bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract C95: Correlated intrinsic breast cancer subtypes and expression of CD47 in BM predicts recurrence
Authors: Nagahara Makoto , Sugihara Kenichi , Mori Masaki
Published: August 2014

Purpose: The intrinsic breast cancer subtypes have shown the prognostic features. Focused on CD47 expression in bone marrow (BM), we previously demonstrated that CD47 is an important biomarker in breast cancer, and functions as a prognostic factor for DFS. The purpose of this study is to identify the correlation between breast cancer subtypes and CD47 expression in BM, which may indicate clinical implications.

Methods: Quantitative real-time PCR was used to evaluate CD47 mRNA expression in bone marrow (BM) and in peripheral blood (PB) from 738 cases of breast cancer. ER, PR, and Her2 scores were obtained from immunohistochemistry (IHC) staining, and Her2 FISH analysis conducted for IHC 2+.

Results: According to ER, PR, and Her2 statuses, the groups of Her2 enriched (ER- and PR- and Her2 3+) and triple negative(ER- and PR- and Her2-) were divided (n=38 and 72). The 5 years disease free survival rates in each subgroup were as followed: p=0.0004; ER+ and PR+ and Her2- 0.94, ER+ and/or PR+ or- and/or Her2+ or- 0.16, Her2 enriched 0.23, triple negative 0.34). In patients with high levels of CD47 expression in the BM, survival was significantly poorer compared to patients with low levels of CD47 expression (DFS of Her2 enriched group: P=0.003, DFS of triple negative group: P=0.002). Furthermore, high CD47 expression group in a multivariate analysis showed significance as an independent variable for poorer prognosis in DFS (P = 0.02). In the PB, however, high CD47 expression in patients was not an independent and significant prognostic factor for DFS and OS in a multivariate analysis.

Conclusions: We confirmed that overexpression of CD47 in BM correlated with the aggressiveness of Her2 enriched and triple negative subgroups. CD47 is a useful prognostic biomarker for predicting survival of Her2 enriched and triple negative subgroups in breast cancer. We suggest that high CD47 expression in BM may represent tumor dormancy in breast cancer involved in postsurgical recurrence and metastasis of Her2 enriched and triple negative subtypes.

Citation Format: Makoto Nagahara, Kenichi Sugihara, Masaki Mori. Correlated intrinsic breast cancer subtypes and expression of CD47 in BM predicts recurrence. [abstract]. In: Proceedings of the AACR Special Conference on Tumor Invasion and Metastasis; Jan 20-23, 2013; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2013;73(3 Suppl):Abstract nr C95.
Abstract IA13: Niches and pathways of latent and overt metastasis
Authors: Massagué Joan
Published: August 2014

The signals and niches that allow disseminated tumor cells (DTCs) to stay viable as metastasis seeds are largely unknown. An understanding of these mechanisms is needed for the development of treatments to eradicate DTCs and prevent metastasis. We have identified molecular mechanisms that prime breast cancer DTCs for survival and stem cell fitness in distant organs. For survival, SRC primes DTCs for a robust PI3K-Akt response to CXCL12 and IGF1 in the bone marrow, and the leukocyte-tethering receptor VCAM-1 primes DTCs for PI3K-Akt activation by leukocyte contacts in the lungs. For stem cell fitness, the extracellular matrix protein tenascin-C (TNC) forms micrometastatic niches that promote WNT and NOTCH signaling. Additionally, a CXCL1-S100A8 paracrine loop protects DTCs from the stresses of metastasis and chemotherapy, providing a basis for these two clinically linked phenomena. In some cases, the pathways driving carcinoma formation additionally mediate metastasis. VHL inactivation leads to HIF signaling as a tumor-initiating event in clear cell renal carcinoma (ccRCC). However, VHL mutation status in ccRCC is not correlated with clinical outcome. We found that during ccRCC progression, cancer cells exploit diverse epigenetic alterations to empower a branch of the VHL-HIF pathway for metastasis through CXCR4 and CYTIP. The strength of this activation is associated with poor clinical outcome. Thus, metastasis can be driven by an epigenetically expanded output of the tumor-initiating pathway as well as by events that are mechanistically uncoupled from the tumor-initiating pathway. In preclinical models, SRC kinase inhibitors, VCAM-1 blocking antibodies, and CXCL1 receptor inhibitors suppress metastasis and augment the efficacy of chemotherapy, providing new avenues for targeting DTCs in the adjuvant setting.

Citation Format: Joan Massagué. Niches and pathways of latent and overt metastasis. [abstract]. In: Proceedings of the AACR Special Conference on Tumor Invasion and Metastasis; Jan 20-23, 2013; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2013;73(3 Suppl):Abstract nr IA13.
Abstract A17: Cancer immunotherapy using naked DNA vaccine targeting p53.
Authors: Soong Rueyshyang , Hung Chien-Fu , Wu T.-C.
Published: August 2014

Background: The pivotal role of p53 as a tumor suppressor protein is illustrated by the fact that this protein is found mutated in more than 50% of human cancers. In most cases, mutations in p53 greatly increase the otherwise short half-life of this protein in normal tissue and cause it to accumulate in the cytoplasm of tumors. The overexpression of mutated p53 in tumor cells makes p53 a potentially desirable target for the development of cancer immunotherapy. However, p53 protein represents an endogenous tumor-associated antigen (TAA), and immunization against self-antigen is challenging because antigen-specific immune response likely only generate low affinity antigen-specific CD8+ T cells. This represents a bottleneck of tumor immunotherapy when targeting endogenous antigens expressed by tumors.

Purpose: The objective of the current study is to develop a safe cancer immunotherapy using a naked DNA vaccine employing xenogenic p53 gene to break immune tolerance resulting in a potent therapeutic antitumor effect against tumors expressing mutated p53. Our study assessed the therapeutic antitumor effect after immunization with DNA encoding human p53 (hp53) or mouse p53 (mp53).

Results: C57BL/6 mice immunized with xenogenic full length of hp53 DNA plasmid intramuscularly followed by electroporation were capable of protecting against challenge with murine colon cancer MC38. In contrast, mice immunized with mp53 DNA were incapable of protecting mice against challenge with MC38 tumors. In a therapeutic model, established MC38 tumors were also well controlled by treatment with hp53DNA therapy in tumor bearing mice compared to mp53 DNA. Mice vaccinated with hp53 DNA plasmid also exhibited an increase in mp53-specific CD8+ T cell precursors compared to vaccination with mp53 DNA. In antibody depletion experiments, we also demonstrated that CD8+ T cells play crucial roles in the antitumor effects.

Conclusion: This study showed intramuscular vaccination with xenogenic hp53 DNA vaccine followed by electroporation is capable of inducing potent antitumor effects against mutated p53 expressing tumors through CD8+ T cells.

Citation Format: Rueyshyang Soong, Chien-Fu Hung, T.-C. Wu. Cancer immunotherapy using naked DNA vaccine targeting p53. [abstract]. In: Proceedings of the AACR Special Conference on Tumor Immunology: Multidisciplinary Science Driving Basic and Clinical Advances; Dec 2-5, 2012; Miami, FL. Philadelphia (PA): AACR; Cancer Res 2013;73(1 Suppl):Abstract nr A17.
Abstract A29: An Ad5 [E1-, E2b-] platform carrying the TAA CEA(6D) induces CEA directed CMI responses in patients with advanced CEA-expressing colorectal cancer in a phase I/II clinical trial..
Authors: Morse Michael A. , Gabitzsch Elizabeth S. , Xu Younong , Rajesh Dua , Nguyen Susan , Balcaitis Stephanie , Balint Joseph P. , Jones Frank
Published: August 2014

First-generation, E1-deleted adenovirus (Ad)-based vectors, although promising platforms for use as cancer vaccines, are limited by the naturally occuring or induced Ad-specific neutralizing antibodies. We hypothesized that Ad5-based vectors with deletions of the E1 and the E2b regions (the latter encoding the DNA polymerase and the preterminal protein) would, by virtue of diminished late phase viral protein expression, avoid immunological clearance and induce more potent immune responses against the encoded tumor antigen transgene in Ad-immune hosts. In this translational study, we demonstrated that delivery of increasing doses of the Ad5 [E1-, E2b-]-CEA(6D) in mice resulted in CEA-specific cell-mediated immune (CMI) responses. In the follow-up human phase I/II study, escalating doses of the vaccine induced higher CEA-specific CMI responses despite the presence of pre-existing Ad5 immunity in a majority (76%) of patients. Importantly, there was minimal toxicity, and overall patient survival (54.1% at 12 months) was similar regardless of pre-existing Ad5 neutralizing antibody titers. The results demonstrate that the novel Ad5 [E1-, E2b-] gene delivery platform can both break tolerance and generate significant CMI responses to the TAA CEA in the setting of both naturally acquired Ad5-specific immunity and/or immunization-induced Ad5 immunity.

Citation Format: Michael A. Morse, Elizabeth S. Gabitzsch, Younong Xu, Dua Rajesh, Susan Nguyen, Stephanie Balcaitis, Joseph P. Balint, Jr., Frank Jones. An Ad5 [E1-, E2b-] platform carrying the TAA CEA(6D) induces CEA directed CMI responses in patients with advanced CEA-expressing colorectal cancer in a phase I/II clinical trial.. [abstract]. In: Proceedings of the AACR Special Conference on Tumor Immunology: Multidisciplinary Science Driving Basic and Clinical Advances; Dec 2-5, 2012; Miami, FL. Philadelphia (PA): AACR; Cancer Res 2013;73(1 Suppl):Abstract nr A29.
Abstract A55: Inhibition of the heat shock protein receptor CD91 abrogates concomitant antitumor immunity.
Authors: Zhou Yu Jerry , Binder Robert J.
Published: August 2014

Most tumors are capable of generating immune response in the bearing host, a phenomenon described as concomitant immunity. Since tumors lack the typical (foreign) PAMPs that provide signal 2 for T cell priming, the mechanism of how concomitant immunity is established remains a mystery. We provide evidence here that, heat shock proteins (HSPs), when released into the tumor microenvironment, is the immunogen necessary for priming concomitant tumor specific immunity. These responses require CD91- the endocytic and signaling receptor for the immunogenic HSPs. In our study, we generated cd11c-specific CD91 knockout mice and showed that tumors grew faster in those knockout mice, compared to CD91+/+ mice. In addition, we abrogated the interaction of tumor-derived HSP with CD91 in vivo by over-expression of Receptor Associated Protein (RAP), an endogenous inhibitor of CD91. The data showed that: first, tumors expressing RAP grew with significantly faster kinetics than the non-RAP expressing counterparts in wild type mice, although this difference was non-existent in immunocompromised mice and the inherent proliferative rates of tumors with or without RAP were identical; second, RAP-expressing tumors, when used as an immunogen in tumor prophylaxis, were significantly less efficient in priming immune responses; third, we examined the competition for CD91 between RAP and tumor-derived HSP in vivo. In the presence of RAP, less amount of tumor-derived EGFP-labeled HSPs were taken up by APCs in draining lymph nodes. In summary, our study demonstrates that the HSP-CD91 pathway is critical for establishment of concomitant immunity. Considering that elevated RAP levels have been reported in colon cancer patients, we propose a novel mechanism of immune evasion for tumors expressing competing ligands for immune receptors such as CD91.

Citation Format: Yu Jerry Zhou, Robert J. Binder. Inhibition of the heat shock protein receptor CD91 abrogates concomitant antitumor immunity. [abstract]. In: Proceedings of the AACR Special Conference on Tumor Immunology: Multidisciplinary Science Driving Basic and Clinical Advances; Dec 2-5, 2012; Miami, FL. Philadelphia (PA): AACR; Cancer Res 2013;73(1 Suppl):Abstract nr A55.
Abstract A7: TARP-TCR-engineered T cells specifically kill HLA-A2 positive prostate and breast cancer cells.
Authors: Hillerdal Victoria , Nilsson Berith , Carlsson Bjorn , Eriksson Fredrik , Essand Magnus
Published: August 2014

In order to produce genetically engineered T cells directed against prostate and breast cancer cells we have cloned the T cell receptor recognizing the human leukocyte antigen (HLA)-A2-restricted TARP4-13 epitope. TARP is a protein exclusively expressed in normal prostate epithelium and in adenocarcinomas of the prostate and breast. Peripheral blood T cells transduced with a lentiviral vector encoding the TARP-TCR proliferated well when exposed to peptide-specific stimuli. They exerted peptide-specific interferon-γ production and cytotoxic activity. Importantly, HLA-A2+ prostate and breast cancer cells expressing TARP were also killed, demonstrating that the TARP4-13 epitope is a physiologically relevant target for T cell therapy of prostate and breast cancer. In conclusion, we present the cloning of a TCR directed against a physiologically relevant HLA-A2 epitope of TARP. To our knowledge this is the first report on engineering of T cells with a TCR directed against an antigen specifically expressed by prostate cells.

Citation Format: Victoria Hillerdal, Berith Nilsson, Bjorn Carlsson, Fredrik Eriksson, Magnus Essand.TARP-TCR-engineered T cells specifically kill HLA-A2 positive prostate and breast cancer cells. [abstract]. In: Proceedings of the AACR Special Conference on Tumor Immunology: Multidisciplinary Science Driving Basic and Clinical Advances; Dec 2-5, 2012; Miami, FL. Philadelphia (PA): AACR; Cancer Res 2013;73(1 Suppl):Abstract nr A7.
Abstract A70: Interactions between dendritic cells and T lymphocytes in a novel 3-D environment.
Authors: Steponavicius-Cruz Karen , Freitas Vanessa Morais , Barbuto José Alexandre Marzagão
Published: August 2014

Introduction: Dendritic cells (DCs) are the major antigen-presenting cells and, in situations of change in homeostasis, are able to initiate immune responses through the activation of T lymphocytes. Their immunomodulatory functions and generation in vitro are of great potential for immunotherapy of cancer. Among the many strategies under investigation, one, which our group has been using, is based on the fusion of DCs and tumor cells for therapeutic vaccination. To better understand the interactions between these fused DC-tumor cell hybrids and the different T cell populations, we are developing a three-dimensional (3D) culture system, whose first results are presented here.

Methods and Results: Monocytes of healthy donors were purified from PBMCs by magnetic beads selection and induced to differentiate into DCs by treatment with GM-CSF and IL-4. Maturation of monocyte-derived DCs (mMo-DCs) was initiated by addition of TNF-alpha 48 hours or 2.5 hours before analysis. Immature DCs were generated for 7 days in the absence of a maturation stimulus. Allogeneic T lymphocytes were purified from PBMCs by magnetic beads selection and were co-cultured for 17 hours in the 3D Biotek scaffold, at 37 °C. These scaffolds are made from polystyrene and have a thickness of 600 μm, fibers of 150 μm and pores of 200 μm. In this system, mMO-DCs TNF-activated for 2.5 h did not interact with allogeneic T lymphocytes, while those activated for 48 hours did. The average time of interaction between mMo-DCs and T lymphocytes was 4 hours, and the average speed of the T lymphocytes in the scaffold containing 48h-activated mMO-DCs was of 0.94 µm/min. Immature DCs interact with allogeneic T lymphocytes and the average time of interaction was 57 minutes. The average speed of the T lymphocytes in the scaffold containing immature DCs was of 0.93 µm/min.

Conclusions: These data indicate that this 3D Biotek scaffold enables interactions between mMO-DCs and lymphocytes and may be useful for the characterization of these interactions, the cellular subtypes and patterns of response induced. Financial Support: FAPESP (2010/18139-7; 2009/54599-5; CNPq).

Citation Format: Karen Steponavicius-Cruz, Vanessa Morais Freitas, José Alexandre Marzagão Barbuto. Interactions between dendritic cells and T lymphocytes in a novel 3-D environment. [abstract]. In: Proceedings of the AACR Special Conference on Tumor Immunology: Multidisciplinary Science Driving Basic and Clinical Advances; Dec 2-5, 2012; Miami, FL. Philadelphia (PA): AACR; Cancer Res 2013;73(1 Suppl):Abstract nr A70.
Abstract B19: Tumor-specific structural rearrangements as potential targets for anticancer vaccines.
Authors: Martin Spencer David , Wick Darin A. , Webb John R. , Holt Robert A. , Nelson Brad H.
Published: August 2014

A hallmark of cancer is the accumulation of mutations that allow cells to proliferate uncontrollably. Every tumor has a unique set of somatic mutations, which can be identified with next generation sequencing. In theory, CD8+ T cells can recognize mutated proteins present in human cancers if the mutations are presented as peptides bound to MHC class I molecules. We hypothesize that personalized, tumor-specific peptide vaccines will activate CD8+ T cells and induce tumor regression. To test this concept, we subjected four mouse mammary tumors to RNA sequencing using the Illumina sequencing platform. We identified 14 somatic point mutations and one fusion transcript unique to these tumors. In addition, we have optimized a vaccination strategy involving long peptides and the adjuvant poly (I:C) that results in massive proliferation of antigen-specific CD8+ T cells. When we used this optimized vaccination protocol to target tumor specific mutations, a strong T cell response was elicited towards the fusion protein, but only weak responses or no responses were elicited towards the point mutations. Currently, we are vaccinating mice with immunogenic mutated-peptides, and assessing whether the T cell response elicited by the vaccines causes regression or rejection of established tumors that harbor the same mutations. As the cost of sequencing the human genome continues to decrease, personalized vaccines that target tumor-specific mutations may become a treatment strategy that is clinically feasible.

Citation Format: Spencer David Martin, Darin A. Wick, John R. Webb, Robert A. Holt, Brad H. Nelson. Tumor-specific structural rearrangements as potential targets for anticancer vaccines. [abstract]. In: Proceedings of the AACR Special Conference on Tumor Immunology: Multidisciplinary Science Driving Basic and Clinical Advances; Dec 2-5, 2012; Miami, FL. Philadelphia (PA): AACR; Cancer Res 2013;73(1 Suppl):Abstract nr B19.
Abstract B29: Improving the identification of beneficial mimotopes using a representative tumor-specific T cell receptor.
Authors: Slansky Jill E. , Buhrman Jonathan D.
Published: August 2014

Vaccination with peptide mimotopes is a promising strategy to improve antitumor immunity. Mimotopes function by eliciting increased numbers of T cells that cross-react with the native tumor antigen. Using a high affinity tumor-specific T cell clone (CT), we have identified a panel of mimotopes for the dominant antigen (AH1) of a mouse colon tumor cell-line (CT26) that elicits a range of tumor protection when used in vaccines. Ex vivo evaluation of the TCRs of T cells initiated by mimotopes that protect against tumor challenge revealed increased representation of a Vβ8.3/Jβ2.6 motif: a small hydrophobic amino acid, followed by a large polar residue, and the tyrosine of the Jβ2.6. Clonotypes with this motif were also identified within the AH1-specific T cell repertoire of naturally responding TIL and following vaccination with the AH1 antigen. This motif was not found in the original CT clone. One particular clone with the motif made up nearly 8.5% of all Vβ8.3 sequences identified following immunization with mimotopes. Thus, we tested the hypothesis that this ‘representative’ TCR improves the frequency of discovery of efficacious mimotopes for cancer immunotherapy relative to the high affinity CT clone in the CT26 model. We have demonstrated that this is the case by screening genetically encoded peptide libraries. These results have important implications when considering the optimal T cells responding to tumor antigens and the improved identification of mimotopes for cancer immunotherapy.

Citation Format: Jill E. Slansky, Jonathan D. Buhrman. Improving the identification of beneficial mimotopes using a representative tumor-specific T cell receptor. [abstract]. In: Proceedings of the AACR Special Conference on Tumor Immunology: Multidisciplinary Science Driving Basic and Clinical Advances; Dec 2-5, 2012; Miami, FL. Philadelphia (PA): AACR; Cancer Res 2013;73(1 Suppl):Abstract nr B29.
Abstract B47: Neural cells labeling with intracerebral ferumoxytol iron oxide nanoparticles in a rat model.
Authors: Wu Yingjen Jeffrey , Muldoon Leslie L. , Neuwelt Edward A.
Published: August 2014

Purpose: To investigate uptake of ferumoxytol iron oxide nanoparticles into rat neural cells for a clinically applicable magnetic resonance (MR) imaging technique.

Materials and Methods: Long Evan rats were injected with intracerebral ferumoxytol (360 μg) iron oxide nanoparticles into the right hemisphere. A needle track in the left hemisphere was used as a sham control. Animals underwent T2-weighted fast spin-echo MRI sequences at 12T 48 h after nanoparticle inoculation. Immediately after MRI, single brain cell suspensions were produced from the right and left hemispheres, and neurospheres were stained for iron using Prussian blue histochemstry.

Results: The ferumoxytol-injected right hemisphere showed decreased T2 signal compared to the sham injected left hemisphere. Positive iron staining was found in neural cells from the right cerebral hemisphere, although <5% of total cell number, while no positive iron staining cells were found in the left hemisphere. The decreasing T2 signal does not correlate the percentage of neural cells labeled with ferumoxytol iron oxide nanoparticles, indicating most of the nanoparticles were not cell-associated. The phenotypes of ferumoxytol-labeled neural cells will be characterized by immunohistochemistry for neural markers. No neural cell in vitro labeling was detected using ferumoxytol in combination with protamine sulfate and heparin.

Conclusion: Iron oxide loading of neural cells in the brain parenchyma can be achieved with the clinically approved agent ferumoxytol. Iron labeling may be utilized to monitor neuro-inflammation and brain tumors after chemotherapy for non-invasive MRI.

Citation Format: Yingjen Jeffrey Wu, Leslie L. Muldoon, Edward A. Neuwelt. Neural cells labeling with intracerebral ferumoxytol iron oxide nanoparticles in a rat model. [abstract]. In: Proceedings of the AACR Special Conference on Tumor Immunology: Multidisciplinary Science Driving Basic and Clinical Advances; Dec 2-5, 2012; Miami, FL. Philadelphia (PA): AACR; Cancer Res 2013;73(1 Suppl):Abstract nr B47.
Abstract B48: Resensitization to CDDP or trail-mediated apoptosis in ovarian cancer cells by Obatoclax.
Authors: Martinez-Paniagua Melisa A. , Romero-Gonzalez Luis A. , Anguiano-Hernandez Yu-Mei , Gonzalez-Torres Carolina , Vega Mario I.
Published: August 2014

Ovarian cancer is the fifth cause of death in women, annually 13,850 deaths are reported in United States. Chemo is the conventional therapy after surgery but the initial response is between 60 to 80%, however, some patients that initially respond well to the therapy suffer recurrent disease or become resistant to the treatment. As a consequence, the overall 5 year survival is only 30%. There is a necessity to identify novel therapies for ovarian cancer or to discover drugs witch re sensitize tumor cells to existence therapy. It has been suggested that the resistance of the tumor cells is because of the increased expression of antiapoptotic proteins especially from the Bcl2 family (Bcl-2, Bcl-XL, Mcl-1). Obatoclax is a small molecule that mimics the BH3 domain of the BH3 only family members, inhibiting the effect of antiapoptotic Bcl-2 members. The objective of this study was to assess the susceptibility of ovarian cancer cell lines to chemo or immune therapy mediated apoptosis by obatoclax and also to elucidate the mechanism. Some ovarian cancer cell lines were treated with different concentrations of Obatoclax, CDDP or TRAIL alone or in combinational treatment to assess the reduction of viability measured by the XTT kit (Roche). Basal and cell treated total protein expressions were assessed by western. Induction of apoptosis was measured by TUNEL. As a single agent, Obatoclax or CDDP inhibited the growth of ovarian cancer cell lines in a dose response manner, TRAIL was no having an evident response showing a plateau with the concentration used. The combinational treatment after selecting the IC50 and IC20 response showed an increased inhibition of the viability. The induction of apoptosis was mediated by a caspase dependent manner. Our data suggest that obatoclax in combination with CDDP or TRAIL can re sensitize resistance cell lines to apoptosis and also it can be an optional treatment of patients with ovarian cancer.

Citation Format: Melisa A. Martinez-Paniagua, Luis A. Romero-Gonzalez, Yu-Mei Anguiano-Hernandez, Carolina Gonzalez-Torres, Mario I. Vega. Resensitization to CDDP or trail-mediated apoptosis in ovarian cancer cells by Obatoclax. [abstract]. In: Proceedings of the AACR Special Conference on Tumor Immunology: Multidisciplinary Science Driving Basic and Clinical Advances; Dec 2-5, 2012; Miami, FL. Philadelphia (PA): AACR; Cancer Res 2013;73(1 Suppl):Abstract nr B48.
Abstract B52: A novel quantitative proteomics approach to identify proteins that elicit antibody responses in vaccinated pancreatic cancer patients.
Authors: Jhaveri Darshil T. , Kim Min-Sik , Huang Lanqing , Sharma Rajni , Anders Robert A. , Pandey Akhilesh , Jaffee Elizabeth M.
Published: August 2014

Pancreatic cancer is the fourth leading cause of cancer deaths in the US. Less than 4% of patients have an overall survival of more than 5 years. Our lab has developed an allogeneic, granulocyte-macrophage colony-stimulating factor (GM-CSF) secreting pancreatic cancer vaccine that has recently completed phase II clinical trials. The application of proteomic approaches has gained ground in the identification of new cancer biomarkers, which are beneficial for both early detection and determination of new targets for development of biologically relevant therapies. We have developed a novel Serum Antibodies based SILAC-Immunoprecipitation (SASI) approach. The SASI approach utilizes immunoprecipitation by serum antibodies, which is coupled to the quantitative stable isotope labeling by amino acids in cell culture (SILAC) to identify proteins that elicit an increased antibody response post-vaccination. Using mass spectrometric analysis, the SASI approach comprehensively identified >40 proteins that elicited at least a 2-fold increase in antibody response post-vaccination, from each of the three patients' sera used. The list of identified proteins includes membrane proteins such as MHC class I complex, transferrin receptor (TFRC) and CD9, secreted proteins such as mesencephalic astrocyte-derived neurotrophic factor (MANF), as well as several cytoplasmic proteins such as regulatory subunit 12A of protein phosphatase 1 (PPP1R12A), galectin 3 and regulatory subunit 8 of the 26S proteasome (PSMC5). Most significantly, the antibody responses detected against these proteins in patients with >3 years disease-free survival suggests an anti-tumor potential of targeting these proteins. We further analyzed PSMC5, TFRC and PPP1R12A for tissue expression in normal, pre-malignant and pancreatic tumor specimens and found these proteins to increase in expression with tumor development. In addition, these proteins were shown to be pancreatic cancer associated antigens that are recognized by post-vaccination antibodies in the sera of patients that received the vaccine and have demonstrated a favorable disease free survival. Overall, our data demonstrates that the novel SASI approach can enable identification of candidate proteins as possible new biomarkers for screening, and as new targets for therapeutic intervention.

Citation Format: Darshil T. Jhaveri, Min-Sik Kim, Lanqing Huang, Rajni Sharma, Robert A. Anders, Akhilesh Pandey, Elizabeth M. Jaffee. A novel quantitative proteomics approach to identify proteins that elicit antibody responses in vaccinated pancreatic cancer patients. [abstract]. In: Proceedings of the AACR Special Conference on Tumor Immunology: Multidisciplinary Science Driving Basic and Clinical Advances; Dec 2-5, 2012; Miami, FL. Philadelphia (PA): AACR; Cancer Res 2013;73(1 Suppl):Abstract nr B52.
Abstract B6: Myeloid-derived suppressor cells from melanoma patients suppress T cell proliferation and are associated with a shorter disease-free interval.
Authors: McCarter Martin D. , Jordan Kimberly
Published: August 2014

Immune therapies such as vaccines, immune stimulators (IL-2), and immune checkpoint regulators (anti-CTLA-4) for melanoma elicit an initial robust immunologic response to the tumor in some patients, however, the majority ultimately go on to die of their disease. This suggests that tumor eventually develop escape mechanisms meant to evade or defeat the intrinsic immune response. Myeloid-derived suppressor cells (MDSCs) have been reported in melanoma and other cancer patients and may contribute to immune suppression or counter-regulatory responses. The aim of this project was to assess the potential role of circulating MDSCs in mediating immune suppression in advanced stage melanoma patients.

Methods: MDSC populations (DC14+ Lin-HLA-DR- , Lin-HLA-DR- CD11b+ CD33+) were isolated from patients with (stage I n=20, stage IV n=35)or without (n=15) melanoma. Isolated MDSC's were mixed with allogeneic CFSE-labeled T cells for a 4 day mixed lymphocyte reaction (MLR). T cell activation and proliferation were quantified.

Results: Stage IV melanoma patients have increased expression (2-3 times) of some MDSC populations (Lin-HLA-DR- CD11b+ CD33+) in the circulation compared to normal and stage I melanoma patients. By MLR, these MDSC's significantly suppressed T cell proliferation (40-50% reduction) and markers of activation (20%) to a greater extent than the same population of MDSC's for normal controls. Inhibiting nitric oxide failed to restore T-cell proliferation. A shorter disease free interval is associated with increased expression of CD33+ MDSC in the circulation.

Conclusion: MDSC populations are increased in the peripheral circulation of stage IV melanoma patients and these cells actively suppress T cell proliferation. Efforts are underway to better understand the melanoma related factors responsible for the induction of MDSC's and the mechanism by which these cells suppress T-cell proliferation.

Citation Format: Martin D. McCarter, Kimberly Jordan. Myeloid-derived suppressor cells from melanoma patients suppress T cell proliferation and are associated with a shorter disease-free interval. [abstract]. In: Proceedings of the AACR Special Conference on Tumor Immunology: Multidisciplinary Science Driving Basic and Clinical Advances; Dec 2-5, 2012; Miami, FL. Philadelphia (PA): AACR; Cancer Res 2013;73(1 Suppl):Abstract nr B6.
Abstract B62: miR-124 systemically enhances antitumor clearance by inhibiting STAT3 signaling and reversing glioma-associated immune suppression.
Authors: Wei Jun , Kong Ling-Yuan , Wang Fei , Xu Shuo , Doucette Tiffany , Ferguson Sherise D. , Yang Yuhui , McEnery Kayla , Jethwa Krishan , Gjyshi Olsi , Qiao Wei , Lang Frederick , Rao Ganesh , Fuller Greg N. , Calin George A. , Heimberger Amy B.
Published: August 2014

MicroRNAs (miRs) have been shown to modulate critical gene transcripts involved in tumorigenesis, but their role in tumor-mediated immune suppression is largely unknown. In this study, we evaluated miRNAs that are preferentially down-regulated in glioblastoma and that interact with immune suppressive pathways as potential new therapeutics. On the basis of miRNA-gene expression libraries of glioblastoma, tissue microarrays, and molecular modeling, we selected miR-124 as the lead candidate for modulating signal transducer and activator of transcription 3 (STAT3) signaling, a key pathway mediating immune suppression of the tumor microenvironment. Upon up regulating miR-124 in glioma cancer stem cells (gCSCs), the STAT3 pathway was inhibited and the miR-124 reversed gCSC-mediated immune suppression of T cell proliferation and Th1 polarization and decreased gCSC-induced Foxp3+ regulatory T cells (Tregs). Treatment of immune-suppressed glioblastoma patient T cells with miR-124 induced marked effector response including up regulation of IL-2, IFN-γ and TNF-α. Both systemic administration of miR-124 or adoptive miR-124 transfected T cell transfers exerted potent anti-glioma therapeutic effects in clonotypic and genetically engineered murine models of glioblastoma and enhanced effector responses in the local tumor microenvironment. These therapeutic effects were ablated in nude murine systems. Our findings highlight the potential application of miR-124 as a novel immunotherapeutic agent for neoplasm treatment by exploiting the immune system to mediate direct tumor cytotoxicity.

Citation Format: Jun Wei, Ling-Yuan Kong, Fei Wang, Shuo Xu, Tiffany Doucette, Sherise D. Ferguson, Yuhui Yang, Kayla McEnery, Krishan Jethwa, Olsi Gjyshi, Wei Qiao, Frederick Lang, Ganesh Rao, Greg N. Fuller, George A. Calin, Amy B. Heimberger. miR-124 systemically enhances antitumor clearance by inhibiting STAT3 signaling and reversing glioma-associated immune suppression. [abstract]. In: Proceedings of the AACR Special Conference on Tumor Immunology: Multidisciplinary Science Driving Basic and Clinical Advances; Dec 2-5, 2012; Miami, FL. Philadelphia (PA): AACR; Cancer Res 2013;73(1 Suppl):Abstract nr B62.
Abstract B67: Galectin-1 expression related to hypoxia in primary human melanoma as a driver for metastatic progression. A target to promote anticancer immunotherapy.
Authors: Ralph Stephen J. , Ito Koichi , Wachter Bernadette , Onturk Selda , Wilmott James , Hersey Peter
Published: August 2014

Galectin-1 is a major protein secreted by the majority of cancer types which is induced by HIF and plays important roles in the tumor microenvironment protecting against anticancer immune cell attack and from reactive oxygen species production. We have shown this protein to be more highly expressed within hypoxic regions of primary melanoma. Galectin-1 expression is critical for protecting endothelial cells in tumor angiogenesis and for promoting tumor metastasis. More recently, we have shown that inhibiting this protein using small drug molecules significantly enhances chemotherapeutic access inside tumors by promoting leakiness and facilitates TIL access inside tumors to kill cancer cells. When galectin-1 blocking drugs are used in combination as a triple therapy with modified whole cancer cell vaccines plus anti-CTLA4 Ig, this provides synergistic enhancements increasing CD8+ CTL anticancer TIL responses to inhibit cancer growth in a range of different mouse tumor models.

Citation Format: Stephen J. Ralph, Koichi Ito, Bernadette Wachter, Selda Onturk, James Wilmott, Peter Hersey. Galectin-1 expression related to hypoxia in primary human melanoma as a driver for metastatic progression. A target to promote anticancer immunotherapy. [abstract]. In: Proceedings of the AACR Special Conference on Tumor Immunology: Multidisciplinary Science Driving Basic and Clinical Advances; Dec 2-5, 2012; Miami, FL. Philadelphia (PA): AACR; Cancer Res 2013;73(1 Suppl):Abstract nr B67.
Abstract B76: CD8+FoxP3+ T cells: A new player in the immune response to ovarian cancer.
Authors: Kost Sara E.F. , deLeeuw Ronald J. , Nelson Brad H.
Published: August 2014

Introduction: Tumor-infiltrating lymphocytes (TIL) are an important prognostic indicator in high-grade serous ovarian carcinoma (HGSC). Certain types of TIL (in particular CD8+ effector T cells) predict better outcomes, whereas others (most notably CD4+CD25+FoxP3+ regulatory T cells or Tregs) predict worse outcomes. An unconventional subset of CD8+FoxP3+ T cells has been widely reported in autoimmunity. While the functional significance of CD8+FoxP3+ TIL remains poorly understood, in a murine tumor model they were associated with effective anti-tumor responses.

Hypothesis: CD8+FoxP3+ TIL are present in HGSC and correlate with effector TIL and increased patient survival.

Experimental Design: Multi-parameter flow cytometry and immunohistochemistry (IHC) were performed on cohorts of 12 and 45 primary HGSC specimens, respectively, to enumerate and locate CD8+FoxP3+ TIL. For IHC, intraepithelial versus stromal location was determined by staining adjacent sections for the epithelial marker cytokeratin. Multi-color staining was resolved using the Nuance™ multispectral imaging system in conjunction with Metamorph software. Survival analysis will be performed by staining a cohort of 200 cases of HGSC.

Results: By flow cytometry, CD8+FoxP3+ cells were found in variable proportions between HGSC cases, representing 1.3 - 7.9% of CD8+ TIL and 1.3 - 25.8% of FoxP3+ TIL. By IHC, CD8+FoxP3+ TIL were found together with CD8+FoxP3- and CD4+FoxP3+ TIL in epithelial and stromal tumor regions and represented up to 19% of the FoxP3+ population. Interestingly, the majority of CD8+FoxP3+ TIL were also found to be TIA-1+KI67-, consistent with an active role in the anti-tumor response.

Conclusions: CD8+FoxP3+ TIL are a component of the host immune response to HGSC. Survival analysis is ongoing and will determine whether CD8+FoxP3+ T cells enhance or inhibit tumor immunity. Improved understanding of this new TIL subset will inform future immunotherapy strategies for this challenging malignancy.

Citation Format: Sara E.F. Kost, Ronald J. deLeeuw, Brad H. Nelson. CD8+FoxP3+ T cells: A new player in the immune response to ovarian cancer. [abstract]. In: Proceedings of the AACR Special Conference on Tumor Immunology: Multidisciplinary Science Driving Basic and Clinical Advances; Dec 2-5, 2012; Miami, FL. Philadelphia (PA): AACR; Cancer Res 2013;73(1 Suppl):Abstract nr B76.
Abstract B91: Tasquinimod targets immuno-suppressive myeloid cell populations and enhances immunotherapy in murine prostate cancer models.
Authors: Shen Li , Miles Kiersten Marie , Ciesielski Michael , Ellis Leigh , Fenstermaker Robert , Pili Roberto
Published: August 2014

Purpose: A major component of the immunosuppressive environment responsible for immune tolerance in cancer is represented by myeloid cell populations such as myeloid derived suppressor cells (MDSCs) and tumor associated macrophages (TAMs). These cells facilitate carcinogenesis and tumor progression by impairing T and NK cells activation and contributing to modulate angiogenesis. Tasquinimod is an anti-angiogenic and immunomodulatory agent that has shown clinical benefit in patients with prostate cancer. This agent has been found to bind to the inflammatory protein S100A9, to block the interaction of S100A9 with myeloid cells surface receptors, and therefore has the potential to affect accumulation and function of these cells.

Experiments and Objectives: We tested tasquinimod in combination with a vaccine therapy in two castration resistant, syngenic transplantable murine prostate cancer models: CR Myc-Cap and TRAMP-C2. This study had the following objectives: 1) to explore tasquinimod's novel immunomodulatory activities and underlining mechanism; 2)To investigate whether tasquinimod inhibits the induction and function of immunosuppressive myeloid cell populations; 3) To assess whether tasquinimod can facilitate vaccine therapy in the castrate resistant prostate cancer models.

Results: We observed that the peripheral MDSCs (in blood and spleen) dramatically increased in tumor bearing mice as compared to naïve mice. The MDSCs number increase correlated with disease progression in the transgenic model. Tasquinimod treatment depleted Gr1-CD11b+, Ly6G-CD11b+ (monocytic MDSCs) myeloid cells and M2 macrophages in peripheral sites, and reduced Gr1+CD11b+ MDSCs infiltrates in the tumors. In addition, tasquinimod (10mg/kg) dramatically improved tumor-specific immune responses, including the granzyme B expression in CD8 cells and cytotoxicity of splenocytes, and enhanced anti-tumor activity of a survivin-mimetic peptide vaccine.

Conclusions: 1. MDSCs accumulate in prostate cancer models suggesting that MDSCs may contribute to immune tolerance in prostate cancer. 2. Tasquinimod reduces immunosuppressive MDSCs and M2 macrophage and may facilitate vaccine therapy by enhancing cell-mediated cytotoxic tumor cell killing and the anti-tumor immune response.

Citation Format: Li Shen, Kiersten Marie Miles, Michael Ciesielski, Leigh Ellis, Robert Fenstermaker, Roberto Pili. Tasquinimod targets immuno-suppressive myeloid cell populations and enhances immunotherapy in murine prostate cancer models. [abstract]. In: Proceedings of the AACR Special Conference on Tumor Immunology: Multidisciplinary Science Driving Basic and Clinical Advances; Dec 2-5, 2012; Miami, FL. Philadelphia (PA): AACR; Cancer Res 2013;73(1 Suppl):Abstract nr B91.
Abstract IA16: Engineered T cell therapies for cancer.
Authors: June Carl H.
Published: August 2014

Chimeric antigen receptors (CARs) combine an antigen recognition domain of a specific antibody with a intracellular signaling domains into a single engineered protein. CD19 is an ideal target for CARs since expression is restricted to normal and malignant B cells. In preclinical studies, inclusion of the CD137 (4-1BB) signaling domain results in remarkable antitumor activity and in vivo persistence of anti-CD19 CARs. We have previously reported dramatic in-vivo expansion, persistence and anti-tumor activity of CAR modified autologous T cells targeted to CD19 (CART19 cells) in 3 patients with CLL with relatively short follow up (Porter, et al NEJM 2011; Kalos et al Sci Trans Med 2011). Currently, 11 patients have been treated with CART19 cells. There were 9 adults (median age 65 years (range 51-78)) treated for relapsed, refractory and high risk CLL and two pediatric cases treated for relapsed refractory pre-B cell ALL. CLL patients had received a median of 5 prior regimens excluding single agent rituxumab (range 2-10) and all had active disease at the time of cell infusion. The pediatric patients with ALL had chemotherapy refractory relapse, and one was resistant to allogeneic transplantation and CD19 directed blinotumomab. A median of 7.5 x 10^8 total cells (range 1.7-50) corresponding to 1.45 (range 0.14-5.9 x 108) genetically modified cells were infused. In the patients in complete remission, CAR modified CART19 cells are detectable in the peripheral blood by flow cytometry at the most recent follow up for at least 2 years. All patients in CR have B cell aplasia. No patient with CR has progressed.

Citation Format: Carl H. June. Engineered T cell therapies for cancer. [abstract]. In: Proceedings of the AACR Special Conference on Tumor Immunology: Multidisciplinary Science Driving Basic and Clinical Advances; Dec 2-5, 2012; Miami, FL. Philadelphia (PA): AACR; Cancer Res 2013;73(1 Suppl):Abstract nr IA16.
Abstract IA23: Cervical tissue T cells in HPV disease.
Authors: Trimble C.L. , Teague J. , Wu T.C. , Clark R.A.
Published: August 2014

Despite the availability of preventative vaccines, human papillomaviruses (HPVs) cause more cancers than any other virus. Like other viral pathogens that cause human cancers, most HPV infections do not cause overt harm, let alone symptoms. Indeed, in immune-competent persons, most genital infections are eventually cleared. Nonetheless, the incidence of HPV-associated cancers in anatomic sites for which no screening algorithms have been validated is rising steadily. Preinvasive, intraepithelial lesions are associated with integration of the HPV genome into the host genome, and subsequent expression of the viral oncoproteins, E6 and E7, which are required for disease initiation and persistence. These true tumor-specific antigens provide non-“self” targets for immune therapies. The relatively indolent clinical course of preinvasive lesions, and tissue tropism of HPV infections provide opportunities to identify and treat early stage disease.

In cervical HPV disease, a subset of high grade dysplastic lesions, cervical intraepithelial neoplasia grade 3 (CIN3), do in fact undergo complete regression, which is presumably immunologically mediated. However, T cell responses to viral antigens, measured in the blood, have not been informative with respect to immune responses in the cervical mucosa.

Our team has been studying strategies to enhance the quality and magnitude of immune responses to HPV antigens. Ongoing clinical trials testing therapeutic vaccination and strategies to manipulate the lesion microenvironment to enhance homing and access of immune effector cells will be discussed. The design of these trials has been informed by studies of tissue-resident T cells in normal cervical mucosa, and by changes identified in dysplastic mucosa, both in lesions that regress, and in those that do not, in the course of a defined, prospective study window. Changes in the composition and distribution of tissue T cells postvaccination will be presented. Strategies to modify the stromal compartment of HPV lesions to promote lesion clearance will also be discussed.

Citation Format: C.L. Trimble, J. Teague, T.C. Wu, R.A. Clark. Cervical tissue T cells in HPV disease. [abstract]. In: Proceedings of the AACR Special Conference on Tumor Immunology: Multidisciplinary Science Driving Basic and Clinical Advances; Dec 2-5, 2012; Miami, FL. Philadelphia (PA): AACR; Cancer Res 2013;73(1 Suppl):Abstract nr IA23.
Abstract IA8: The interplay of inflammation and cancer.
Authors: Dranoff Glenn
Published: August 2014

Efficacious cancer immunotherapies will likely require combinations of strategies that enhance tumor antigen presentation and antagonize negative immune regulatory circuits. We demonstrated that vaccination with irradiated, autologous melanoma cells engineered to secrete GM-CSF followed by antibody blockade of CTLA-4 accomplishes clinically significant tumor destruction with minimal toxicity in a majority of stage IV metastatic melanoma and some advanced ovarian carcinoma patients. The detailed analysis of patients achieving sustained clinical benefits from these therapies illustrated the importance of a coordinated cellular and humoral anti-tumor immune response. Indeed, the extent of tumor necrosis in post-treatment biopsies was linearly related to the natural logarithm of the ratio of infiltrating CD8+ effector T cells to FoxP3+ Tregs. Moreover, therapy induced a potent antibody response to several key tumor cell surface and secreted proteins; these antibodies manifest functional activity in vitro, antagonizing tumor cell survival, invasive potential, and angiogenesis.

Through an analysis of cytokine deficient mice, we delineated a critical role for GM-CSF in Treg homeostasis. GM-CSF is required for the expression of the phosphatidylserine binding protein MFG-E8 in antigen presenting cells, whereas the uptake of apoptotic cells by phagocyte-derived MFG-E8 maintains peripheral Treg activity. While blockade of MFG-E8 can enhance therapeutic immunity in the context of cancer vaccines, loss of Treg homeostasis with MFG-E8 deficiency can lead to chronic inflammation. Mice deficient in GM-CSF and IFN-γ develop at high frequency germinal center derived lymphomas and nonsmall cell lung carcinomas. The pathogenesis of these tumors reflects a combination of tumor-promoting inflammation and a loss of protective tumor immunity. Taken together, these findings highlight dual roles for immunity in cancer pathogenesis and therapy.

Citation Format: Glenn Dranoff. The interplay of inflammation and cancer. [abstract]. In: Proceedings of the AACR Special Conference on Tumor Immunology: Multidisciplinary Science Driving Basic and Clinical Advances; Dec 2-5, 2012; Miami, FL. Philadelphia (PA): AACR; Cancer Res 2013;73(1 Suppl):Abstract nr IA8.