Chemoprotective effects of recombinant human IL-1 alpha in cyclophosphamide-treated normal and tumor-bearing mice. Protection from acute toxicity, hematologic effects, development of late mortality, and enhanced therapeutic efficacy

In this study, recombinant human IL-1 alpha (rhIL-1 alpha) was used to protect normal and tumor-bearing BALB/c mice from the acute toxicity caused by lethal doses of cyclophosphamide (Cy) and 5-fluorouracil. Pretreatment of mice for 7 days with 10,000 U/day of rhIL-1 alpha protected 70 to 100% of mice from the acute death induced by lethal doses of both Cy (380 mg/kg) and 5-fluorouracil (250 mg/kg). In contrast, post-treatment of mice with single or multiple doses of rhIL-1 alpha was not chemoprotective. Pretreatment of mice with rhIL-1 alpha increased the acute LD90 of Cy from 380 mg/kg to greater than 500 mg/kg in normal mice, an LD90 dose-modifying effect of at least 1.25, was accompanied by a more rapid recovery from neutropenia and a less severe reduction in the number of bone marrow single lineage monocyte, myeloid, or myelomonocytic colonies. Some of the mice (10 to 50%) that were successfully protected by pretreatment with rhIL-1 alpha died after day 50. These mice consistently presented with extensive pulmonary inflammation and fibrosis at death. Mice bearing murine renal cancer (Renca) were also protected from the acute toxic effects of Cy (450 mg/kg) by pretreatment with rhIL-1 alpha. Renca-bearing mice pretreated with rhIL-1 alpha and either sublethal (300 mg/kg) or lethal (450 mg/kg) doses of Cy exhibited enhanced survival times over those of untreated Renca-bearing mice. Interestingly, the cause of death in Renca-bearing mice that ultimately failed treatment with rhIL-1 alpha plus 300 mg/kg Cy was recurrent tumor, whereas most mice treated with rhIL-1 alpha plus 450 mg/kg Cy had no detectable tumor, although several died from late pulmonary inflammation and fibrosis. Thus, the dose escalation of Cy in rhIL-1 alpha-pretreated mice results in greater antitumor effects of Cy. However, the dose escalation of some cytotoxic agents allowed by the use of myelostimulatory agents can result in late fatal complications not detected in acute toxicity testing.     

Tumor Vasculature-Targeted Recombinant Mutated Human TNF-α Enhanced the Antitumor Activity of Doxorubicin by Increasing Tumor Vessel Permeability in Mouse Xenograft Models

Objective

Increasing evidence suggests that, when used in combination, tumor necrosis factor-α (TNF-α) synergizes with traditional chemotherapeutic drugs to exert a heightened antitumor effect. The present study investigated the antitumor efficacy of recombinant mutated human TNF-α specifically targeted to the tumor vasculature (RGD-rmhTNF-α) combined with the chemotherapeutic agent doxorubicin in 2 murine allografted tumor models.

Methods

Mice bearing hepatoma or sarcoma allografted tumors were treated with various doses of RGD-rmhTNF-α alone or in combination with doxorubicin (2 mg/kg). We then evaluated tumor growth and tumor vessel permeability as well as intratumoral levels of RGD-rmhTNF-α and doxorubicin.

Results

RGD-rmhTNF-α treatment enhanced the permeability of the tumor vessels and increased intratumoral doxorubicin levels. In addition, intratumoral RGD-rmhTNF-α levels were significantly higher than that of rmhTNF-α. In both of the tested tumor models, administering RGD-rmhTNF-α in combination with doxorubicin resulted in an enhanced antitumor response compared to either treatment alone. Double-agent combination treatment of doxorubicin with 50,000 IU/kg RGD-rmhTNF-α induced stronger antitumor effects on H22 allografted tumor-bearing mice than the single doxorubicin agent alone. Moreover, doxorubicin with 10,000 IU/kg RGD-rmhTNF-α synergized to inhibit tumor growth in S180 allografted tumor-bearing mice.

Conclusions

These results suggest that targeted delivery of low doses of RGD-rmhTNF-α into the tumor vasculature increases the antitumor efficacy of chemotherapeutic drugs.     

IFN-gamma mediates CD4+ T-cell loss and impairs secondary antitumor responses after successful initial immunotherapy

Protective cell-mediated immune responses in cancer are critically dependent on T-helper type 1 (T(H)1) cytokines such as interferon-gamma (IFN-gamma). We have previously shown that the combination of CD40 stimulation and interleukin-2 (IL-2) leads to synergistic antitumor responses in several models of advanced metastatic disease. We now report that after this treatment and other immunotherapy regimens, the CD4+ T-cell population, in contrast to CD8+ T cells, did not significantly increase but rather exhibited a substantial level of apoptosis that was dependent on IFN-gamma. Mice immunized with tumor cells and treated with an immunotherapy regimen that was initially protective were later unable to mount effective memory responses compared with immunized mice not receiving immunotherapy. Immunotherapy given to tumor-bearing Ifngr-/- mice resulted in restoration of secondary responses. Thus, although immunotherapeutic regimens inducing strong IFN-gamma responses can lead to successful early antitumor efficacy, they may also impair the development of durable antitumor responses.     

Synergistic antitumor effect of anti-PD-L1 combined with oxaliplatin on a mouse tumor model

Oxaliplatin (OXP) can change tumor microenvironment from immune-suppressive toward the immune-favorable condition. Almost all of the antitumor agents cannot totally cure cancer as monotherapy. So the current focus of cancer research became combining therapy using different treatment regimen, especially chemotherapy with checkpoint blockers. In this study, we assessed the activity of combining regimen using anti-PD-L1 with OXP in CT26 tumor-bearing BALB/c mice. We further analyzed the immune cell phenotypes in tumor site, lymph nodes, and spleen by flow cytometry analysis. Our study showed that combination therapy with OXP and anti-PD-L1 significantly increased survival in vivo and inhibited tumor growth of tumor-bearing mice. Inconsistent with better antitumor activity, our combination therapy led to an increase in tumor-infiltrating activated CD8+ T cells. In draining lymph nodes and spleen, regulatory T cells decreased significantly. Mice receiving either anti-PD-L1 or OXP alone had a larger tumor and lower survival rate in comparison with combination therapy receiving group. The time and order of administration of each component of the combination therapy affected antitumor response.     

Anti-CD40 antibody induces antitumor and antimetastatic effects: the role of NK cells

We assessed the effect of the stimulatory anti-CD40 Ab on NK cell activation in vivo and the therapeutic potential of activated NK cells in tumor-bearing mice. Single-dose i.p. injection of the anti-CD40 Ab resulted in production of IL-12 and IFN-gamma in vivo, followed by a dramatic increase in NK cell cytolytic activity in PBLs. NK cell activation by anti-CD40 Ab was also observed in CD40 ligand knockout mice. Because NK cells express CD40 ligand but not CD40, our results suggest that NK activation is mediated by increased cytokine production upon CD40 ligation of APCs. Treatment of tumor-bearing mice with anti-CD40 Ab resulted in substantial antitumor and antimetastatic effects in three tumor models. Depletion of NK cells with anti-asialo GM1 Ab reduced or abrogated the observed antitumor effects in all the tested models. These results indicate that a stimulatory CD40 Ab indirectly activates NK cells, which can produce significant antitumor and antimetastatic effects.     

Progesterone antagonists and progesterone receptor modulators in the treatment of breast cancer

Progesterone antagonists (PAs) (antiprogestins) or progesterone receptor modulators (PRMs) form an interesting category of new hormonal agents in the treatment of breast cancer. In vitro, antiproliferative effects of different PAs are mainly observed in estrogen-stimulated growth of PR-positive tumor cell lines. Both progestin agonist/antagonist actions on mammary tumor cells are dependent on the type of cell line, culture medium and concentrations of the PAs used, and type of biologic response measured. In various experimental animal tumor models, different PAs showed a greater antitumor activity than tamoxifen or high-dose progestins. Most interestingly, combination treatment of different PAs (mifepristone, ORG 31710, onapristone) or PRMs with different antiestrogens (tamoxifen, droloxifen, ICI 164384) or with an aromatase inhibitor (atemestane) showed greater antitumor efficacy than treatment with each single type of drug alone. These additive antiproliferative effects were demonstrated in various experimental in vitro and in vivo models. In some studies, these effects were accompanied by additive effects on several cell biologic parameters. In pretreated postmenopausal patients with metastatic breast cancer, objective responses have been observed in 10-12%, and stable disease in 42-46% of the patients; in previously untreated patients objective response rates of 11 and 56% have been reported. The clinical development of onapristone was stopped because of liver toxicity. At the present time, apart from development of new pure potent PAs, clinical investigation of combined therapy of PAs with antiestrogens are urgently needed.     

Interleukin-21 triggers both cellular and humoral immune responses leading to therapeutic antitumor effects against head and neck squamous cell carcinoma

BACKGROUND: Interleukin-21 (IL-21) plays important roles in the regulation of T, B, and natural killer (NK) cells. We hypothesized that the cytokine may provide a novel immunotherapy strategy for cancer by stimulating both Th1 and Th2 immune responses. In this context, antitumor immunity induced by IL-21 was examined in mice bearing subcutaneous head and neck squamous cell carcinomas (HNSCC). METHODS: A plasmid vector encoding murine IL-21 was injected intravenously into mice with pre-established HNSCC tumors, either alone or in combination with a vector construct expressing IL-15. Cytotoxic T lymphocyte (CTL) and NK killing activities were evaluated by chrome release assays, while HNSCC-specific antibody was examined by flow cytometry and ELISA. RESULTS: Significant antitumor effects were obtained by repeated transfection with either the IL-21 or the IL-15 gene. Co-administration of both cytokine genes resulted in increased suppression of tumor growth, significantly prolonging the survival periods of the animals. Thirty percent of the tumor-bearing mice that received the combination therapy survived for more than 300 days, completely rejecting rechallenge with the tumor at a distant site. IL-21 induced significant elevation of HNSCC-specific CTL activity, while IL-21 and IL-15 augmented NK activity in an additive manner. IL-21 gene transfer also promoted the production of tumor-specific IgG. CONCLUSIONS: In vivo transduction of the IL-21 gene elicits powerful antitumor immunity, including both humoral and cellular arms of the immune response, and results in significant suppression of pre-established HNSCC. Co-transfer of the IL-15 gene further improved the therapeutic outcome, mainly by augmenting NK tumoricidal activity. The biological effects of IL-21 may be in sharp contrast to those of conventional Th1 and Th2 cytokines, suggesting intriguing implications of this cytokine for the classical concept of Th1 vs. Th2 paradigm.     

Potent antitumor effects of combination therapy with IFNs and monocytes in mouse models of established human ovarian and melanoma tumors

Interferon-activated monocytes are known to exert cytocidal activity against tumor cells in vitro. Here, we have examined whether a combination of IFN-α2a and IFN-γ and human monocytes mediate significant antitumor effects against human ovarian and melanoma tumor xenografts in mouse models. OVCAR-3 tumors were treated i.t. with monocytes alone, IFN-α2a and IFN-γ alone or combination of all three on day 0, 15 or 30 post-tumor implantation. Mice receiving combination therapy beginning day 15 showed significantly reduced tumor growth and prolonged survival including complete regression in 40% mice. Tumor volumes measured on day 80 in mice receiving combination therapy (206 mm(3)) were significantly smaller than those of mice receiving the IFNs alone (1,041 mm(3)), monocytes alone (1,111 mm(3)) or untreated controls (1,728 mm(3)). Similarly, combination therapy with monocytes and IFNs of much larger tumor also inhibited OVCAR-3 tumor growth. Immunohistochemistry studies showed a large number of activated macrophages (CD31(+)/CD68(+)) infiltrating into OVCAR-3 tumors and higher densities of IL-12, IP10 and NOS2, markers of M1 (classical) macrophages in tumors treated with combination therapy compared to the controls. Interestingly, IFNs-activated macrophages induced apoptosis of OVCAR-3 tumor cells as monocytes alone or IFNs alone did not mediate significant apoptosis. Similar antitumor activity was observed in the LOX melanoma mouse model, but not as profound as seen with the OVCAR-3 tumors. Administration of either mixture of monocytes and IFN-α2a or monocytes and IFN-γ did not inhibit Lox melanoma growth; however, a significant inhibition was observed when tumors were treated with a mixture of monocytes, IFN-α2a and IFN-γ. These results indicate that monocytes and both IFN-α2a and IFN-γ may be required to mediate profound antitumor effect against human ovarian and melanoma tumors in mouse models.     

Preclinical Combination Therapy of Thiarabine Plus Various Clinical Anticancer Agents

Thiarabine is undergoing clinical trials. In support of that effort combination therapy of thiarabine plus six clinical anticancer agents was evaluated using various human tumor xenograft models. The antitumor activity of thiarabine in combination appeared to be greater than additive with irinotecan (DLD-1 colon), paclitaxel (PC-3 prostate), cisplatin (PC-3 prostate), or cyclophosphamide (RL lymphoma), additive with irinotecan (NCI-H460 NSCLC), cisplatin (NCI-H460 NSCLC) or methotrexate (CCRF-CEM leukemia), and less than additive with irinotecan (HT29 colon), paclitaxel (NCI-H460 NSCLC) or cisplatin (NCI-H23 NSCLC). Combining thiarabine with irinotecan, paclitaxel, cisplatin, or cyclophosphamide should receive consideration in the clinical treatment of cancer.     

Combination treatment with MEK and AKT inhibitors is more effective than each drug alone in human non-small cell lung cancer in vitro and in vivo

AZD6244 and MK2206 are targeted small-molecule drugs that inhibit MEK and AKT respectively. The efficacy of this combination in lung cancer is unknown. Our previous work showed the importance of activated AKT in mediating resistance of non-small cell lung cancer (NSCLC) to AZD6244. Thus we hypothesized that dual inhibition of both downstream MEK and AKT pathways would induce synergistic antitumor activity. In this study, we evaluated the efficacy of AZD6244 and MK2206 individually on a large panel of lung cancer cell lines. Then, we treated 28 human lung cancer cell lines with a combination of AZD6244 and MK2206 at clinically applicable drug molar ratios. The AZD6244-MK2206 combination therapy resulted in a synergistic effect on inhibition of lung cancer cell growth compared to the results of single drug treatment alone. MK2206 enhanced AZD6244-induced Bim overexpression and apoptosis in A549 and H157 cells. When we tested the combination of AZD6244 and MK2206 at ratios of 8∶1, 4∶1, 2∶1, and 1∶8, we found that the synergistic effect of the combination therapy was ratio-dependent. At ratios of 8∶1, 4∶1, and 2∶1, the drug combination consistently demonstrated synergy, whereas decreasing the ratio to 1∶8 resulted in a loss of synergy and produced an additive or antagonistic effect in most cell lines. Furthermore, the AZD6244-MK2206 combination therapy showed synergy in the suppression of A549 and H157 xenograft tumor growth and increased mean animal survival time. The AZD6244-MK2206 combination therapy resulted in effective inhibition of both p-ERK and p-AKT expression in tumor tissue. In addition, a significant increase of apoptosis was detected in tumor tissue from mice treated with AZD6244-MK2206 compared with that from the single agent treated mice. Our study suggests that the combination of AZD6244 and MK2206 has a significant synergistic effect on tumor growth in vitro and in vivo and leads to increased survival rates in mice bearing highly aggressive human lung tumors.     

Effect of in vitro interferon treatment on the rapid clearance of murine leukemia cells in vivo.

Previous work showed that interferon (IFN) can protect target cells from NK mediated lysis in vitro. In the present study we investigate the effect of IFN alpha/beta or IFN gamma treatment of three different murine leukemia cell lines. For this purpose FLC-745 (susceptible to the antiproliferative activity of IFN alpha/beta and gamma), FLC-3C18 (IFN alpha/beta -resistant and IFN gamma - susceptible) of DBA/2 origin and EL-4 (IFN alpha/beta - susceptible and IFN gamma - resistant) leukemia of C57B1/6 origin were treated with IFN alpha/beta or gamma in vitro and assayed for their susceptibility to natural resistance measured in vivo as organ rapid clearance 4 hr after iv injection into syngeneic mice. Using young or Poly I:C stimulated hosts, but not mice with low levels of natural resistance (i.e. older animals or mice treated with cyclophosphamide), slower elimination of treated cells was observed with: (a) FLC-745 cells treated with IFN alpha/beta and IFN gamma and (b) FLC 3C18 treated with IFN gamma. Such a delayed clearance was not observed with: (a) FLC-3C18 cells treated with IFN alpha/beta and (b) EL-4 leukemia cells preincubated with IFN alpha/beta or IFN gamma. These results suggest that under selected conditions IFNs can protect leukemic cells from in vivo natural reactivity.     

Induction of interferons (IFNs) and tumor necrosis factor (TNF) in mice by a novel glycolipid trehalose 2,3,6'-trimycolate from Rhodococcus aurantiacus (Gordona aurantiaca).

The immunomodifying activity of a novel mycoloyl glycolipid, trehalose 2,3,6'-trimycolate (GaGM), from a unique psychrophilic acid-fast bacterium, Rhodococcus aurantiacus, was examined. ICR mice were primed intravenously (i.v.) or intraperitoneally (i.p.) with liposomes containing GaGM (300 micrograms/mouse), and were administered LPS dissolved in saline (25 micrograms/mouse, i.v.) 2 weeks later. Two hours after injection of LPS, interferons (IFNs) and tumor necrosis factor (TNF) were induced significantly in mice sera. The increase in activities of IFNs and TNF was approximately paralleled with granuloma formation in spleen of mice primed with GaGM. However, IFNs and TNF were not induced either in mice primed with GaGM but not elicited with LPS, or in those primed with GaGM and elicited by GaGM. Both activities induced were lower in mice primed with trehalose mono- or dimycolate from R. aurantiacus (GaTMM, GaTDM) or TDM from Nocardia rubra than in GaGM-primed mice. Time course study showed that the maximum activity of each interferon (alpha, beta, or gamma) was observed at different stages after LPS administration; IFN-alpha, IFN-beta, and IFN-gamma appeared 3, 2, and 6 hours most abundantly after LPS administration, respectively.     

Additive and synergistic effects of a novel antiestrogen, toremifene (Fc-1157a), and human interferons on estrogen responsive MCF-7 cells in vitro

The effect of human interferons alpha and gamma alone and in combination with a novel antiestrogen toremifene were studied in vitro using MCF-7 cell line, an estrogen receptor positive and antiestrogen sensitive cell line. The effects were evaluated by a simple bioluminescence method with which the number of living cells was obtained as cellular adenosine triphosphate (ATP) content. The growth of MCF-7 cells was inhibited both by interferon alpha and interferon gamma. At least additive effect was evident when the cells were exposed to combination of interferons and toremifene: the combination was additive with interferon gamma + toremifene and synergistic with interferon alpha + toremifene. The combination of toremifene and interferons may have clinical importance.     

Combined immunostimulation in the prevention of tumor take in mice using endotoxins,their derivatives,and other immune adjuvants

Summary In this study a variety of immunostimulatory agents was tested alone or in combination with other agents to measure their ability to enhance nonspecific resistance to challenge with the TA3-Ha transplantable murine ascites tumor. Lipopolysaccharides (LPS) were effective alone in protecting mice from tumor. Their lipid-free, nontoxic, polysaccharide-rich hydrolytic products (PS) showed a much reduced, but still significant effect in anti-TA3-Ha resistance induction. Other agents that stimulated tumor resistance included nontoxic native hapten and trehalose-6,6-dimycolate (P3) of Ribi, synthetic glycolipids, cord factor, and killed BCG preparations. Simultaneous pretreatment with a combination of any of these agents and either LPS or PS resulted in a significantly higher level of tumor resistance. Administration of LPS, PS, or native hapten to mice that had been previously infected with viable BCG resulted in the strongest antitumor effect. These studies demonstrate that nonspecific resistance can be enhanced in an additive or synergistic manner by using combinations of agents, which presumably stimulate different cell populations of the host's immune system.     

Susceptibility of different leukemic cell lines to the anticellular and antiviral effects of interferons

A total of 18 different types of human leukemic cell lines were tested for their susceptibility to the anticellular and antiviral effects of interferons (IFNs) alpha, beta and gamma. In general, only the three myelogenous leukemic cell lines U937, KG-1 and HL-60 were found to be highly susceptible to the anticellular effect of the different IFNs while cells of the other lineages were relatively resistant. In order to determine whether the cell lines were sensitive to the antiviral effects of IFNs, the cells were first screened for their ability to support the replication of vesicular stomatitis virus (VSV), sindbis virus (SBV) and semliki forest virus (SFV). Unexpectedly, only three cell lines--Raji, K562 and U937 were highly susceptible to SFV while other cell lines were relatively refractory to all three viruses. Using SFV as indicator virus, the antiviral activity of all IFNs could be detected in all three cell lines and their relative efficiency was in the order of alpha greater than beta greater than gamma. The significance of these results were discussed.     

Sensitivity of chronic myeloid leukemia hemopoietic progenitors to PTT-119 in combination with human recombinant interferon alpha and gamma

PTT-119, a new synthetic alkylating compound, has shown a marked "in vitro" inhibitory effect on chronic myeloid leukemia (CML) granulo-monocytic precursors (CFU-GM) at doses greater than 5 micrograms/ml. Based on previous experiences of synergistic associations between alkylating drugs and biological modifiers, we tested the effects of low doses of PTT-119 (from 0.1 to 1 microgram/ml) in concert with alpha, gamma, or alpha + gamma interferons and compared to IFNs alone, in order to investigate an alternative choice for treatment of CML patients in chronic phase. Our results showed a significantly higher CFU-GM cloning inhibition after addition of 100 or 1,000 U/ml of alpha IFN to 0.1 microgram/ml PTT-119 (from 39.6% +/- 26.6 SD to 80.7% +/- 10 SD and 91.5% +/- 8 SD, respectively), while gamma IFN resulted in only a slight increase in colony growth inhibition when compared to the drug used alone. The association of alpha plus gamma IFN coupled with PTT-119 treatment did not significantly improve the results observed after exposure of leukemic progenitors to PTT-119 and alpha IFN alone. We conclude that a combined treatment with PTT-119 and IFN is probably worth testing both for purging methods before autologous bone marrow transplantation and for in vivo administration in chronic myeloid leukemia.     

Paraproteins and primary lymphoma in SJL mice. I. Individuality of idiotypes on paraproteins

The isotype distribution and idiotypic determinants of paraproteins (PP) found in sera from aging SJL mice were studied. One or more PP were found in 79% of 9-11 month old mice; the isotypes were gamma 1 greater than or equal to alpha greater than or equal to gamma 2a much greater than gamma 3 much greater than gamma 2b. There was a significant tendency for the PP to occur in pairs or triplets, multiple PP being found in 35% of the mice without an evident association between any two isotypes. Rabbit anti-Id was prepared against four isolated PP, two gamma 1 and two gamma 2a. Inhibition of anti-Id-Id interaction was specific for the immunizing PP only and was absent with all other PP studied (8 gamma 1 and 10 gamma 2 PP) and with normal SJL Ig. It is concluded that SJL PP fail to exhibit cross-reactive idiotypes and show a wider range of isotype distribution than was previously suspected, although PP of mu, delta, and epsilon isotypes are absent.     

Cell-associated double-stranded RNA enhances antitumor activity through the production of type I IFN

The efficacy of tumor cell vaccination largely depends on the maturation and activation status of the dendritic cell. Here we investigated the ability of soluble and tumor cell-associated dsRNA to serve as an adjuvant in the induction of protective adaptive antitumor responses. Our data showed that cell-associated dsRNA, but not soluble dsRNA, enhanced both tumor-specific CD8(+) and CD4(+) T cell responses. The cell-associated dsRNA increased the clonal burst of tumor-specific CD8(+) T cells and endowed them with an enhanced capacity for expansion upon a secondary encounter with tumor Ags, even when the CD8(+) T cells were primed in the absence of CD4(+) T cell help. The adjuvant effect of cell-associated dsRNA was fully dependent on the expression of TLR3 by the APCs and their subsequent production of type I IFNs, as the adjuvant effect of cell-associated dsRNA was completely abrogated in mice deficient in TLR3 or type I IFN signaling. Importantly, treatment with dsRNA-associated tumor cells increased the number of tumor-infiltrating lymphocytes and enhanced the survival of tumor-bearing mice. The data from our studies suggest that using cell-associated dsRNA as a tumor vaccine adjuvant may be a suitable strategy for enhancing vaccine efficacy for tumor cell therapy in cancer patients.     

EBV-induced molecule 1 ligand chemokine (ELC/CCL19) promotes IFN-gamma-dependent antitumor responses in a lung cancer model

The antitumor efficacy of EBV-induced molecule 1 ligand CC chemokine (ELC/CCL19) was evaluated in a murine lung cancer model. The ability of ELC/CCL19 to chemoattract both dendritic cells and T lymphocytes formed the rationale for this study. Compared with diluent-treated tumor-bearing mice, intratumoral injection of recombinant ELC/CCL19 led to significant systemic reduction in tumor volumes (p < 0.01). ELC/CCL19-treated mice exhibited an increased influx of CD4 and CD8 T cell subsets as well as dendritic cells at the tumor sites. These cell infiltrates were accompanied by increases in IFN-gamma, MIG/CXCL9, IP-10/CXCL10, GM-CSF, and IL-12 but a concomitant decrease in the immunosuppressive molecules PGE(2) and TGFbeta. Transfer of T lymphocytes from ELC/CCL19 treated tumor-bearing mice conferred the antitumor therapeutic efficacy of ELC/CCL19 to naive mice. ELC/CCL19 treated tumor-bearing mice showed enhanced frequency of tumor specific T lymphocytes secreting IFN-gamma. In vivo depletion of IFN-gamma, MIG/CXCL9, or IP-10/CXCL10 significantly reduced the antitumor efficacy of ELC/CCL19. These findings provide a strong rationale for further evaluation of ELC/CCL19 in tumor immunity and its use in cancer immunotherapy.