus IT-DC administered to one tumor site led to tumor regression at distant sites, including multiple lung metastases. CONCLUSIONS: PDT + IT-DC induces potent systemic antitumor immunity in mice and should be evaluated in the treatment of human cancer.     

Induction of antitumor immunity by transduction of CD40 ligand gene and interferon-gamma gene into lung cancer

CD40-CD40 ligand (CD40L) interaction is an important costimulatory signaling pathway in the crosstalk between T cells and antigen-presenting cells. This receptor-ligand system is known to be essential in eliciting strong cellular immunity. Here we demonstrate that murine lung cancer cells (3LLSA) transduced with the CD40L gene (3LLSA-CD40L) were rejected in syngeneic C57BL/6 mice, but grew in CD40-deficient mice to the same extent as control tumor cells. Immunohistochemical study showed that inflammatory cells, including CD4+, CD8+ T cells and NK cells, infiltrated into the inoculated 3LLSA-CD40L tumor tissue. Inoculation of 3LLSA-CD40L cells into mice resulted in the induction of 3LLSA-specific cytotoxic T-cell immunity, and the growth of parental 3LLSA tumors was inhibited when 3LLSA cells were inoculated into C57BL/6 mice mixed with 3LLSA-CD40L cells or when they were rechallenged 4 weeks after 3LLSA-CD40L cells were rejected. Furthermore, co-inoculation of interferon (IFN)-gamma-transduced cells (3LLSA-IFNgamma) with 3LLSA-CD40L cells enhanced the antitumor immunity efficiently in vivo. These results indicate that the in vivo priming with CD40L- and IFN-gamma gene-transduced lung cancer cells is a promising strategy for inducing antitumor immunity in the treatment of lung cancer.     

Efficient antitumor immunity derived from maturation of dendritic cells that had phagocytosed apoptotic/necrotic tumor cells

Dendritic cells (DCs) that acquired antigen from apoptotic tumor cells are able to induce major histocompatibility complex (MHC) class I-restricted cytotoxic T lymphocytes and antitumor immunity. In the present study, we investigated the efficiency of antitumor immunity derived from DCs that had phagocytosed apoptotic/necrotic BL6-10 melanoma cells compared with that of DCs pulsed with the tumor mTRP2 peptide. Our data showed that phagocytosis of apoptotic/necrotic tumor cells resulted in maturation of DCs with up-regulated expression of proinflammatory cytokines [interleukin (IL)-1beta, IL-6, tumor necrosis factor-alpha, interferon-gamma and granulocyte-macrophage colony-stimulating factor], chemokines (MIP-1alpha, MIP-1beta and MIP-2), the CC chemokine receptor CCR7 and the cell surface molecules (MHC class II, CD11b, CD40 and CD86), and down-regulated expression of the CC chemokine receptors CCR2 and CCR5. These mature DCs displayed enhanced migration toward the CC chemokine MIP-3beta in a chemotaxis assay in vitro and to the regional lymph nodes in an animal model in vivo. Our data also showed that vaccination with DCs that had phagocytosed apoptotic/necrotic BL6-10 cells was able to (i) more strongly stimulate allogeneic T-cell proliferation in vitro, (ii) induce an in vivo Th1-type immune response leading to more efficient tumor-specific cytotoxic CD8(+) T-cell-mediated immunity and (iii) eradicate lung metastases in all 6 vaccinated mice compared with mice vaccinated with DCs pulsed with the tumor mTRP2 peptide, in which lung metastases were reduced (mean number of 16 per mouse) but not completely eradicated. Therefore, DCs that had phagocytosed apoptotic/necrotic tumor cells appear to offer new strategies in DC cancer vaccines.     

Antitumor activity of alpha-galactosylceramide, KRN7000, in mice with the melanoma B16 hepatic metastasis and immunohistological study of tumor infiltrating cells

Liver metastasis of primary tumors is clinically a major problem. We examined the antitumor activity of KRN7000, an alpha-galactosylceramide, in mice with liver metastasis of the B16 melanoma. KRN7000 significantly inhibited tumor growth in the liver, and its potency was similar to that of interleukin-12. The KRN7000 administration resulted in a high percentage of cured mice, which acquired tumor-specific immunity. To study what kinds of antitumor effector cells participated in killing tumor cells, we then performed immunohistological analysis of tumor-infiltrating cells, and found that KRN7000 induced marked invasion of NK1.1+ cells, CD8+ cells, and F4/80+ cells (macrophages) into B16 tumor nodules. In addition, it appeared that KRN7000-treated, liver-associated macrophages possessed strong lytic activity against tumor cells. These results suggest that NK cells, NK1.1+ T (NKT) cells, cytotoxic T lymphocytes, and macrophages play an important role in killing tumor cells in the liver, and that KRN7000 may be useful for the treatment of cancer liver metastasis.     

Therapeutic vaccination against murine lymphoma by intratumoral injection of recombinant fowlpox virus encoding CD40 ligand

The interaction between CD40 ligand (CD40L, CD154) and its receptor CD40 on antigen-presenting cells is essential for the initiation of cell-mediated and humoral immune responses. Malignant B cells also express CD40 and respond to CD40L by enhancing expression of costimulatory molecules. In this study, we investigated the therapeutic antitumor effect of intratumoral administration of recombinant fowlpox virus encoding murine CD40L (rF-mCD40L) in a murine B-cell lymphoma model. BALB/c mice with established s.c. and widely metastatic A20 lymphoma tumors were treated with intratumoral injections of rF-mCD40L together with systemic chemotherapy. This combined chemoimmunotherapy resulted in complete tumor regression and long-term survival of the mice. Some tumor cells in the injected sites expressed the CD40L transgene and had increased expression of the CD80 and CD86 costimulatory molecules. The therapeutic effect was dependent on CD8 but not on CD4 T cells. Moreover, there was a requirement that the recombinant CD40L virus be injected directly into the tumor, as opposed to peritumoral or distant sites. Thus, rF-mCD40L injected directly into the tumor microenvironment enhances the immunogenicity of tumor B cells. The results support future plans for intratumoral injection of rF-mCD40L in patients with lymphoma.     

腺病毒介导的胞嘧啶脱氨酶自杀基因联合IL—2基因疗法的肿瘤治 …

以腺病毒作为载体,将大肠杆菌胞嘧啶脱氨酶基因与小鼠ＩＬ－２基因联合转移,研究其体内抗肿瘤作用及免疫机理。方法小鼠皮下接种黑色素瘤Ｂ１６Ｆ１０细胞后３天,肿瘤局部 注射表达ＩＬ－２的重要腺病毒ＡｄＩＬ－２和表达的ＣＤ的重组腺病毒ＡｄＣＤ,然后连续０天给予５－氟胞嘧啶３００ｍｇ／ｋｇ进行治疗。     

Treatment of hepatocellular carcinoma with adenoviral vector-mediated Flt3 ligand gene therapy

Fms-like tyrosine kinase 3 ligand (Flt3L) plays an important role in development and activation of dendritic cells (DCs) and natural killer cells (NK). It has been shown that administration of either tumor cells transfected in vitro with Flt3L vectors or soluble Flt3L fusion protein in a high dose can enhance host antitumor immunity in animal model systems. In this study, we developed a recombinant defective adenovirus with an insert of gene encoding extracellular domain of mouse Flt3L (Ad-mFlt3L) under control of cytomegalovirus promoter and investigated its biological efficacy in eliciting tumor-specific immune response against hepatocellular carcinoma in mouse hepatoma model. The constructed Ad-mFlt3L efficiently infected hepa 1-6 hepatoma cells both in vitro and in vivo, leading to a high production of mFlt3L proteins in association with accumulation of DCsNK cells and lymphocytes in local tumor tissues. Tumor cells infected with Ad-mFlt3L lost tumorigenicity and became more immunogenic in syngeniec animal models. Intratumoral injection of Ad-mFlt3L (10(9) expression-forming unit) x 3 significantly inhibited tumor growth with elicitation of long-lasting antitumor immunity, which is both preventive and curative. The tumor-specific immunity can be partially abrogated by depletion of either CD3+CD4+ T cells or NK cells and can be also re-established in na?ve animals by adoptive transfer of splenocytes from treated mice. The results suggest that adenovirus-mediated Flt3L gene therapy may provide a useful strategy for treatment of cancers.     

Adenovirus-mediated combined P16 gene and GM-CSF gene therapy for the treatment of established tumor and induction of antitumor immunity

Antitumor effects of combined transfer of P16 and cytokine genes were investigated in this study. The adenovirus harboring the P16 gene (AdP16) and murine granulocyte-macrophage colony-stimulating factor gene (AdGM-CSF) were utilized for the treatment of established tumors. The mice were inoculated subcutaneously with Renca cells and, 6 days later, received an intratumoral injection of AdP16 in the presence or absence of AdGM-CSF. The results demonstrated that tumor-bearing mice treated with AdP16 in combination with AdGM-CSF showed more potent inhibition of tumor growth and survived much longer than did mice treated with AdP16, AdGM-CSF, adenovirus expressing beta-galactosidase, or phosphate-buffered saline alone (P<.01). The tumor mass showed obvious necrosis and inflammatory cell infiltration, and more CD(4)(+) and CD(8)(+) T cells infiltrating the tumor after combined therapy. After combined therapy, the expression of MHC-1 (H-2K(d)) and Fas molecules on freshly isolated tumor cells increased greatly. The activity of specific cytotoxic T lymphocytes was also found to be induced more significantly after the combined therapy (P<.01). Our results demonstrated that combined therapy with P16 and GM-CSF genes can inhibit the growth of established tumors in mice significantly and induce antitumor immunity of the host efficiently.     

Glucocorticoid-induced tumor necrosis factor receptor stimulation enhances the multifunctionality of adoptively transferred tumor antigen-specific CD8+ T cells with tumor regression

We have reported for the first time the significance of effector T-cell multifunctionality in antitumor immunity, suggesting that the appearance of multifunctional/polyfunctional tumor-specific CD8⁺ T cells in vivo is a critical determinant of the success of antitumor immunotherapy, and a strategy to induce multifunctionality in effector cells is required for the successful immunotherapy of hosts with progressing tumor. Glucocorticoid-induced tumor necrosis factor receptor (GITR) stimulation has been shown to enhance antitumor immune response. However, its functional impact on adoptively transferred T cells remains unclear. Here, we analyzed the impact of GITR stimulation in vivo on the functional profiles of adoptively transferred CD8⁺ T cells specific for murine fibrosarcoma CMS5. GITR stimulation was found to enhance multifunctionality (interferon (IFN)-γ and tumor necrosis factor (TNF)-α production and CD107a mobilization as a degranulation marker) in transferred cells at the single-cell level. These cells exhibited upregulated expression of CD25 in draining lymph nodes and increased infiltration in tumor. Mice that received T-cell therapy with GITR stimulation showed reduced Foxp3⁺CD4⁺ T cells among tumor infiltrating lymphocytes and increased in vivo cytotoxic T lymphocytes (CTL) activity even with progressing tumor, resulting in enhanced tumor regression. These data strengthen the idea that effector T-cell multifunctionality is a sensitive immune correlate for successful immunotherapy against malignancy and provide an immunological rationale for effective T-cell therapy combined with GITR stimulation. ( Cancer Sci 2009; 100: 1317–1325)     

Immunomodulatory gene therapy with interleukin 12 and 4-1BB ligand: long- term remission of liver metastases in a mouse model

BACKGROUND: The success of immunomodulatory cancer therapy is frequently hampered by the transient nature of the antitumor immune response. We have shown previously in a mouse model that interleukin 12 (IL-12) generates a strong natural killer (NK) cell-mediated antitumor response and reduces liver metastases induced by a colon carcinoma cell line. However, only a small percentage of the treated animals developed the cytotoxic T-lymphocytic response required for a long-term systemic antitumor immunity. 4-1BB is a co-stimulatory molecule expressed on the surface of activated T cells. Interaction of 4-1BB with its natural ligand (4-1BBL) has been shown to amplify T-cell (especially CD8+)-mediated immunity. In this study, we investigated the effects of adenovirus-mediated gene therapy delivering both IL-12 and 4-1BBL genes on mice with hepatic metastases induced by colon cancer cells. METHODS: Syngeneic BALB/c mice received intrahepatic injection of poorly immunogenic MCA26 colon cancer cells. Various combinations of replication-defective adenoviruses expressing IL-12 and 4-1BBL genes were injected into the established liver tumors. Changes in tumor size and animal survival were then monitored. All statistical tests were two-sided. RESULTS: The long-term survival rate of mice treated with the combination of IL-12 and 4-1BBL was significantly improved over that of animals in the control group (P =.0001). In vivo depletion of NK cells or CD8+ T cells completely abolished the long-term survival advantage of the IL-12 plus 4-1BBL-treated animals (P<.002). Moreover, the systemic immunity induced by this combination treatment protected these animals against a subcutaneous challenge with parental MCA26 cells. CONCLUSION: Adenovirus-mediated transfer of IL-12 and 4-1BBL genes directly into liver tumors resulted in tumor regression that required both NK and CD8+ T cells and generated a potent, long-lasting antitumor immunity.     

Induction of potent antitumor immunity by intratumoral injection of interleukin 23-transduced dendritic cells

Dendritic cells (DCs) are potent antigen-presenting cells that play a critical role in priming immune responses to tumor. Interleukin (IL)-23 can act directly on DC to promote immunogenic presentation of tumor peptide in vitro. Here, we evaluated the combination of bone marrow-derived DC and IL-23 on the induction of antitumor immunity in a mouse intracranial glioma model. DCs can be transduced by an adenoviral vector coding single-chain mouse IL-23 to express high levels of bioactive IL-23. Intratumoral implantation of IL-23-expressing DCs produced a protective effect on intracranial tumor-bearing mice. The mice consequently gained systemic immunity against the same tumor rechallenge. The protective effect of IL-23-expressing DCs was comparable with or even better than that of IL-12-expressing DCs. IL-23-transduced DC (DC-IL-23) treatment resulted in robust intratumoral CD8(+) and CD4(+) T-cell infiltration and induced a specific TH1-type response to the tumor in regional lymph nodes and spleen at levels greater than those of nontransduced DCs. Moreover, splenocytes from animals treated with DC-IL-23 showed heightened levels of specific CTL activity. In vivo lymphocyte depletion experiments showed that the antitumor immunity induced by DC-IL-23 was mainly dependent on CD8(+) T cells and that CD4(+) T cells and natural killer cells were also involved. In summary, i.t. injection of DC-IL-23 resulted in significant and effective systemic antitumor immunity in intracranial tumor-bearing mice. These findings suggest a new approach to induce potent tumor-specific immunity to intracranial tumors. This approach may have therapeutic potential for treating human glioma.     

Induction of therapeutic T-cell immunity by tumor targeting with soluble recombinant B7-immunoglobulin costimulatory molecules.

Tumor targeting with immunomodulatory molecules is an attractive strategy to enhance the host's antitumor response. Expression of CD80 (B7-1) and CD86 (B7-2) costimulatory molecules in tumor cells has proven to be an efficient way to enhance their immunogenicity. Here, we studied the effects of tumor targeting with biotinylated recombinant soluble B7-1- and B7-2 immunoglobulin G molecules (bio-B7-IgG) using a pretargeting approach based on the sequential use of a biotinylated antitumor monoclonal antibody and avidin. Mouse RMA T-lymphoma cells bearing either bio-B7-1-IgG or bio-B7-2-IgG on their surface prime in vitro naive CD8+ CTLs, which are highly effective in adoptive immunotherapy, and induce therapeutic immunity when injected in tumor-bearing animals. In vivo targeting of established RMA tumors with bio-B7-IgG either cures tumor-bearing mice or significantly prolongs their survival. The antitumor response induced by targeted bio-B7-IgG depends on both CD4+ and CD8+ T cells. Moreover, tumor targeting with bio-B7-IgG in vivo is critical for both expansion in lymphoid organs and mobilization into the tumor of tumor-specific CD8+ CTLs. When targeting is performed on poorly immunogenic TS/A mammary adenocarcinoma, only bio-B7-1-IgG primes naive CTLs in vitro and cures or significantly prolongs the survival of tumor-bearing mice in vivo, confirming that the two costimulatory molecules are not redundant with this tumor. Altogether, these data suggest that tumor avidination and targeting with soluble bio-B7-IgG may represent a promising strategy to enhance the antitumor response in the host.