Tumor Lysate-pulsed Dendritic Cells Research Articles

Abstract 5029: Microglia differentiated from induced pluripotent stem cells treat glioma tumors in mice

Abstract Glioblastoma Multiforme (GBM), a grade IV astrocytoma is the most aggressive primary brain tumor. GBM patients have the worst prognoses with median overall survival of less than 15 months from the time of diagnosis. The standard of care consists of maximal surgical resection followed by chemotherapy, given concomitantly with and following radiotherapy. Despite these measures, disease progression or recurrence is virtually inevitable and occurs at a median time of approximately 7 months. Among the more promising experimental treatments for recurrent GBM is the “vaccination” of patients with dendritic cells to elicit anti-tumor immune responses. The safety of this approach has been well demonstrated, but the clinical response rate of patients has ranged from 8%-30%. To improve upon these responses, we investigated the ability of microglia; the resident immune cells of the CNS, to better activate cytotoxic T lymphocytes within the tumor microenvironment. Using a murine syngeneic intracranial tumor model, we found that combined vaccination with dendritic cells and microglia is more effective than either alone. Combination treated mice had sustained regression of tumors over a four-week imaging period and the median survival of the animals was greater than one year. In contrast, the median survivals of the various control groups were ≤ 52 days. Translation of our findings to the clinical setting requires a source of patient-specific microglia. Here, we report the sequential differentiation of human induced pluripotent stem cells (hiPSC) into hematopoietic progenitor-like cells, and then into cells with a phenotype and gene expression profile resembling that of primary brain-isolated microglia. Similarly, human iPSC-derived microglia (hiPS-MG) exhibit phagocytic activity; produce inflammatory cytokines and reactive oxygen species. We have also differentiated mouse induced pluripotent stem cells into microglia (miPS-MG) with gene expression profile and phenotype similar to primary neonatal microglia. Mice treated with a combination therapy of tumor lysate-pulsed miPS-MG and dendritic cells show median survival greater than one year, whereas the survival for the control group was less than 50 days. For direct clinical translation of our findings, the ability to readily differentiate hiPSC into microglia in vitro will allow for the generation of patient-specific cells for use in a GBM immunotherapy protocol. hiPSC-derived microglia may also be useful as modulatory cells for a variety of CNS diseases and serve as gene and protein delivery vehicles to the CNS. Acknowledgement: This research was supported by the Intramural Research Program of the NIH [NINDS]. Citation Format: Hetal Pandya, David M. Ichikawa, Michael J. Shen, Andrea B. Sedlock, Kory R. Johnson, Dragan Maric, Dorian B. McGavern, Harry L. Malech, John K. Park. Microglia differentiated from induced pluripotent stem cells treat glioma tumors in mice. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 5029. doi:10.1158/1538-7445.AM2015-5029

Dendritic cells combined with anti-GITR antibody produce antitumor effects in osteosarcoma.

We attempted to enhance the antitumor effects of tumor lysate-pulsed dendritic cells by eliminating regulatory T cells. The combinatorial effects of dendritic cells and agonist anti-glucocorticoid-induced tumor necrosis factor receptor (anti-GITR) antibodies were investigated with respect to enhancement of the systemic immune response, elimination of regulatory T cells, and inhibition of tumor growth. To determine whether the combination of dendritic cells and anti‑GITR antibodies could enhance systemic immune responses and inhibit primary tumor growth in a murine osteosarcoma (LM8) model. We established the following 4 groups of C3H mice (20 mice in total): i), control IgG-treated mice; ii), tumor lysate-pulsed dendritic cell‑treated mice; iii), agonist anti-GITR antibody-treated mice; and iv), agonist anti-GITR antibody- and tumor lysate-pulsed dendritic cell‑treated mice.The mice that received the agonist anti-GITR antibodies and tumor lysate-pulsed dendritic cells displayed inhibited primary growth, prolonged life time, reduced numbers of regulatory T lymphocytes in the spleen, elevated serum interferon-γ levels, increased number of CD8+ T lymphocytes. The mice that received combined therapy had reduced level of immunosuppressive cytokines in tumor tissue and serum. Combining agonist anti-GITR antibodies with tumor lysate-pulsed dendritic cells enhanced the systemic immune response. These findings provide further support for the continued development of agonist anti-GITR antibodies as an immunotherapeutic strategy for osteosarcoma. We suggest that our proposed immunotherapy could be developed further to improve osteosarcoma treatment.

Cytokine induced killer cell-based immunotherapies in patients with different stages of renal cell carcinoma

Cytokine induced killer (CIK) cell-based treatments have shown antitumor activity against renal cell carcinoma (RCC) in vitro and in vivo. But the therapeutic options and benefits of various CIK cells were unknown for different stages of RCC. In this random clinical trial, the 3-year disease free survival (DFS) of operable RCC patients treated with autologous tumor lysate-pulsed dendritic cells co-cultured with cytokine induced killer (Ag-DC-CIK) was 96.7% compared with 57.7% in the control group. Ag-DC-CIK immunotherapy decreased the risk of post-operative disease progression and relapse (P = 0.0418). In inoperable RCC patients treated with CIK, the 3-year overall survival (OS) and progression-free survival (PFS) were significantly longer than the control group (P = 0.0116 and P = 0.0212). The CD4+/CD8+ T cell ratio in peripheral blood increased after the last cell infusion in the CIK treatment group, and especially further increased in the Ag-DC-CIK treatment group (P = 0.002). No severe toxicity was observed after infusion of CIK cells. Therefore, tumor antigen-sensitized Ag-DC-CIK cells might be more efficient and personalized for the patients with tumor resection, and CIK cells could improve the prognosis for those inoperable patients. According to the stages of RCC patients, different CIK cell-based immunotherapies would help to achieve more beneficial effects.

P0241 : CD40/CD40Ligand interaction between tumor lysate-pulsed dendritic cells induces a Th1-environment and enhances the cytotoxic activity of immunologic effector cells towards human hepatocellular carcinoma and cholangiocellular carcinoma cells

P0241 : CD40/CD40Ligand interaction between tumor lysate-pulsed dendritic cells induces a Th1-environment and enhances the cytotoxic activity of immunologic effector cells towards human hepatocellular carcinoma and cholangiocellular carcinoma cells

ITOC2 – 017. Prolonged survival for patients with recurrent glioblastoma multiforme who are treated with tumour lysate-pulsed autologous dendritic cells

<h3>Background</h3> Recurrent glioblastoma multiforme (rGBM) is a life threatening condition, with a mortality rate of approximately 100%. Despite recent advances in therapy, the survival rate in rGBM patients has not meaningfully changed in the past several decades. We have treated two cohorts of rGBM patients, one consisting of patients with early progression, and one consisting of patients with progression following several cycles of adjuvant temozomolide chemotherapy, with autologous dendritic cells pulsed with autologous tumour cell lysate (DCVax®-L). Such treatment is intended to activate the immune system against the tumour cells, with the expectation that the ensuing immune attack may delay progression and time to death. <h3>Aim</h3> To determine survival characteristics of patients with rGBM who are treated with DCVax-L. <h3>Methods</h3> Disease progression in patients with GBM was determined through magnetic resonance imaging. Progression needed to be determined by independent review on two consecutive scans for early progressor classification to avoid enrolling patients with pseudoprogression. Assessment of progression was done using modified RANO criteria. <h3>Results</h3> Nineteen (19) patients diagnosed with GBM were determined to have rapid recurrence following radiation therapy with concomitant temozolomide chemotherapy. The life expectancy for such patients is typically poor, and they were provided the DCVax-L treatment as part of a compassionate use approach in parallel to ongoing late stage clinical trials. Median overall survival in this cohort from initial GBM diagnosis is 15.1months (95% confidence interval (CI): 10.5–17.2), and the range is 8.1–>31months. A literature search revealed six publications with comparable populations of patients (Brandes et al., 2008; Roldan et al., 2009; Sanghera et al., 2010; Kang et al., 2010; Gunjur et al., 2011; Linhares et al., 2013). The table below demonstrates that these patients typically have a life expectancy of 8–10months. Reference (<i>n</i>) Population Median Survival (95% CI) DCVax-L (19) post RT+chemo, confirmed 2months later 15.1months (10.5–17.2) Brandes et al. (2008) (18) PD at 4weeks pos RT+chemo, confirmed after two more tmz cycles 10.2 months (n.a.) Roldan et al. (2009) (10) PD at 4–6weeks post RT+chemo, confirmed after <mml:math><mml:mrow><mml:mo>⩾</mml:mo></mml:mrow></mml:math>1 more tmz cycle(s) 9.1months (4.9–19.1) Kang et al. (2010) (10) PD at two consecutive scans post RT+chemo 10.8 (n.a.) Sanghera et al. (2010) (29) PD at two consecutive scans within 8weeks post RT+chemo8.3months (n.a.) Gunjur et al. (2011 (27) PD at two consecutive scans within 3months post RT+chemo, or clinical deterioration 10.4months (n.a.) Linhares et al. (2013) (13) PD at two consecutive scans within 3months post RT+chemo 9.0 months (3.7–14.3) A second cohort of patients with rGBM consisted of eight patients with recurrence following several adjuvant temozolomide treatment cycles. Median overall survival (OS) for these patients is 14.7months from the time of surgery for first recurrence. The comparator for this cohort is derived from Friedman et al. (2009), who reported mOS of 8.7–9.3months from the time of first recurrence for rGBM patients treated with Bevacizumab with or without irinotecan. <h3>Results and conclusion</h3> The results obtained in two cohorts of patients with rGBM suggest that treatment with autologous DC pulsed with autologous tumour cell lysate can extend survival by 5months or more.

Anti-CD27 Antibody Potentiates Antitumor Effect of Dendritic Cell-Based Vaccine in Prostate Cancer-Bearing Mice.

In the current study, we investigated whether anti-CD27 monoclonal antibody can enhance the antitumor efficacy of a dendritic cell–based vaccine in prostate cancer–bearing mice. The overall therapeutic effect of a dendritic cell–based vaccine for prostate cancer remains moderate. A prostate cancer model was established by subcutaneous injection of RM-1 tumor cells into male C57BL/6 mice on day 0. After 4 days, tumor-bearing mice were treated with RM-1 tumor lysate–pulsed dendritic cells (i.e., dendritic cell–based vaccine), anti-CD27 monoclonal antibody, or a combination of RM-1 tumor lysate–pulsed dendritic cells with anti-CD27 monoclonal antibody. Mice were killed at 21 days after tumor cell implantation. Tumor size was measured for assessment of antitumor effect. Spleens were collected for analysis of antitumor immune responses. The antitumor immune responses were evaluated by measuring the proliferation and activity of T cells, which have the ability to kill tumor cells. The combination therapy with RM-1 tumor lysate–pulsed dendritic cells and anti-CD27 antibody significantly enhanced T-cell proliferation and activity, and significantly reduced tumor growth, compared with monotherapy with RM-1 tumor lysate–pulsed dendritic cells or anti-CD27 antibody. Our results suggest that combined treatment can strengthen antitumor efficacy by improving T-cell proliferation and activity.

Mgn-3/biobran enhances generation of cytotoxic CD8+ T cells via upregulation of dec-205 expression on dendritic cells.

Arabinoxylan rice bran (MGN-3/Biobran) has been shown to be a potent biological response modifier (BRM) that activates different arms of the immune system, including dendritic cells (DCs), which prime CD4+ helper T-cell responses. The present study explores the ability of MGN-3-activated DCs to prime CD8+ T cells and examines the mechanisms underlying its effect. Human monocyte-derived DCs were treated with MGN-3 (20 and 40 &#956;g/ml). Results indicate that treatment with MGN-3 caused DCs to prime higher granzyme B-expressing CD8+ T cells. Tumor lysate-pulsed MGN-3 DC also increased tumor cell killing compared to DC-stimulated CD8+ T cells. This was associated with: i) increased expression of DEC-205 in MGN-3-activated DCs in a dose-dependent manner; and ii) MGN-3 induced significant production of Type III interferon, IL29, but not Type I IFNs &#945; and &#946;. These results suggest that MGN-3 is a potent natural adjuvant that efficiently activates DCs and may therefore be useful for mounting an efficient immune response against infections and cancer.

Chitosan as an adjuvant-like substrate for dendritic cell culture to enhance antitumor effects

To induce monocyte differentiation into dendritic cells (DCs) is the essential protocol for the DC-mediated cancer immunotherapy. In this study, monocytes isolated from mouse bone marrow were cultured on chitosan substrate to evaluate the effect of the chitosan culture system on the induction and tumor protection of DCs. Compared to tissue culture polystyrene (TCPS), the chitosan culture system could enhance monocyte aggregation and detachment with increased MTT reduction activity and expression of DC marker CD11c and LPS co-receptor CD14. Moreover, compared to TCPS, chitosan could enhance lipopolysaccharides (LPS)-stimulated DCs to secrete higher amount of IL-12. More importantly, vaccination of tumor lysate-pulsed DCs harvested from chitosan could increase cytotoxic T-lymphocyte (CTL) activity and showed significantly enhanced anti-tumor effect than those from TCPS. Therefore, the current study demonstrated that a protocol to culture DCs on a less-adherent chitosan substrate followed by treatment with tumor lysate has the potential in future DC-based vaccine application.

Autologous Tumor Lysate-Pulsed Dendritic Cell Immunotherapy with Cytokine-Induced Killer Cells Improves Survival in Gastric and Colorectal Cancer Patients

Gastric and colorectal cancers (GC and CRC) have poor prognosis and are resistant to chemo- and/or radiotherapy. In the present study, the prophylactic effects of dendritic cell (DC) vaccination are evaluated on disease progression and clinical benefits in a group of 54 GC and CRC patients treated with DC immunotherapy combined with cytokine-induced killer (CIK) cells after surgery with or without chemo-radiotherapy. DCs were prepared from the mononuclear cells isolated from patients using IL-2/GM-CSF and loaded with tumor antigens; CIK cells were prepared by incubating peripheral blood lymphocytes with IL-2, IFN-γ, and CD3 antibodies. The DC/CIK therapy started 3 days after low-dose chemotherapy and was repeated 3–5 times in 2 weeks as one cycle with a total of 188.3±79.8×106 DCs and 58.8±22.3×108 CIK cells. Cytokine levels in patients' sera before and after treatments were measured and the follow-up was conducted for 98 months to determine disease-free survival (DFS) and overall survival (OS). The results demonstrate that all cytokines tested were elevated with significantly higher levels of IFN-γ and IL-12 in both GC and CRC cohorts of DC/CIK treated patients. By Cox regression analysis, DC/CIK therapy reduced the risk of post-operative disease progression (p<0.01) with an increased OS (<0.01). These results demonstrate that in addition to chemo- and/or radiotherapy, DC/CIK immunotherapy is a potential effective approach in the control of tumor growth for post-operative GC and CRC patients.

Postoperative dendritic cell vaccine plus activated T-cell transfer improves the survival of patients with invasive hepatocellular carcinoma

The recurrence rate after surgery in patients with hepatocellular carcinoma (HCC) is very high, while prognosis is quite poor. However, there is no standard treatment to prevent recurrence of HCC after a curative operation. In this study, we investigated the clinical utilization of an autologous tumor lysate-pulsed dendritic cell vaccine plus ex vivo activated T cell transfer (ATVAC) in an adjuvant setting for postoperative HCC as a non-randomized controlled trial. Ninety-four patients with invasive HCC received informed consent information regarding the study, and 42 opted to have the ATVAC after surgery. Their recurrence-free survival (RFS) and overall survival (OS) were measured after 5 years and compared with those of 52 patients who selected to have the curative operation alone. The median RFS and OS were 24.5 months and 97.7 months in the patients receiving adjuvant ATVAC and 12.6 months and 41.0 months in the group receiving surgery alone (P = 0.011 and 0.029). In the treated group, patients with positive delayed-type hypersensitivity (DTH) had a better prognosis (RFS P = 0.019, OS P = 0.025). No adverse events of grade 3 or more were observed.A postoperative dendritic cell vaccine plus activated T cell transfer would be a feasible and effective treatment for preventing recurrence in HCC patients and achieving long-term survival especially in DTH positive patients.

Identification of pancreatic cancer-associated tumor antigen from HSP-enriched tumor lysate-pulsed human dendritic cells.

PurposeVaccine strategies utilizing dendritic cells (DCs) to elicit anti-tumor immunity are the subject of intense research. Although we have shown that DCs pulsed with heat-treated tumor lysate (HTL) induced more potent anti-tumor immunity than DCs pulsed with conventional tumor lysate (TL), the underlying molecular mechanism is unclear. In order to explore the molecular basis of this approach and to identify potential antigenic peptides from pancreatic cancer, we analyzed and compared the major histocompatibility complex (MHC) ligands derived from TL- and HTL-pulsed dendritic cells by mass spectrophotometry.Materials and MethodsHuman monocyte-derived dendritic cells were pulsed with TL or HTL prior to maturation induction. To delineate differences of MHC-bound peptide repertoire eluted from DCs pulsed with TL or HTL, nanoflow liquid chromatography-electrospray ionization-tandem mass spectrometry (nLC-ESI-MS-MS) was employed.ResultsHTL, but not TL, significantly induced DC function, assessed by phenotypic maturation, allostimulation capacity and IFN-γ secretion by stimulated allogeneic T cells. DCs pulsed with TL or HTL displayed pancreas or pancreatic cancer-related peptides in context of MHC class I and II molecules. Some of the identified peptides had not been previously reported as expressed in pancreatic cancer or cancer of other tissue types.ConclusionOur partial lists of MHC-associated peptides revealed the differences between peptide profiles eluted from HTL-and TL-loaded DCs, implying that induced heat shock proteins in HTL chaperone tumor-derived peptides enhanced their delivery to DCs and promoted cross-presentation by DC. These findings may aid in identifying novel tumor antigens or biomarkers and in designing future vaccination strategies.

CD137 stimulation and p38 MAPK inhibition improve reactivity in an in vitro model of glioblastoma immunotherapy.

Dendritic cell vaccination has become an interesting option for cancer immunotherapy. Tumor-lysate-pulsed dendritic cells (DC) can prime naïve T cells and induce the regression of established tumors including gliomas as shown in various animal models. Despite hopeful results even in clinical studies, the outcome for many patients is still unsatisfying. In the present study, we tested the combination of tumor-lysate-pulsed dendritic cells (TPDC) with a monoclonal antibody against CD137, a monoclonal antibody against CD25 (daclizumab) and a specific p38 mitogen-activated protein kinase (p38 MAPK) inhibitor (SB203580) for improving immunostimulation in an in vitro model of immunotherapy for human gliomas. We observed a higher secretion of interferon gamma by TPDC-primed peripheral blood mononuclear cells (PBMC) that were incubated with an antibody against CD137 or the p38 MAPK inhibitor. In addition, we observed higher specific lysis of tumor cells after incubation of PBMC with the p38 MAPK inhibitor or the anti-CD137 antibody. In contrast, incubation of TPDC-primed PBMC with the anti-CD25 antibody did enhance neither interferon gamma secretion nor cellular cytotoxicity. Cell depletion experiments demonstrated that the immune reaction induced by TPDC is strongly dependent on CD4-positive and CD8-positive cells. Incubation of DC during maturation and antigen loading with the anti-CD137 antibody did not enhance cytotoxicity and interferon gamma secretion in comparison with application of the anti-CD137 antibody during priming. In conclusion, our data suggest that p38 MAPK inhibition and anti-CD137 antibodies can enhance the immune response against glioblastoma cells.

Immunotherapy for High-Grade Glioma: How to Go Beyond Phase I/II Clinical Trials

Evaluation of: Lasky JL 3rd, Panosyan EH, Plant A et al. Autologous tumor lysate-pulsed dendritic cell immunotherapy for pediatric patients with newly diagnosed or recurrent high-grade gliomas. Anticancer Res. 33, 2047-2056 (2013). Immunotherapy for children and adults with high-grade glioma (HGG) is an emerging innovative treatment approach, which aims at stimulating the body's own immune system against HGG by using autologous dendritic cells pulsed with autologous tumor lysate as a therapeutic vaccine. This is the third report on immunotherapy for HGG in children, bringing additional knowledge and experience to the scientific community. However, at the same time, this and other manuscripts urge for the next step in treatment development.

Improving immunotherapy of hepatocellular carcinoma (HCC) using dendritic cells (DC) engineered to express IL‐12 in vivo

Interleukin 12 (IL-12), one of the most potent Th1-cytokines, has been used to improve dendritic cells (DC)-based immunotherapy of cancer. However, it failed to achieve clinical response in patients with hepatocellular carcinoma (HCC). In this study, improved conditions of immunotherapy with DC engineered to express IL-12 were studied in murine subcutaneous HCC. Tumour-lysate pulsed DC were transduced with IL-12-encoding adenoviruses or cultivated with recombinant (r)IL-12. DC were injected intratumourally, subcutaneously or intravenously at different stages of tumour-development. Dendritic cell overexpressing IL-12 by adenoviruses showed enhanced expression of costimulatory molecules and stronger priming of HCC-specific effector cells than DC cultured with rIL-12. Intratumoural but not systemic injections of IL-12-DC induced the strongest antitumoural effects reaching complete regressions in 75% of early-staged tumours and in 33% of advanced tumours. Importantly, antitumoural effects could be further enhanced through combination with sorafenib. Analysing the tumour-environment, IL-12-DC increased the levels of Th1-cytokines/chemokines and of CD4(+) -, CD8(+) -T- and NK-cells. Induced immunity was tumour-specific and sustained since all tumour-free animals were protected towards hepatic tumour-cell rechallenge. However, IL-12-DC also enhanced immunosuppressive cytokines, regulatory T cells and even myeloid-derived suppressor cells within the tumours. Induced IL-12-overexpression by adenoviral vectors can effectively immunostimulate DC. Intratumoural but not systemic injection of activated IL-12-DC was crucial for effective tumour regression. The mechanism of this approach seems to be the induction of a sufficient Th1 tumour-environment allowing the recruitment of effector cells rather than the inhibition of tumour immunosuppression. Thus, improved immunotherapy with IL-12-DC represents a promising approach towards HCC.

Examination of MARCO Activity on Dendritic Cell Phenotype and Function Using a Gene Knockout Mouse

We have reported the upregulation of MARCO, a member of the class A scavenger receptor family, on the surface of murine and human dendritic cells (DCs) pulsed with tumor lysates. Exposure of murine tumor lysate-pulsed DCs to an anti-MARCO antibody led to loss of dendritic-like processes and enhanced migratory capacity. In this study, we have further examined the biological and therapeutic implications of MARCO expression by DCs. DCs generated from the bone marrow (bm) of MARCO knockout (MARCO-/-) mice were phenotypically similar to DCs generated from the bm of wild-type mice and produced normal levels of IL-12 and TNF-α when exposed to LPS. MARCO-/- DCs demonstrated enhanced migratory capacity in response to CCL-21 in vitro. After subcutaneous injection into mice, MARCO-/- TP-DCs migrated more efficiently to the draining lymph node leading to enhanced generation of tumor-specific IFN-γ producing T cells and improved tumor regression and survival in B16 melanoma-bearing mice. These results support targeting MARCO on the surface of DCs to improve trafficking and induction of anti-tumor immunity.

Abstract LB-321: Combination dendritic cell and conventional chemoradiation therapy improves survival for rectal cancer in a murine model.

Abstract The goal of neoadjuvant therapy in rectal cancer is to downstage or preferably eradicate the tumor prior to resection. Achieving complete pathologic response (CPR) using neoadjuvant chemoradiation is associated with improved overall and disease free survival. The aim of this study was to achieve improved complete pathologic response rates and survival by combining intra-tumoral injection of dendritic cells with conventional therapy. A syngeneic rectal cancer cell line (CT26) was subcutaneously injected into the thigh of immunocompetent Balb/c mice. Mice either received no treatment, conventional chemoradiation (CXRT) (5-fluorouracil + XRT) for 3 weeks, or CXRT plus tumor-lysate pulsed dendritic cells (CXRT/DC). Specific anti-CT26 cytotoxic T cell (CTL) activity was determined using standard chromium release assay. Flow cytometry for immune cell subsets was performed on cells isolated from spleens and tumors. Animals in the control group showed predictable uninhibited growth and all were euthanized due to tumor size. CXRT animals showed improved survival but none achieved complete response. Thirty three percent of the animals in CXRT/DC group exhibited CPR. Median survival for the control, CXRT, and CXRT/DC treated animals was 24, 46.5, 67 days, (p&amp;lt;0.0001). Splenocytes from CXRT/DC treated mice exhibited higher specific anti-CT26 CTL activity (28%) when compared to CXRT (17%), untreated control (11%) and naïve mice (&amp;lt;1%). Flow cytometry demonstrated an increase in regulatory T cells (Tregs), macrophages, granulocytic myeloid derived suppressor cells (MDSCs) and monocytic MDSCs in tumor bearing animals, (p&amp;lt;0.05). Dendritic cell activity stimulates anti-tumor CTL responses, but may also decrease MDSC and Treg mediated immunosuppression. Addition of dendritic cells to conventional chemoradiation therapy for rectal cancer increases complete pathologic response and thereby survival via immune modulation. Further analysis will allow subset comparisons between treatment groups. Citation Format: Swapnil D. Kachare, Jennifer M. Cayer, F. Melinda Carver, Zachary P. Englert, Kathryn M. Verbanac, Emmanuel E. Zervos, Nasreen A. Vohra. Combination dendritic cell and conventional chemoradiation therapy improves survival for rectal cancer in a murine model. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr LB-321. doi:10.1158/1538-7445.AM2013-LB-321

175 ENHANCING DENDRITIC CELL VACCINES WITH LIGANDS FOR DECTIN-1 AND TOLL-LIKE RECEPTOR-9

You have accessJournal of UrologyKidney Cancer: Basic Research (I)1 Apr 2013175 ENHANCING DENDRITIC CELL VACCINES WITH LIGANDS FOR DECTIN-1 AND TOLL-LIKE RECEPTOR-9 Yanping Wang, Nargess Hassanzadeh-Kiabi, Helen Goodridge, David Underhill, and Hyung Kim Yanping WangYanping Wang Los Angeles, CA More articles by this author , Nargess Hassanzadeh-KiabiNargess Hassanzadeh-Kiabi Los Angeles, CA More articles by this author , Helen GoodridgeHelen Goodridge Los Angeles, CA More articles by this author , David UnderhillDavid Underhill Los Angeles, CA More articles by this author , and Hyung KimHyung Kim Los Angeles, CA More articles by this author View All Author Informationhttps://doi.org/10.1016/j.juro.2013.02.1555AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookTwitterLinked InEmail INTRODUCTION AND OBJECTIVES Cancer vaccines have the potential to target tumors while sparing normal tissue. Dectin-1 receptor plays an important role in activating a cellular immune response against fungal infection, providing a rational for evaluating Dectin-1 ligand as an adjuvant for cancer vaccines. Activation of toll-like receptors (TLRs), particularly TLR-3, 4 and 9, has been reported to produce robust antitumor immune responses. Therefore, Dectin stimulation was evaluated alone or in combination with TLR stimulate. METHODS To assess CD8 T cell responses, B16 lysate-pulsed DCs were co-cultured with lymphocytes from pmel-1 mice. Pmel-1 transgenic mice express CD8 T cells with specificity for a Db-restricted epitope from the melanoma tumor antigen gp10025-33. The effect of stimulating DCs with ligands for Dectin-1, TLR, or both was assessed by monitoring pmel-1 lymphocyte proliferation and activation. Antitumor activity was assessed in vivo using syngeneic mouse tumor models for melanoma and renal cell carcinoma. RESULTS Treating DCs with Dectin-1 ligand (β-glucan) alone did not increase CD8 T cell proliferation or IFN-ψ production. Furthermore, there was no significant effect on CD8 T cells when DCs were treated with β-glucan combined with TLR4 ligand (lipopolysaccharide) or TLR3 ligand (poly I:C). However, the combination of β-glucan and TLR9 ligand (unmethylated CpG oligonucleotides, CpG) increased CD8 lymphocyte proliferation significantly more than CpG or β-glucan alone. In fact, the increase in CD8 proliferation was greater than with β-glucan plus all 3 TLR ligands. The combination of β-glucan and CpG was significantly more effective than β-glucan alone in stimulating IFN-ψ production. Based on these results, the combination of β-glucan and CpG was evaluated for enhancing DC vaccines in vivo. Bone marrow derived DCs were pulsed with tumor lysate and stimulated with β-glucan, CpG or both, prior to injection in mice. Following two DC vaccine treatments, mice were subcutaneously challenged with B16 tumors. A tumor prevention model was used to unequivocally demonstrate that antitumor effects were immune mediated with no possibility of direct cytotoxic effect by β-glucan or CpG. None of the mice treated with β-glucan plus CpG grew tumor while all control mice rapidly grew tumor. These findings were replicated using the in vivo RENCA model for renal cell carcinoma. CONCLUSIONS Preclinical data suggests that tumor lysate-pulsed DC vaccines can be enhanced by stimulation of Dectin-1 and TLR-9. © 2013 by American Urological Association Education and Research, Inc.FiguresReferencesRelatedDetails Volume 189Issue 4SApril 2013Page: e72 Advertisement Copyright & Permissions© 2013 by American Urological Association Education and Research, Inc.MetricsAuthor Information Yanping Wang Los Angeles, CA More articles by this author Nargess Hassanzadeh-Kiabi Los Angeles, CA More articles by this author Helen Goodridge Los Angeles, CA More articles by this author David Underhill Los Angeles, CA More articles by this author Hyung Kim Los Angeles, CA More articles by this author Expand All Advertisement Advertisement PDF downloadLoading ...

Development and validation of a fully GMP-compliant production process of autologous, tumor-lysate pulsed dendritic cells

One of the major challenges of DC vaccination is the recruitment of further cell therapy facilities and the establishment of validated and harmonized DC-production protocols in order to conduct larger, randomized clinical trials. Here, we report about the transfer and validation of a former successfully used open DC-generation method into a closed-system, GMP-compatible protocol.

Enhancement of antitumor immunity by combining anti-cytotoxic T lymphocyte antigen-4 antibodies and cryotreated tumor lysate-pulsed dendritic cells in murine osteosarcoma

Immunotherapy with tumor lysate-loaded dendritic cells (DCs) is one of the most promising strategies to induce antitumor immune responses. However, the antitumor activity of cytotoxic Tcells may be restrained by their expression of the inhibitory T-cell coreceptor cytotoxic Tlymphocyte antigen-4 (CTLA-4). By relieving this restraint, CTLA-4-blocking antibodies promote tumor rejection, but the full scope of their most suitable applications has yet to be fully determined. In the present study, we offer proof of a preclinical concept in a C3Hmouse osteosarcoma (LM8) model that CTLA-4 blockade cooperates with cryotreated tumor lysate-pulsed DCs in a primary tumor to prevent the outgrowth of lung metastasis. To evaluate immune response activation, we established the following four groups of C3Hmice (60mice in total): i)control immunoglobulinG (IgG)-treated mice; ii)tumor lysate-pulsed DC-treated mice; iii)anti-CTLA-4 antibody-treated mice and iv)tumor lysate-pulsed DC- and anti-CTLA-4 antibody-treated mice. The mice that received the tumor lysate-pulsed DCs and anti-CTLA-4 antibody displayed reduced numbers of regulatory Tlymphocytes and increased numbers of CD8+Tlymphocytes inside the metastatic tumor, inhibition of metastatic growth, a prolonged lifetime, reduced numbers of regulatory Tlymphocytes in the spleen and high serum interferon-γ levels. Combining an anti-CTLA-4 antibody with tumor lysate-pulsed DCs enhanced the systemic immune response. To the best of our knowledge, these findings document for the first time an effect of the combination of tumor lysate-pulsed DCs and CTLA-4-blocking antibodies in osteosarcoma. We suggest that cryotreated tumor lysate-pulsed DCs, although insufficient on their own, may mediate the rejection of metastatic lesions and prevent recurrence of the disease when combined with CTLA-4 blockade in osteosarcoma patients in the clinical setting.

Addition of tumor-lysate pulsed dendritic cells to chemoradiation increases complete pathologic response and imparts lasting immunity in a murine model of rectal cancer

Addition of tumor-lysate pulsed dendritic cells to chemoradiation increases complete pathologic response and imparts lasting immunity in a murine model of rectal cancer