Intratumoral expression of macrophage-derived chemokine induces CD4+ T cell-independent antitumor immunity in mice

Macrophage-derived chemokine is chemotactic for a variety of leukocytes, and has been shown to be involved in T 2-mediated cellular immunity. To evaluate the role of this chemokine in tumor immunity in vivo, an adenovirus vector encoding the human macrophage-derived chemokine cDNA (AdMDC) was administered to established murine tumors. Gene transfer with AdMDC significantly inhibited tumor growth and prolonged animal survival. AdMDC was not directly cytotoxic to tumor cells, but splenocytes from animals that received intratumoral AdMDC were able to lyse syngeneic tumor cells, and purified splenic CD8 cells secreted interferon-gamma in a tumor-specific manner. The antitumor activity of AdMDC was lost in mice lacking CD8 T lymphocytes, but surprisingly, it was preserved in animals lacking CD4 cells, as was the systemic cytotoxic T lymphocyte response. Systemic NK cells did not play a role in the antitumor immune response induced by AdMDC. Experiments using knockout mice demonstrated that host expression of MHC Class I, but not Class II, IL-4, or IL-12, was necessary for AdMDC to exert its antitumor effect, and immunohistochemistry demonstrated infiltrates of CD8 and CD86 cells, but not CD4 cells in treated tumors. These studies highlight a new function for macrophage-derived chemokine by demonstrating that it possesses in vivo antitumor activity with CD8 T cells as the effector cells, and interestingly, that the CD4 cell/MHC II pathway of CD8 cell activation is not required for the antitumor effects of this chemokine.(H)     

Expression of the chemokines IP-10 and Mig in IL-10 transduced tumors

Several laboratories have reported marked tumor inhibition when the cytokine interleukin-10 (IL-10) is overexpressed as a transgene in a variety of tumor cells. To identify critical effector molecules, we compared the expression of the chemokine crg-2, the murine homolog of human inducible protein 10 (human IP-10) in murine mammary tumors derived from the transplantation of six IL-10 expressing clones of tumor cell line 66.1, parental 66.1, or 66-neo-cells. We observed increased levels of IP-10 mRNA in all IL-10-expressing tumors examined in comparison to 66-neo. IP-10 mRNA was not detected in parental 66.1 tumors. The closely related chemokine Mig (monokine induced by interferon-gamma [IFN-gamma]) was also induced in all IL-10-expressing tumors. Studies of cultured tumor cells in vitro show that mammary epithelial tumor cells, in the absence of host elements, can express IP-10 and Mig in response to induction with either lipopolysaccharide (LPS) or IFN-gamma alone. The combination of LPS plus IFN-gamma resulted in even greater induction of IP-10 RNA. The kinetics of induction differ somewhat for the two chemokines, with IP-10 showing slower induction and less rapid decline. Because both Mig and IP-10 are chemotactic for tumor-infiltrating lymphocytes, we examined the presence of CD4+ and CD8+ lymphocytes in these tumors. Consistent with the upregulation of Mig and IP-10, we saw significantly increased numbers of CD8+ cells and a lesser increase in CD4+ cells in tumors with elevated levels of both chemokines.     

Role in nude mice of interferon and natural killer cells in inhibiting the tumorigenicity of human hepatocellular carcinoma cells infected with hepatitis B virus

The human hepatoma cell line, PLC/PRF/5, which is persistently infected with hepatitis B virus (HBV), has integrated HBV-DNA, secretes HBV surface antigen (HBsAg), and does not grow readily in congenitally athymic (nu/nu) mice. The present investigation was undertaken to ascertain whether the low tumorigenicity of this cell line was governed by a host immune response and/or was related to expression of HBsAg. Subcutaneous injection of 4-5 X 10(6) cells into BALB/c nude mice produced localized encapsulated tumors with morphologic features of primary hepatocellular carcinoma in 25% of the animals within 29-40 d. No tumor growth was observed at lower cell inocula. In contrast, SK-HEP-1, an HBV-negative human hepatoma cell line, produced tumors at 1-5 X 10(6) cells inocula in 66% of the animals. Immunosuppression of mice with antilymphocyte serum (ALS) or irradiation increased tumor incidence in mice inoculated with 1 X 10(6) PLC/PRF/5 cells to almost 100% and produced local invasiveness. Immunosuppression also reduced the latency, i.e., time to tumor appearance, and increased mean tumor weight. These results suggest that tumorigenicity was limited by the host immune response.The nature of the response was delineated by treating nude mice challenged with tumor cells with sheep anti-mouse interferon globulin (anti-IFN). When 2 X 10(6) cells were injected, tumor growth occurred in 75% of anti-IFN-treated mice, whereas controls injected with the same number of cells, but not receiving anti-IFN, failed to develop tumors. The tumors in the anti-IFN-treated mice were highly invasive and the latency period until tumor appearance was reduced to 3-5 d. An inverse correlation was found between susceptibility of the hepatoma cells to natural killer (NK) activity in vitro and resistance to tumor growth in vivo. In vitro cytotoxicity for PLC/PRF/5 cells was eliminated by anti-NK 1.1 and complement, establishing the effector cell as an NK cell. NK cell activity 14 d after inoculation of mice with PLC/PRF/5 cells was augmented against PLC/PRF/5 target cells but not against SK-HEP-1 cells. Treatment of mice with ALS, irradiation, or anti-IFN abolished NK activity against PLC/PRF/5 cells. Co-cultivation of nude mouse spleen cells with PLC/PRF/5 but not with HBsAg or SK-HEP-1 cells induced secretion of murine IFNalpha. These results suggest that the IFN/NK cell system may play a role in limiting tumorigenicity and invasiveness of HBV-infected human hepatocellular carcinoma cells by a mechanism similar to that found for other cells persistently infected with viruses.     

卵巢肿瘤干细胞中趋化因子表达与细胞侵袭后的转移能力

背景：利用趋化因子或趋化性多肽招募免疫效应细胞汇集于肿瘤,激发抗肿瘤免疫的方法有望成为肿瘤治疗的新策略。目的：探讨卵巢肿瘤细胞中趋化因子的表达与细胞侵袭转移能力关系。方法：采用RT-PCR方法检测卵巢肿瘤细胞系SW626和Anglne细胞株中CXCR4的表达,然后在Transwell小室检测CXCL12与CXCL12/CXCR4对SW626细胞趋化活性和侵袭活性的影响。结果与结论：RT-PCR检测结果提示SW626卵巢细胞株中有CXCR4分子的表达,而在Anglne细胞中CXCR4无表达。CXCR4的表达能促使卵巢肿瘤细胞中发生趋化与侵袭转移的细胞数目增多,而加入CXCR4中和抗体时,能抑制上述效果。结果可见卵巢肿瘤细胞干细胞趋化因子CXCL12对SW626细胞有明显的趋化活性和侵袭转移活性,其活性是通过其受体CXCR4介导的。     

The CXCR3 targeting chemokine CXCL11 has potent antitumor activity in vivo involving attraction of CD8+ T lymphocytes but not inhibition of angiogenesis

The IFN-gamma-inducible and CXCR3-targeting human CXC chemokines CXCL9 (Mig) and CXCL10 (IP10) have potent antitumor activity through attraction of cytotoxic T lymphocytes and inhibition of angiogenesis. The more recently identified CXCR3-targeting chemokine CXCL11 (I-TAC/IP9) proved to be a more potent chemokine than CXCL9 and CXCL10 in vitro, both in chemotaxis assays with CXCR3+ T lymphocytes and in calcium mobilization experiments. However, its antitumor activity in vivo has not been shown so far. To investigate this, mice were challenged with EL4 T-cell lymphoma cells, genetically modified to produce murine CXCL11. Tumor growth curves showed complete rejection of CXCL11-producing tumors but not of control tumors. Tumor infiltrate analysis by flow cytometry showed a clear correlation between rejection of CXCL11-producing tumors and an increase of tumor-infiltrating CD8+CXCR3+ as well as CD8+CXCR3- T lymphocytes. In vivo CD8 T-cell depletion completely abrogated the antitumor effect. No difference in angiogenesis between control and CXCL11-producing tumors was observed. In survivors, rechallenge experiments with wild-type tumor cells suggested development of protective antitumor immunity involving tumor-specific IFN-gamma production by CD8+ T lymphocytes. These experiments show, for the first time, antitumor activity of CXCL11 in vivo, which warrants exploration for its potential role in anticancer immunotherapy.     

Requirement of the chemokine receptor CXCR3 for acute allograft rejection

Chemokines provide signals for activation and recruitment of effector cells into sites of inflammation, acting via specific G protein-coupled receptors. However, in vitro data demonstrating the presence of multiple ligands for a given chemokine receptor, and often multiple receptors for a given chemokine, have led to concerns of biologic redundancy. Here we show that acute cardiac allograft rejection is accompanied by progressive intragraft production of the chemokines interferon (IFN)-gamma-inducible protein of 10 kD (IP-10), monokine induced by IFN-gamma (Mig), and IFN-inducible T cell alpha chemoattractant (I-TAC), and by infiltration of activated T cells bearing the corresponding chemokine receptor, CXCR3. We used three in vivo models to demonstrate a role for CXCR3 in the development of transplant rejection. First, CXCR3-deficient (CXCR3(-/)-) mice showed profound resistance to development of acute allograft rejection. Second, CXCR3(-/)- allograft recipients treated with a brief, subtherapeutic course of cyclosporin A maintained their allografts permanently and without evidence of chronic rejection. Third, CXCR(+/+) mice treated with an anti-CXCR3 monoclonal antibody showed prolongation of allograft survival, even if begun after the onset of rejection. Taken in conjunction with our findings of CXCR3 expression in rejecting human cardiac allografts, we conclude that CXCR3 plays a key role in T cell activation, recruitment, and allograft destruction.     

Interleukin-10 inhibits tumor metastasis through an NK cell-dependent mechanism

Interleukin-10 (IL-10) is a recently described pleiotropic cytokine secreted mainly by type 2 helper T cells. Previous studies have shown that IL-10 suppresses cytokine expression by natural killer (NK) and type 1 T cells, thus down-regulating cell-mediated immunity and stimulating humoral responses. We here report that injected IL-10 protein is an efficient inhibitor of tumor metastasis in experimental (B16-F10) and spontaneous (M27 and Lox human melanoma) metastasis models in vivo at doses that do not have toxic effects on normal or cancer cells. Histological characterization after IL-10 treatment confirmed the absence of CD8+ and CD4+ T cells and macrophages at the sites of tumor growth, but abundant NK cells were localized at these sites. This unexpected finding was confirmed by showing that IL-10 inhibits most B16-F10 and Lox metastases in mice deficient in T or B cells (SCID and nu/nu mice), but not in those deficient in NK cells (beige mice or NK cell-depleted mice). However, IL-10 downregulation of pro-inflammatory cytokine production and/or recruitment of additional effector cells may also be involved in the anti-tumor effect at higher local concentrations of IL-10, since transfected B16 tumor cells expressing high amounts of IL-10 were rejected by normal, nu/nu, or SCID mice at the primary tumor stage, and there was still a 33% inhibition of tumor metastasis in beige mice.     

Adenovirus-mediated lymphotactin gene transfer improves therapeutic efficacy of cytosine deaminase suicide gene therapy in established murine colon carcinoma

Lymphotactin (Ltn) is the sole member of C chemokines which attracts T cells and NK cells specially. Ltn gene was transferred in vivo to improve the antitumor efficacy of cytosine deaminase (CD) gene therapy. Upregulation of CD80 and CD54 on murine CT26 colon carcinoma cells was observed after combined transfection with adenovirus encoding CD (AdCD) and adenovirus encoding murine Ltn (AdLtn) followed by administration of 5-fluorocytosine (5FC) in vitro. AdCD/5FC treatment also increased the expression of CD95 and induced obvious apoptosis of CT26 cells. After combined treatment with AdLtn and AdCD/5FC, the pre-established murine model with subcutaneous CT26 colon carcinoma exhibited most significant tumor growth inhibition, and four of eight tumor-bearing mice were tumor free, while tumors in other mice grew more progressively. Examination of lymphocyte infiltration and cytokine gene expression in tumor tissue revealed that tumors from AdLtn/AdCD/5FC-or AdLtn-treated mice were heavily infiltrated with CD4+, CD8+ T cells and NK cells, and IL-2 and IFN-gamma mRNA expression were present in parallel with T cell and NK cell infiltration. Splenic NK and CTL activities increased significantly after the combination therapy. In vivo depletion analysis showed that NK cells, CD4+ T cells and CD8+T cells participated in the antitumor effect of the host with CD8+T cells being the main T cell subset responsible for the enhanced antitumor immune response. These findings suggested that increased immunogenicity and induction of apoptosis of the tumor cells, and efficient induction of local and systemic antitumor immunity of the host might contribute to the enhanced antitumor effects of the combined Ltn and CD suicide therapy. Gene Therapy (2000) 7, 329-338.     

SDF-1及其受体CXCR4与前列腺癌的研究进展

前列腺癌远处转移是影响患者死亡率的重要原因。多项研究显示趋化因子受体CXCR4和其配体基质细胞衍生因子SDF-1能调控肿瘤细胞的器官特异性转移。近来也发现SDF-1能增强前列腺癌细胞的粘附、迁移和侵袭,提示CXCR4参与了前列腺癌转移。作为趋化因子受体CXCR4特异性阻滞剂:AMD3100、T22、T140以及人scFv在实验中已被证明能够抑制体外癌细胞的转移,在临床阻止和治疗前列腺癌转移上应用前景广阔。     

A chemokine-to-cytokine-to-chemokine cascade critical in antiviral defense

Macrophage inflammatory protein 1alpha (MIP-1alpha) promotes natural killer (NK) cell inflammation in livers during murine cytomegalovirus (MCMV) infections, and NK cell-produced interferon gamma (IFN-gamma) contributes to defense against MCMV infections. A specific role for local NK cell IFN-gamma production, however, has not been established. The importance of MIP-1alpha and NK cell-produced IFN-gamma in shaping endogenous immune responses and defense in different compartments was examined. MIP-1alpha deficiency profoundly decreased resistance to MCMV and was associated with dramatically reduced NK cell accumulation and IFN-gamma production in liver. MIP-1alpha-independent IFN-gamma responses were observed in serum and spleen, and infection-induced elevations in blood NK cell populations occurred in absence of the factor, but peak liver expression of another chemokine, the monokine induced by IFN-gamma (Mig), depended upon presence of MIP-1alpha, NK cells, and IFN-gamma. The Mig response was also important for viral resistance. Thus, serum cytokine responses are insufficient; MIP-1alpha is critical for NK cell migration and IFN-gamma delivery to mediate protection; and Mig induction in tissues is a downstream protective response resulting from the process. These results define a critical chemokine-to-cytokine-to-chemokine cascade required for defense during a viral infection establishing itself in tissues.     

HSV oncolytic therapy upregulates interferon-inducible chemokines and recruits immune effector cells in ovarian cancer.

Cooperation between oncolytic herpes simplex virus (HSV) and host effector immune mechanisms has been previously described. In the present study, we investigated the mechanism underlying such cooperation in a murine syngeneic model of ovarian carcinoma. Therapeutic administration of HSV-1716, a replication-restricted mutant, resulted in significant reduction of tumor growth and a significant survival advantage. Intratumoral injection of HSV-1716 induced expression of IFN-gamma, MIG, and IP-10 in the tumor. This was accompanied by a significant increase in the number of tumor-associated NK and CD8+ T cells expressing CXCR3 and CD25. Ascites from HSV-1716-treated animals efficiently induced in vitro migration of NK and CD8+ T cells, which was dependent on the presence of MIG and IP-10. Murine monocytes and dendritic cells (DCs) were responsible for the production of MIG and IP-10 upon HSV-1716 infection. In monocytes, this was partially abrogated by neutralizing antibodies against IFN-alpha and -beta, thus indicating a role for type-1 IFNs in the reported effect. Human ovarian carcinomas showed high numbers of monocytes and DCs. Upon HSV-1716 infection, human monocyte-derived DCs produced large amounts of IFN-gamma and upregulated MIG and IP-10 expression. These results indicate that HSV-1716 induces an inflammatory response that may facilitate antitumor immune response upon oncolytic therapy.     

Immunotherapy of murine hepatocellular carcinoma by alpha-fetoprotein DNA vaccination combined with adenovirus-mediated chemokine and cytokine expression

Most hepatocellular carcinomas (HCCs) express oncofetal alpha-fetoprotein (AFP). We and others have demonstrated efficient tumor control mediated by cellular immune responses in mice bearing subcutaneous tumors derived from the AFP-expressing murine HCC cell line Hepa 1-6 by DNA vaccination against AFP. In the present study, we examined AFP DNA vaccination in the AFP-expressing primary murine HCC model BW7756. In this model AFP DNA vaccination resulted in only minimal lymphocytic infiltration and failed to control tumor growth. To augment the AFP-specific cellular immune response, intratumoral expression of chemokine IP-10 (interferon-inducible protein-10) and the proinflammatory cytokine interleukin (IL)-12 by adenoviral vectors (AdmIL-12 and AdmIP-10) was analyzed. Intratumoral injection of AdmIL-12 and AdmIP-10 resulted in transient tumor regressions, without prolongation of animal survival. By contrast, AFP DNA vaccination followed by intratumoral injection of AdmIL-12 and AdmIP-10 resulted in tumor regression in all animals and in prolongation of animal survival; in 25% of animals the tumors became undetectable. This study demonstrates for the first time that activation of effector cells against a tumor antigen induced by the combination of DNA vaccination and intratumoral chemokine and cytokine expression is superior to the respective treatment strategies alone. This effect may be mediated by attraction of activated effector cells to the tumor tissue.     

Natural killer cell immunosuppressive function requires CXCR3-dependent redistribution within lymphoid tissues

NK cell suppression of T cells is a key determinant of viral pathogenesis and vaccine efficacy. This process involves perforin-dependent elimination of activated CD4⁺ T cells during the first 3 days of infection. Although this mechanism requires cell-cell contact, NK cells and T cells typically reside in different compartments of lymphoid tissues at steady state. Here, we showed that NK cell suppression of T cells is associated with transient accumulation of NK cells within T cell–rich sites of the spleen during lymphocytic choriomeningitis virus infection. The chemokine receptor CXCR3 was required for this relocation and suppression of antiviral T cells. Accordingly, NK cell migration was mediated by type I IFN–dependent promotion of CXCR3 ligand expression. In contrast, adenoviral vectors that weakly induced type I IFN and did not stimulate NK cell inhibition of T cells also did not promote measurable redistribution of NK cells to T cell zones. Exogenous IFN rescued NK cell migration during adenoviral vector immunization. Thus, type I IFN and CXCR3 were critical for properly positioning NK cells to constrain antiviral T cell responses. Development of strategies to curtail migration of NK cells between lymphoid compartments may enhance vaccine-elicited immune responses.     

CD4+CD25+ regulatory T cells inhibit natural killer cell functions in a transforming growth factor-beta-dependent manner

Tumor growth promotes the expansion of CD4+CD25+ regulatory T (T reg) cells that counteract T cell-mediated immune responses. An inverse correlation between natural killer (NK) cell activation and T reg cell expansion in tumor-bearing patients, shown here, prompted us to address the role of T reg cells in controlling innate antitumor immunity. Our experiments indicate that human T reg cells expressed membrane-bound transforming growth factor (TGF)-beta, which directly inhibited NK cell effector functions and down-regulated NKG2D receptors on the NK cell surface. Adoptive transfer of wild-type T reg cells but not TGF-beta-/- T reg cells into nude mice suppressed NK cell-mediated cytotoxicity, reduced NKG2D receptor expression, and accelerated the growth of tumors that are normally controlled by NK cells. Conversely, the depletion of mouse T reg cells exacerbated NK cell proliferation and cytotoxicity in vivo. Human NK cell-mediated tumor recognition could also be restored by depletion of T reg cells from tumor-infiltrating lymphocytes. These findings support a role for T reg cells in blunting the NK cell arm of the innate immune system.     

Suppression of tumor formation in lymph nodes by L-selectin-mediated natural killer cell recruitment

Natural killer (NK) cells are known to reject certain tumors in vivo; however, the ability of NK cells to prevent metastasis of tumors into secondary lymphoid organs has not been addressed. Here, we report that in tumor-bearing hosts, NK cells are recruited to regional lymph nodes in wild-type mice, but not in mice deficient for L-selectin or L-selectin ligands. By adoptive transfer and complete Freund's adjuvant stimulation experiments, we demonstrated that L-selectin on NK cells and L-selectin ligands on endothelial cells are essential for NK cell recruitment to lymph nodes. Furthermore, freshly isolated resident lymph node NK cells lysed tumors efficiently, and metastasis of B16 melanoma cells to draining lymph nodes was suppressed in wild-type or Rag-1-deficient mice, but not when NK cells were depleted. Although L-selectin-deficient NK cells efficiently lysed tumor cells in vitro, NK cell-dependent suppression of tumor metastasis was diminished in mice deficient for L-selectin or L-selectin ligands because of insufficient NK cell recruitment to lymph nodes. Moreover, tumor metastasis was substantially inhibited in L-selectin-deficient mice reconstituted with wild-type NK cells. These findings indicate that L-selectin-mediated NK cell recruitment plays a crucial role in the control of tumor metastasis into secondary lymphoid organs.     

RNAi targeting CXCR4 inhibits tumor growth through inducing cell cycle arrest and apoptosis

CXC chemokine receptor 4 (CXCR4) is involved in many human malignant tumors and plays an important role in tumor growth and metastasis. To explore the effects of CXCR4 expression on the malignant cells of oral squamous cell carcinoma (OSCC), Tca8113 and SCC-9 cell lines, as well as their xenograft models, of nude mice were used to detect cancer cell proliferation alteration. This study also examined the corresponding molecular mechanism after CXCR4 knockdown using a recombinant lentiviral vector expressing small interference RNA (siRNA) for CXCR4. RNA interference-mediated knockdown of CXCR4 in highly aggressive (Tca8113 and SCC-9) tumor cells significantly inhibited the proliferation of the two cell lines in vitro and in vivo. The expression levels of >1,500 genes involved in cell cycle, apoptosis, and multiple signaling pathways were also altered. These results provide new evidence of CXCR4 as a promising tumor gene therapeutic target.     

Myeloid-derived suppressor cells are implicated in regulating permissiveness for tumor metastasis during mouse gestation

Metastasis depends on the ability of tumor cells to establish a relationship with the newly seeded tissue that is conducive to their survival and proliferation. However, the factors that render tissues permissive for metastatic tumor growth have yet to be fully elucidated. Breast tumors arising during pregnancy display early metastatic proclivity, raising the possibility that pregnancy may constitute a physiological condition of permissiveness for tumor dissemination. Here we have shown that during murine gestation, metastasis is enhanced regardless of tumor type, and that decreased NK cell activity is responsible for the observed increase in experimental metastasis. Gene expression changes in pregnant mouse lung and liver were shown to be similar to those detected in premetastatic sites and indicative of myeloid cell infiltration. Indeed, myeloid-derived suppressor cells (MDSCs) accumulated in pregnant mice and exerted an inhibitory effect on NK cell activity, providing a candidate mechanism for the enhanced metastatic tumor growth observed in gestant mice. Although the functions of MDSCs are not yet understood in the context of pregnancy, our observations suggest that they may represent a shared mechanism of immune suppression occurring during gestation and tumor growth.     

Interaction between CXCR4 and CCL20 Pathways Regulates Tumor Growth

The chemokine receptor CXCR4 and its ligand CXCL12 is overexpressed in the majority of tumors and is critically involved in the development and metastasis of these tumors. CXCR4 is expressed in malignant tumor cells whereas its ligand SDF-1 (CXCL12) is expressed mainly by cancer associated fibroblasts (CAF). Similarly to CXCR4, the chemokine CCL20 is overexpressed in variety of tumors; however its role and regulation in tumors is not fully clear. Here, we show that the chemokine receptor CXCR4 stimulates the production of the chemokine CCL20 and that CCL20 stimulates the proliferation and adhesion to collagen of various tumor cells. Furthermore, overexpression of CCL20 in tumor cells promotes growth and adhesion in vitro and increased tumor growth and invasiveness in vivo. Moreover, neutralizing antibodies to CCL20 inhibit the in vivo growth of tumors that either overexpress CXCR4 or CCL20 or naturally express CCL20. These results reveal a role for CCL20 in CXCR4-dependent and -independent tumor growth and suggest a therapeutic potential for CCL20 and CCR6 antagonists in the treatment of CXCR4- and CCL20-dependent malignancies.     

Lymphotactin cotransfection enhances the therapeutic efficacy of dendritic cells genetically modified with melanoma antigen gp100

Lymphotactin (Lptn) is a C chemokine that attracts T cells and NK cells. Dendritic cells (DC) are highly efficient, specialized antigen-presenting cells and antigen-pulsed DC has been regarded as promising vaccines in cancer immunotherapy. The aim of our present study is to improve the therapeutic efficacy of DC-based tumor vaccine by increasing the preferential chemotaxis of DC to T cells. In this study, Lptn and/or melanoma-associated antigen gp100 were transfected into mouse bone marrow-derived DC, which were used as vaccines in B16 melanoma model. Immunization of C57BL/6 mice with DC adenovirally cotransfected with Lptn and gp100 (Lptn/gp100-DC) could enhance the cytotoxicities of CTL and NK cells, increase the production of IL-2 and interferon-gamma significantly, as compared with immunization with gp100-DC, Lptn-DC, LacZ-DC, DC or PBS counterparts. The Lptn/gp100-DC immunized mice exhibited resistance to tumor challenge most effectively. It was found that the tumor mass of mice vaccinated by Lptn/gp100-DC showed obvious necrosis and inflammatory cell infiltration. In vivo depletion analysis demonstrated that CD8(+) T cells are the predominant T cell subset responsible for the antitumor effect of Lptn/gp100-DC and CD4(+) T cells were necessary in the induction phase of tumor rejection, while NK cells were less important although they participated in the antitumor response either in the induction phase or in the effector phase. In the murine model with the pre-established subcutaneous B16 melanoma, immunization with Lptn/gp100-DC inhibited the tumor growth most significantly when compared with other counterparts. These findings provide a potential strategy to improve the efficacy of DC-based tumor vaccines.