Long-lasting Antitumor Effects Research Articles

A Case Report of Putative Autoimmunity to Prostate Cancer After Immune-Mediated Rejection of Melanoma Through a Shared Tumor Antigen

Patients with metastatic melanoma or castration-resistant metastatic prostate cancer have limited life expectancy. We report the case of a patient with metastatic melanoma who was free of disease progression for 18 years and whose prostate cancer, diagnosed 14 years after his melanoma diagnosis, entered spontaneous (though transient) remission. We found that the patient produced antibodies and T cells specific for prostate-specific membrane antigen (PSMA), suggesting that he developed auto-immunity to prostate cancer after immune-mediated rejection of the melanoma. We propose that an immune response to a shared tumor antigen resulted in long-lasting anti-tumor effects against both malignancies.

Selected anti-tumor vaccines merit a place in multimodal tumor therapies

Multimodal approaches are nowadays successfully applied in cancer therapy. Primary locally acting therapies such as radiotherapy (RT) and surgery are combined with systemic administration of chemotherapeutics. Nevertheless, the therapy of cancer is still a big challenge in medicine. The treatments often fail to induce long-lasting anti-tumor responses. Tumor recurrences and metastases result. Immunotherapies are therefore ideal adjuncts to standard tumor therapies since they aim to activate the patient's immune system against malignant cells even outside the primary treatment areas (abscopal effects). Especially cancer vaccines may have the potential both to train the immune system against cancer cells and to generate an immunological memory, resulting in long-lasting anti-tumor effects. However, despite promising results in phase I and II studies, most of the concepts finally failed. There are some critical aspects in development and application of cancer vaccines that may decide on their efficiency. The time point and frequency of medication, usage of an adequate immune adjuvant, the vaccine's immunogenic potential, and the tumor burden of the patient are crucial. Whole tumor cell vaccines have advantages compared to peptide-based ones since a variety of tumor antigens (TAs) are present. The master requirements of cell-based, therapeutic tumor vaccines are the complete inactivation of the tumor cells and the increase of their immunogenicity. Since the latter is highly connected with the cell death modality, the inactivation procedure of the tumor cell material may significantly influence the vaccine's efficiency. We therefore also introduce high hydrostatic pressure (HHP) as an innovative inactivation technology for tumor cell-based vaccines and outline that HHP efficiently inactivates tumor cells by enhancing their immunogenicity. Finally studies are presented proving that anti-tumor immune responses can be triggered by combining RT with selected immune therapies.

Absence of LTB4/BLT1 Axis Promotes Generation of Long-Lasting Antitumor Memory Responses Induced by Administration of GM-CSF Gene-Transduced Tumor Cells, in a CD4+ T Cell-Dependent Manner,

Abstract 3246GM-CSF (Granulocyte macrophage-colony stimulating factor) gene-transduced leukemia cell vaccine has been reported to be promising approach to enhance antitumor immune response in leukemia patients. The enhancement of this specific antitumor immunity is considered to be clinically beneficial to maintain complete remission for long time after chemotherapy without stem cell transplantion. Leukotriene B4 (LTB4) is an extremely potent lipid inflammatory mediator derived from membrane phospholipids, known to recruit and activate leukocytes including neutrophils, mediated by the same class of receptors, the GPCR (G-protein coupled receptor) superfamily, BLT1 and BLT2. So far, the role of LTB4 in tumor immunology is not well known. Previously, we demonstrated that the GM-CSF gene transduction into murine monocytic leukemia cell line of WEHI3B (WGM) eliminated the tumorigenicity in subcutaneous challenge model using wild type (WT) BALB/c mice. At day 50 after the challenge, we rechallenged WEHI3B cells into the opposite flank of both WT and BLT1-KO mice which had rejected WGM cells (WT/WGM, KO/WGM, respectively). Intriguingly, more KO/WGM mice re-rejected the rechallenged WEHI3B cells and showed significantly prolonged survival compared with WT/WGM mice.To clarify this unique mechanism of the long-lasting antitumor effects observed in KO/WGM mice, we performed following immunological assays. Our in vivo immune cell depletion assays (NK, CD4+ T, CD8+ T cell) showed that KO/WGM mice treated with anti-CD4 antibody displayed rather enhanced tumor growth compared with WT/WGM mice. Thus, we next compared the rates of different subsets of memory T cells, so called memory stem T cell subsets (TSCM), central memory T cell subsets (TCM) and effector memory T cell subsets (TEM) in TDLNs (Tumor draining lymph nodes) between WT/WGM mice and KO/WGM mice at the day 46 after tumor challenge. Expectedly, results showed that TDLNs harvested from KO/WGM mice exhibited higher subset ratios of CD44+CD62L+ (TCM) cell to CD4+ T cells as well as CD44+CD62L− or (TEM) cell to CD4+ T cells than those from WT/WGM mice. In the case of use of CD122 for memory marker, similar results were obtained. Of note, TDLNs from KO/WGM mice exhibited increased ratios of CD44+CD122+CD62L− (TSCM) cell. In addition, the cell numbers of immunosuppressive CD3+ CD4+ PD-1+, CD3+ CD4+ GITR+ and CD3+ CD4+ CTLA4+ cells were reduced in both TDLNs and spleen derived from KO/WGM mice compared with those from WT/WGM mice. Furthermore, our results of CBA (Cytometric Bead Array) assay using splenocytes harvested from day 2 to day 15 showed that the Th2 cytokine production level of IL-4 and IL-5 from splenocyte harvested from KO/WGM mice were higher than those from WT/WGM mice. In regard to IL-2 and IFN-g (Th1 cytokine), the production level at both day 7 and day10 from splenocytes harvested from KO/WGM mice were also higher than those from WT/WGM mice, implicating that loss of LTB4/BLT1 signaling promotes systemic activation of both tumor antigens specific Th1 and Th2 CD4+ T cell subsets. Finally, our flow cytometric analyses demonstrated that exogenously GM-CSF-driven upregulated MFI of CD40+, CD80+ and CD86+ DCs in TDLNs from KO/WGM mice were further significantly increased compared with those from WT/WGM mice (p<0.05), and more numerous number of CD86+ DCs that had phagocytosed TAAs of WEHI3B cells was detected in TDLNs harvested from KO/WGM mice than that from WT/WGM mice.In conclusion, we for the first time demonstrate that loss of LTB4/BLT1 axis can sustain long-lasting memory CD4+ T cell-dependent antitumor immunity against syngeneic tumor cells long after GM-CSF triggering tumor rejection, allowing us to expect the possibility that the strategy of blocking LTB4/BLT1signaling may be useful to maintain antitumor immunity induced by GM-CSF gene-transduced leukemia cell vaccine in clinical settings. Disclosures:No relevant conflicts of interest to declare.

Memory Type 2 Helper T Cells Induce Long-Lasting Antitumor Immunity by Activating Natural Killer Cells

Functionally polarized helper T cells (Th cells) play crucial roles in the induction of tumor immunity. There is considerable knowledge about the contributions of IFN-producing Th1 cells that supports the role of cytotoxic cluster of differentiation (CD8) T cells and natural killer (NK) cells, but much less is known about how IL-4-producing Th2 cells contribute to tumor immunity. In this study, we investigated the cellular and molecular mechanisms employed by memory Th2 cells in sustaining tumor immunity by using a mouse model system wherein ovalbumin (OVA) is used as a specific tumor antigen. In this model, we found that OVA-specific memory Th2 cells exerted potent and long-lasting antitumor effects against NK-sensitive OVA-expressing tumor cells, wherein antitumor effects were mediated by NK cells. Specifically, NK cell cytotoxic activity and expression of perforin and granzyme B were dramatically enhanced by the activation of memory Th2 cells. Interleukin 4 (IL-4) produced by memory Th2 cells in vivo was critical for the antitumor effects of the NK cells, which IL-4 directly stimulated to induce their perforin- and granzyme-B-dependent cytotoxic activity. Our findings show that memory Th2 cells can induce potent antitumor immunity through IL-4-induced activation of NK cells, suggesting potential applications in cellular therapy for cancer patients.

517 INTRAVESICAL LIPOSOME-MEDIATED INTERLEUKIN-15 GENE THERAPY ENHANCES TUMOR-SPECIFIC CYTOTOXICITY IN ORTHOTOPIC MURINE BLADDER CANCER MODEL

You have accessJournal of UrologyBladder Cancer: Basic Research1 Apr 2011517 INTRAVESICAL LIPOSOME-MEDIATED INTERLEUKIN-15 GENE THERAPY ENHANCES TUMOR-SPECIFIC CYTOTOXICITY IN ORTHOTOPIC MURINE BLADDER CANCER MODEL Kazuhiro Matsumoto, Eiji Kikuchi, Minoru Horinaga, Akira Miyajima, Ken Nakagawa, and Mototsugu Oya Kazuhiro MatsumotoKazuhiro Matsumoto Tokyo, Japan More articles by this author , Eiji KikuchiEiji Kikuchi Tokyo, Japan More articles by this author , Minoru HorinagaMinoru Horinaga Saitama, Japan More articles by this author , Akira MiyajimaAkira Miyajima Tokyo, Japan More articles by this author , Ken NakagawaKen Nakagawa Tokyo, Japan More articles by this author , and Mototsugu OyaMototsugu Oya Tokyo, Japan More articles by this author View All Author Informationhttps://doi.org/10.1016/j.juro.2011.02.1213AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookTwitterLinked InEmail INTRODUCTION AND OBJECTIVES Targeted cytokine therapies are required to improve BCG-related cytotoxic efficacy and reduce its side effects for the treatment of non-muscle invasive bladder cancer. Interleukin-15 (IL-15) is known to stimulate the proliferation of CD8+T-cells and NK cells, and also to maintain a memory CD8+T-cell, suggesting that it may be of value in cytokine treatment for bladder cancer. In the present study, we evaluated the therapeutic effect of in situ IL-15 gene therapy in an orthotopic bladder cancer model, and the long-lasting tumor-specific antitumor effect. METHODS We established an orthotopic bladder cancer model by implanting 5 x 105 MBT-2 cells into female C3H/HeN mice through the urethra. Intravesical instillation of lipoplex containing 5 μg of IL-15 gene plasmid was repeated 6 times at 2-day intervals starting from day 5 after tumor implantation, and we investigated its antitumor effect. Surviving mice treated with in situ IL-15 gene delivery were rechallenged with 5 x 105 MBT-2 cells in their right lower abdominal quadrant, and 1 X106 FM3A cells into the opposite side at the same time on day 50 to evaluate the tumor specific immunologic memory against MBT-2 cells. RESULTS On day 23, the bladder weights in the medium alone group, β-gal gene delivery control group, and IL-15 gene therapy group were 196 ± 36, 201 ± 35, and 96 ± 29 mg, respectively (P<0.05), demonstrating the anti-tumor effect of intravesical IL-15 gene therapy in this model. In the bladders treated with IL-15 gene plasmid instillation, histological analysis revealed many inflammatory cells were induced around the tumors. Immunohistochemical analysis confirmed that there was predominant infiltration of CD8+ T-cells around the tumor nest. In surviving mice after the intravesical IL-15 gene therapy, the growth of rechallenged subcutaneous MBT-2 cells was inhibited again, although newly implanted FM3A cells in the same mice were not rejected. Cytotoxic T lymphocyte (CTL) assay using CD8+ T cells isolated from their splenocytes also showed a tumor-specific CTL response against MBT-2 cells. CONCLUSIONS Our results showed that intravesical IL-15 plasmid transfection in situ exhibits significant tumor suppression in an orthotopic bladder cancer model and they had the potential to acquire tumor specific immunologic memory against MBT-2 cells. In conclusion, the present data indicate that IL-15 gene therapy may be a promising new adjuvant therapy for non-muscle invasive bladder cancer and a useful substitute for BCG treatment. © 2011 by American Urological Association Education and Research, Inc.FiguresReferencesRelatedDetails Volume 185Issue 4SApril 2011Page: e210 Advertisement Copyright & Permissions© 2011 by American Urological Association Education and Research, Inc.MetricsAuthor Information Kazuhiro Matsumoto Tokyo, Japan More articles by this author Eiji Kikuchi Tokyo, Japan More articles by this author Minoru Horinaga Saitama, Japan More articles by this author Akira Miyajima Tokyo, Japan More articles by this author Ken Nakagawa Tokyo, Japan More articles by this author Mototsugu Oya Tokyo, Japan More articles by this author Expand All Advertisement Advertisement PDF downloadLoading ...

Abstract 2441: An innovative treatment modality of antibody therapy against intracellular tumor antigens

Abstract Therapy with mAb directed against cell surface antigens of cancer is now a well-established treatment modality for human cancer. However, many tumor antigens of great interest have an intracellular location, and therefore not considered suitable targets for antibody therapy. On the other hand, it has been reported that mAbs targeted against specific tumor antigens induce cellular immune response including CD8+ T cells. This raises the possibility that the anti-tumor effects of mAb therapy may partially depend on enhanced cross-priming mediated by APCs following the uptake of antigen released from tumors complexed with Ab (immune-complex). Here, we investigated whether mAb therapy could be applicable to intracellular antigens using a transplantable colon carcinoma CT-26 expressing NY-ESO-1 (a well-studied human cancer/testis antigen), an intracellular antigen lacking cell surface expression. Immune responses to NY-ESO-1 in the murine model closely parallel NY-ESO-1 humoral and cellular immune responses in humans. To accentuate the natural release of intracellular NY-ESO-1 from dying tumor cells to facilitate immune-complex formation, we employed the effect the anti-cancer drug on the response to NY-ESO-1 mAb. The combination of NY-ESO-1 mAb and 5-fluorouracil exhibited a strong and long-lasting anti-tumor effect that could not be achieved by treatment with either 5-fluorouracil or mAb alone. The anti-tumor effect induced by combination therapy was no longer observed when Fc-depleted Fab antibodies were used, indicating a Fc-dependent action on APCs. Furthermore, augmented anti-tumor effects observed by the combination therapy was associated with strong NY-ESO-1-specific CD8+ T cells in draining lymph nodes and tumor sites, and upregulation of maturation markers on dendritic cells. The opsoniziation of released intracellular tumor antigens with injected mAb facilitates uptake by APC and presentation to CD8+ T cells, leading to accelerated and augmented active immunization. The data indicate that mAb to intracellular molecules of cancer represents a promising approach to cancer immunotherapy. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 2441.

Antitumor activity of anti-CTLA-4 monoclonal antibody (mAb) in combination with ixabepilone in preclinical tumor models

3048 Background: The antitumor activity of a homolog of ipilimumab, a CTLA-4 blocking agent, was investigated in combination with ixabepilone, the first in a new class of microtubule-stabilizing drugs. We hypothesized that this combinatorial approach may produce therapeutic synergy based on their unique mechanism of action and cellular targets. Ixabepilone induces tumor cell necrosis thus providing a source of tumor antigens and changes in tumor architecture that facilitate T-cell priming and infiltration, whereas CTLA-4 blocking mAb promotes expansion and infiltration of tumor-primed cytolytic T cells. Methods: Efficacy studies were conducted in 3 different tumor lines: SA1N fibrosarcoma, M109 lung carcinoma and EMT-6 mammary carcinoma. Ixabepilone and CTLA-4 mAb were administered at their optimal dose and schedule: ixabepilone, 8 mg/kg; CTLA-4 mAb, 20 mg/kg every 4 days for 3 doses. For the combination group, anti-CTLA-4 mAb was administered one day after each ixabepilone treatment. In selected studies, animals with complete tumor regressions were rechallenged with a lethal dose of tumor cells to determine the level of immune protection. In other studies, the combined efficacy of CTLA-4 mAb + ixabepilone was compared directly with CTLA-4 mAb + paclitaxel. Results: The combination of the ipilimumab homolog CTLA-4 mAb and ixabepilone showed a synergistic antitumor effect in all tumor models tested causing long-lasting complete responses in 70–100% of the animals, demonstrating superior efficacy compared to each treatment alone (p<0.05). Furthermore, animals treated with ixabepilone + CTLA-4 mAb rejected a subsequent tumor rechallenge suggesting the development of a protective memory immune response. Conversely, ixabepilone- treated animals showed only partial protection, as evident by a delay in tumor growth. Combination treatments were well-tolerated. The CTLA-4 mAb +ixabepilone combination yielded superior efficacy than CTLA-4+paclitaxel. Conclusions: These findings provide evidence that the combination of ixabepilone and ipilimumab homolog CTLA-4 blocking mAb elicits effective and long-lasting antitumor effects and warrant investigation of ipilimumab and ixabepilone regimen in clinical trials. Author Disclosure Employment or Leadership Consultant or Advisory Role Stock Ownership Honoraria Research Expert Testimony Other Remuneration Bristol-Myers Squibb Bristol-Myers Squibb

Dendritic cell therapy with interferon-α synergistically suppresses outgrowth of established tumors in a murine colorectal cancer model

Both dendritic cell (DC)-based immunotherapy and interferon (IFN)-alpha therapy have been proved to have potent long-lasting antitumor effects. In anticipation of synergistic antitumor effects, we performed combination therapy with DCs and IFN-alpha gene-transduced murine colorectal cancer MC38 cells (MC38-IFN-alpha). DCs incubated with MC38-IFN-alpha, but not neomycin-resistance gene-transduced MC38 cells (MC38-Neo), effectively enhanced proliferation of allogeneic splenocytes in vitro. In 12 of 17 mice, DCs in combination with MC38-IFN-alpha prevented the development of a parental tumor, while DCs and MC38-Neo did in only three of 17 mice (P=0.008). In a therapeutic model of an established parental tumor, inoculation of DCs and MC38-IFN-alpha suppressed the growth of the established parental tumors significantly compared with the administration of DCs with MC38-Neo or naive splenocytes with MC38-IFN-alpha (P=0.016 and 0.024, respectively). Analyses of immunohistochemistry and tumor-infiltrating mononuclear cells showed that CD8(+), CD11c(+), and NK1.1(+) cells markedly infiltrated the established tumors of mice treated with DCs and MC38-IFN-alpha. From the results of observation of parental tumor outgrowth in immune cell-depleted mice, CD8(+) cells, and asialo-GM-1(+) cells were thought to contribute to the antitumor effects induced by the combination therapy. Furthermore, MC38-specific cytolysis was detected when splenocytes of mice inoculated with DCs and MC38-IFN-alpha cells were stimulated with MC38-IFN-alpha cells in vitro. Since DC-based immunotherapy in combination with IFN-alpha-expressing tumor cells induces potent antitumor cellular immune responses, it should be considered for clinical application.

A possible mechanism for the long-lasting antitumor effect of the macromolecular conjugate DE-310: mediation by cellular uptake and drug release of its active camptothecin analog DX-8951

DE-310, a new macromolecular prodrug, was designed to enhance the pharmacological profiles of a novel camptothecin analog (DX-8951f), and a single treatment with DE-310 exhibits a similar or greater therapeutic effect than do optimally scheduled multiple administrations of DX-8951f in several types of tumors. In this study, the drug-release mechanism by which DE-310 excites antitumor activity was investigated in Meth A cells, a malignant ascites model of murine fibrosarcoma. A single i.v. injection of DE-310 at the maximum tolerated dose (MTD) prolonged survival of Meth A-bearing mice by 300%. DX-8951 and glycyl-8951 (G-DX-8951), enzymatic cleavage products of DE-310, were detected in serum and ascites fluid, and also in the culture medium of Meth A ascites cells incubated in vitro with DE-310. The total amounts of DX-8951, G-DX-8951, and conjugated DX-8951 in Meth A tumor cells were three times higher than that in macrophages. Furthermore, DX-8951-related fluorescence was observed in Meth A ascites cells obtained from Meth A-bearing mice that had received DE-310 or CM-Dex-PA-DX-8951 that does not release free DX-8951. DX-8951-related fluorescence was also observed at the site of lysosomes in cells incubated in vitro with DE-310 at 37 degrees C, but not in those incubated at 4 degrees C. Drugs were released from DE-310 by cysteine proteinase prepared from Meth A tumor tissue. These results suggest that the mechanism by which DX-8951 is released from DE-310 in vivo is involved in the process of uptake of DE-310 into tumor or macrophages, digestion by intracellular lysosomal cysteine proteinase, and subsequent secretion of the drugs.

Triptycenes

In contrast to their inactive parent compound triptycene (code name TT0), several triptycene (TT) analogs (code names TT1 to TT13), most of them new compounds, were synthesized and shown to prevent L1210 leukemic cells from synthesizing macromolecules and growing in vitro. The most potent rigid tetracyclic quinones synthesized so far are TT2 and its C2-brominated derivative, TT13. The antitumor activity of TT2 has been compared to that of daunomycin (DAU), a clinically valuable anthracycline antibiotic which is structurally different from TT2 but also contains a quinone moiety. TT2 inhibits the proliferation (IC50: 300 nM at day 2 and 150 nM at day 4) and viability (IC50: 250 nM at day 2 and 100 nM at day 4) of L1210 cells to the same maximal degree as DAU, suggesting that the cytostatic and cytotoxic activities of TT2 are a combination of drug concentration and duration of drug exposure. Since TT2 does not increase the mitotic index of L1210 cells at 24 h like vincristine, it is unlikely to be an antimitotic drug that disrupts microtubule dynamics. Like DAU, a 1.5-3 h pretreatment with TT2 is sufficient to inhibit the rates of DNA, RNA and protein syntheses determined over 30-60 min periods of pulse-labeling in L1210 cells in vitro (IC50: 6 microM). In contrast to DAU, which is inactive, a 15 min pretreatment with TT2 has the advantage of also inhibiting the cellular transport of nucleosides occuring over a 30 s period in vitro (IC50: 6 microM), suggesting that TT2 prevents the incorporation of [3H]thymidine into DNA because it rapidly blocks the uptake of [3H]thymidine by the tumor cells. After 24 h, TT2 induces as much DNA cleavage as camptothecin and DAU, two anti-cancer drugs producing DNA strand breaks and known to respectively inhibit DNA topoisomerase I and II activities. Interestingly, the abilities of TT2 to block nucleoside transport, inhibit DNA synthesis and induce DNA fragmentation are irreversible upon drug removal, suggesting that this compound may rapidly interact with various molecular targets in cell membranes and nuclei to disrupt the functions of nucleoside transporters and nucleic acids, and trigger long-lasting antitumor effects which persist after cessation of drug treatment. Because inhibition of nucleoside transport is highly unusual among DNA-damaging drugs, the use of bifunctional TTs with antileukemic activity in the nM range in vitro might provide a considerable advantage in polychemotherapy to potentiate the action of antimetabolites and sensitize multidrug-resistant tumor cells.