bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 1414: The role of RUNX3 and RUNX3-related chemokines in lung cancer-induced bone resorption.
Authors: Kim Hyun-Jeong , Chung Won-Yoon , Park Kwang-Kyun
Published: August 2014

Lung cancer frequently metastasizes to bone, which is associated with significant morbidity and a dismal prognosis. Runt-related transcription factor 3 (RUNX3) functions as a tumor suppressor in many cancers, and may be also important in the development of lung cancer. In this study, we found that RUNX3 knockdown increased the proliferation, migration and invasion of human lung cancer A549 cells. Interestingly, the receptor activator of nuclear factor-kappaB ligand (RANKL)/osteoprotegerin (OPG) ratio, an index of osteoclastogenic stimulation, was significantly increased in human osteoblastic hFOB1.19 cells treated with RUNX3 knockdown A549 cell-derived conditioned medium (CM) as compared to CM of RUNX3-expressing A549 cells. In the chemokine and receptor PCR array to identify RUNX3-regulated factors that mediate bone invasion of lung cancer cells, RUNX3 knockdown led to upregulation of CCL5 and downregulation of CCL19 and CXCL11 in A549 cells. CCL5 increased the proliferation, migration and invasion of A549 cells but CCL19 and CXCL11 had not a remarkable effect. CCL5 increased RANKL/OPG ratio in hFOB1.19 cells but CCL19 and CXCL11 decreased. CCL5 promoted RANKL-induced osteoclast differentiation in BMMs, while CCL19 and CXCL11 reduced. In the mouse model that RUNX3-expressing or RUNX3 knockdown A549 cells were injected subcutaneously over the calvaria, RUNX3 knockdown resulted in the increased tumor size and severe bone destruction. Additionally, in the osteolytic bone invasion model that the cells were inoculated into bone marrow of mouse tibia, bone lesions appeared more seriously in mice injected with RUNX3 knockdown A549 cells. The serum level of CCL5 was increased, while those of CCL19 and CXCL11 were decreased, in RUNX3 knockdown A549 cell-inoculated mice as compared with RUNX3-expressing A549 cell-inoculated mice. Taken together, bone invasion of lung cancer cells may be closely related with RUNX3 and RUNX3-regulated chemokines.

Citation Format: Hyun-Jeong Kim, Won-Yoon Chung, Kwang-Kyun Park. The role of RUNX3 and RUNX3-related chemokines in lung cancer-induced bone resorption. [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 1414. doi:10.1158/1538-7445.AM2013-1414
Abstract 1417: CXCR2 plays critical roles in colitis and colitis-associated cancer through trafficking of myeloid-derived suppressor cells.
Authors: Katoh Hiroshi , Wang Dingzhi , Kim Sun-Hee , DuBois Raymond N.
Published: August 2014

A large body of evidence indicates that chronic inflammation is a risk factor for colorectal cancer (CRC). However, all of the mechanisms by which inflammation promotes CRC development are not fully understood. Our lab previously showed that PGE2 promotes CRC progression through angiogenic chemokine CXCL1 (Wang et al., 2006). CXCL1 exerts its downstream functions via binding to its receptor, CXCR2, which is a G-protein-coupled receptor. Our previous study showed that CXCR2 expression is also elevated mainly in endothelial and immune cells of human colorectal carcinomas. CXCR2 is expressed on neutrophils and involved in neutrophil trafficking. These results prompted us to hypothesize that CXCR2 may be one of mediators connecting inflammation and CRC. To address this question, preliminary experiments were performed to determine whether CXCR2 is involved in inflammation-enhanced CRC progression.

Our results revealed that deletion of CXCR2 significantly attenuated AOM/DSS induction of chronic colonic inflammation and tumor burden, suggesting that CXCR2 is required for promoting chronic inflammation and tumor formation and growth in the colon. Among immunocytes we tested, loss of CXCR2 only significantly suppressed a massive infiltration of MDSCs (CD11b+Ly6G+) into the colonic mucosa in both normal (water treatment) and chronic inflammatory (DSS treatment) conditions. We further found that the status of CXCR2 did not affect MDSC populations in the bone marrow of mice treated with either water or DSS. Interestingly, treatment with DSS more dramatically reduced MDSC populations in the circulatory system of WT mice than of CXCR2 knockout mice, suggesting that CXCR2-deficient MDSCs have less ability to migrate to local organs. Our in vitro results further demonstrated that treatment with CXCL1 or CXCL5 were able to induce chemotaxis of WT-mice derived MDSCs but not MDSCs isolated from CXCR2-/- mice. Collectively, these results suggest that CXCR2-expressing MDSCs contribute to chronic inflammation and colitis-associated tumor formation and growth. Our findings are novel and provide a rationale for development of new therapeutic approaches to subvert tumor-induced immunosuppression by using CXCR2 antagonists and neutralized antibodies.

Citation Format: Hiroshi Katoh, Dingzhi Wang, Sun-Hee Kim, Raymond N. DuBois. CXCR2 plays critical roles in colitis and colitis-associated cancer through trafficking of myeloid-derived suppressor cells. [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 1417. doi:10.1158/1538-7445.AM2013-1417
Abstract 1430: The role of IL27 as a driver of skin carcinogenesis.
Authors: Dibra Denada , Newman Melissa , Cutrera Jeffry , Li Shulin
Published: August 2014

Although the genetic components that drive carcinogenesis and the stem cell involvement in cancer initiation have been established, the mechanism through which the immune response/cytokine milieu within the tumor microenvironment promotes cancer formation and stem cell behavior is an exciting yet understudied area of research. This statement is specifically true with an important regulator of the immune response, IL27, which is known to have both pro- and anti-inflammatory properties. During an inflammatory response, IL27 has been shown to regulate two canonical cytokines IL10 and IL17, and to affect multiple inflammatory and infectious diseases. Yet, the function of IL27 at physiological levels during the instigation of carcinogenesis remains unknown. To determine the role of IL27 during the early stages of cancer formation, the well-established two step skin carcinogenesis model (DMBA followed by the promoter benzoyl peroxide (BP)) was used. Interestingly, mice that lack IL27 signaling (IL27RA-/-) are protected during the initiation of carcinogenesis as they have lower incidence of papilloma formation over time when compared to wildtype mice. Additionally, wildtype mice treated with IL27 via gene therapy have a higher incidence and develop more papillomas when compared to control counterparts. Bone marrow transfer studies showed that IL27 signaling in both hematopoietic and non-hematopoietic cells is needed to drive skin carcinogenesis. Additionally, IL27 signaling induces loss of epithelial stem cell homeostasis/maintenance. In mice treated with DMBA/BP, 23% of the wildtype mice treated had ulcerations that failed to heal over time, while none of IL27RA-/- showed any defects in the skin. In mice that went under bone marrow transfer followed by DMBA/BP, 40% of wildtype ones developed severe skin ulcerations, while none of the IL27RA-/- developed any skin abnormalities. And lastly, mice undergoing the tumorigenesis protocol that were treated with IL27 developed hair loss, which was mirrored by loss of hair follicles. These phenotypes are like due to the loss of epithelial stem cell maintenance and/or homeostasis by IL27. These finding propose an unrecognized role of IL27 in promoting papilloma formation and disrupting epithelial cell maintenance and/or homeostasis.

Citation Format: Denada Dibra, Melissa Newman, Jeffry Cutrera, Shulin Li. The role of IL27 as a driver of skin carcinogenesis. [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 1430. doi:10.1158/1538-7445.AM2013-1430
Abstract 1460: Quantification of circulating tumor cells using multiplexed Surface-Enhanced Raman Scattering Nanoparticles.
Authors: Wang Xu , Qian Ximei , Beitler Jonathan J. , Chen Zhuo (Georgia) , Khuri Fadlo R. , Lewis Melinda M. , Shin Hyung J. , Nie Shuming , Shin Dong M.
Published: August 2014

Background: Detection, quantification, and molecular characterization of CTCs may aid physicians in treatment decision making and optimizing. However, the major challenge in the detection and characterization of CTCs is the lack of a highly specific and sensitive assay to isolate and identify the rare population of CTCs in complex blood samples. Moreover, current approaches for CTC detection including CellSearch (FDA approved CTC detection method) which primarily relays on single biomarker are suboptimal because of tumor heterogeneity. Here we develop a multiplexed single-tube assay for directly detection of CTCs in blood sample using two targeted Surface-Enhanced Raman scattering (SERS) nanoparticles. Methods: SERS nanoparticles (60 nm) were prepared by spectral encoding with a Raman reporter molecule (QSY or BHQ) and then stabilized and protected by low molecular weight polyethylene glycols (PEGs). A targeting ligand, epidermal growth factor (EGF) peptide or epithelial cell adhesion molecule (EpCAM) antibody, was covalently conjugated to PEG layer of SERS nanoparticle. The functionalized EGF- or EpCAM- SERS nanoparticle was able to identify CTCs in the peripheral blood with high specificity and affinity. Results: We collected and examined fresh blood samples from 20 SCCHN patients with various histologies and successfully detected positive SERS signals that were greater than background in 9 of 12 (75%) of patients. This technique was demonstrated to be capable of identifying rare CTCs by two biomarkers simultaneously, with a range of 5-720 cells per ml of whole blood. Blood samples from 3 normal controls did not show any CTCs. In one patient who had 0 count of CTC detected by EGFR-targeting SERS tag showed 170 CTCs/ml of blood detected by EpCAM-targeting SERS tags. Conclusion: Special design of the two SERS nanoparticles, distinguished with unique spectral signatures and bio-conjugation of EGF or EpCAM antibody as targeting molecule, allowed simultaneous detection of epidermal growth factor receptor (EGFR) or EpCAM on CTCs in one blood sample. Our technique based on two biomarkers rather than those relayed on single biomarker may improve CTC assessment in clinical settings. (This research was supported by the National Cancer Institute award 5 P50 CA128613 and U54CA119338-04).

Citation Format: Xu Wang, Ximei Qian, Jonathan J. Beitler, Zhuo (Georgia) Chen, Fadlo R. Khuri, Melinda M. Lewis, Hyung J. Shin, Shuming Nie, Dong M. Shin. Quantification of circulating tumor cells using multiplexed Surface-Enhanced Raman Scattering Nanoparticles. [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 1460. doi:10.1158/1538-7445.AM2013-1460
Abstract 1465: Prognostic significance of stem cell and EMT phenotype in Circulating Tumor Cells.
Authors: Strati Areti D. , Georgoulias Vasilis , Lianidou Evi
Published: August 2014

Background: Circulating tumor cells (CTCs) are associated with prognosis and can be used as a liquid biopsy for repeated follow up examinations. The subpopulation of CTCs that successfully establish distant metastases share EMT and stemness features. These cells have been found to be associated with resistance to anti-cancer therapies and treatment failure.

Patients and methods: Peripheral blood (20 mL in EDTA) was obtained and after density gradient centrifugation, immunomagnetic Ber-EP4 coated capture beads were used to enrich for epithelial cells from 102 patients with early breast cancer and 23 patients with verified metastasis. Messenger RNA was isolated from enriched epithelial cells using oligo (dT)25 coated magnetic beads. After cDNA synthesis the expression of CK-19, MAGE-A3, HER-2, PBGD and CD24, CD44, ALDH1, HPRT was tested, using two different quadruplex RT-qPCR assays. Moreover we performed RT-qPCR for the expression of TWIST1, hMammaglobin and hTERT α+β+.

Results: The presence of circulating tumor cells was evaluated using CK-19, MAGE-A3, HER-2, hTERT α+β+, and mammaglobin gene transcripts. In 102 operable breast cancer patients 31 patients (30.4%) were found positive for the presence of CTCs. CTC biomarkers were significantly correlated with lymph nodes (P=0.032). In the CTC+ group, 10 patients (32.3%) were positive for TWIST1 and 16 (51.6%) were positive for CD44+/CD24-/low and ALDH1+ cell phenotype. We found a significant correlation between HER2 negative tumors with TWIST1 expression (P=0.027) and Stem Cell phenotype (P=0.012). The detection of both EMT and Stem Cell characteristics was associated with shorter disease-free survival (P=0.024). In the CTC positive group of patients with verified metastasis (39.1%) 1 patient (11.1%) was positive for TWIST1 and 4 (44.4%) were positive for CD44+/CD24-/low and ALDH1+ cell phenotype. The detection of CK19+, Stem Cell and Stem Cell and/or EMT phenotype was associated with shorter overall-free survival (P=0.036, P=0.001 and P=0.002).

Discussion: CTCs in breast cancer patients share both stem cell and/or EMT phenotype. The detection and molecular characterization of CTCs with an EMT or stem cell-like characteristics could be a powerful diagnostic tool for the determination of prognosis and designing new therapeutic drugs for the elimination of minimal residual disease.

Citation Format: Areti D. Strati, Vasilis Georgoulias, Evi Lianidou. Prognostic significance of stem cell and EMT phenotype in Circulating Tumor Cells. [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 1465. doi:10.1158/1538-7445.AM2013-1465
Abstract 1489: The T-box transcription factor Brachyury blocks cell cycle progression and mediates tumor resistance to chemotherapy and radiation.
Authors: Huang Bruce K. , Cohen Joseph , Fernando Romaine I. , Hamilton Duane H. , Litzinger Mary T. , Hodge James W. , Palena Claudia M.
Published: August 2014

The T-box transcription factor Brachyury, a molecule frequently detected in human cancers but seldom found in normal adult tissue, has recently been proposed as a significant determinant of the epithelial-mesenchymal transition (EMT) in human carcinomas. In the current investigation, we present data demonstrating that in three different human lung carcinoma models, expression of Brachyury is associated with a mesenchymal phenotype. Additionally, elevated Brachyury expression is shown to strongly correlate with increased in vitro resistance to cytotoxic therapies, such as chemotherapy and radiation. Further investigation showed that chemotherapy treatment in vitro selected tumor cells that were high in Brachyury, and that the degree of resistance to therapy was comparable to the level of Brachyury expression. We also demonstrate that in vitro and in vivo, human lung carcinoma cells with greater levels of Brachyury divide at slower rates than those with lower levels of Brachyury, a phenomenon associated with marked downregulation of cyclin D1, phosphorylated Rb (pRb), and CDKN1A (p21). ChIP and luciferase resporter assays revealed that Brachyury represses p21 expression in carcinoma cells by directly binding to a half T-box consensus site located within the promoter region of the p21 gene, indicating a potential mechanism for the observed therapy resistance associated with Brachyury expression. Finally, we observed that in vivo treatment of tumor xenografts with chemotherapy resulted in the selective growth of resistant tumors that were high in Brachyury. Altogether, these results suggest that in addition to being a driver of EMT, Brachyury expression may attenuate cell cycle progression, and enable tumor cells to become less susceptible to chemotherapy and radiation in human carcinomas.

Citation Format: Bruce K. Huang, Joseph Cohen, Romaine I. Fernando, Duane H. Hamilton, Mary T. Litzinger, James W. Hodge, Claudia M. Palena. The T-box transcription factor Brachyury blocks cell cycle progression and mediates tumor resistance to chemotherapy and radiation. [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 1489. doi:10.1158/1538-7445.AM2013-1489
Abstract 1507: Iron upregulates HLA-G mRNA expression of the human MCF-7 breast cancer cell line.
Authors: Head Jonathan F. , Jiang Xian P. , Elliott Robert L.
Published: August 2014

HLA-G is a nonclassical MHC Class I molecule that protects the fetus from the maternal immune system, and it also plays a role in immune evasion of cancer. Previously we have shown that iron can suppress the cytolytic function of natural killer cells to human breast cancer cell lines. We have found that breast cancer cells over express HLA-G mRNA and proteins in both cell lines and breast carcinomas. In order to determine if iron concentration affects HLA-G expression, we cultured the human breast cancer cell line MCF-7 in media with 100 mM, 400 mM, and 1600 mM of iron for 24 hours. We used RT-PCR to examine HLA-G mRNA expression of the MCF-7 cells. The PCR primers were designed so that they could detect all alternatively spliced HLA-G mRNA isoforms, 1000 bp (HLA-G1 and HLA-G5), 600 bp (HLA-G2 and HLA-G4), and 300 bp (HLA-G3). RT-PCR showed that the MCF-7 cells cultured with control medium only had one band of 300 bp mRNA expression. However, the MCF-7 cells cultured in medium containing 100 mM, 400 mM, and 1600 mM of iron had three bands of 300 bp, 600 bp, and 1000 bp HLA-G mRNA expression. Moreover, the intensity of the 300 bp band from the iron-treated MCF-7 cells is significantly higher than the cells cultured with control medium. In summary, iron upregulates the HLA-G mRNA expression of MCF-7 cells, and this probably contributes to cancer immunosuppression and to the cytolytic dysfunction of natural killer cells.

Citation Format: Jonathan F. Head, Xian P. Jiang, Robert L. Elliott. Iron upregulates HLA-G mRNA expression of the human MCF-7 breast cancer cell line. [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 1507. doi:10.1158/1538-7445.AM2013-1507
Abstract 1511: Stromal gene signature regulated by TGFβ signaling predicts poor clinical outcome in ovarian cancer .
Authors: Cheon Dong-Joo , Tong Yunguang , Sim Myung-Shin , Cui Xiaojiang , Berel Dror , Tighiouart Mourad , Wiedemeyer Wolf Ruprecht , Karlan Beth , Orsulic Sandra
Published: August 2014

Despite the clinical and histopathological similarities in initial disease presentation, patients with advanced ovarian cancer rapidly develop chemoresistance and succumb to the disease. In order to identify genes that are associated with poor patient survival, we analyzed microarray datasets of 795 advanced ovarian cancer patients and correlated gene expression levels with clinical data. We identified a 60-gene signature that was strongly associated with poor survival. Many genes in the signature were stromal genes, including POSTN, FN1, COL11A1, COL6A1, COL3A1, LOX, TIMP3, and VCAN. The same stromal signature was enriched in ovarian cancer metastases and in a cisplatin-resistant A2780 human ovarian cancer cell line. Expression of many stromal genes was induced by TGFβ1 and the effect was reversed by A83-01, a potent TGFβ1 inhibitor. Interestingly, we found that ovarian cancer stem cells (CSC) also express TGFβR2 and several TGFβ1-target genes, including COL6A1 and COL3A1, which were previously implicated in cisplatin resistance. Culturing of A2780 cells on collagen substrate resulted in increased cisplatin resistance and expansion of the CSC population. Our results suggest that TGFβ1 signaling induces stromal gene expression and forms a favorable microenvironment for ovarian CSC, thereby leading to cisplatin resistance and poor patient survival.

Citation Format: Dong-Joo Cheon, Yunguang Tong, Myung-Shin Sim, Xiaojiang Cui, Dror Berel, Mourad Tighiouart, Wolf Ruprecht Wiedemeyer, Beth Karlan, Sandra Orsulic. Stromal gene signature regulated by TGFβ signaling predicts poor clinical outcome in ovarian cancer . [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 1511. doi:10.1158/1538-7445.AM2013-1511
Abstract 153: Hypothyroidism and hyperthyroidism in relation to endometrial cancer: A prospective study.
Authors: Kueck Angela , Kang Jae Hee , Stevens Richard , Curhan Gary , Alexander Erik , Rosner Bernard , DeVivo Immaculata , Tworoger Shelley
Published: August 2014

Background

Thyroid hormones may be involved in tumorigenesis and metastasis in hormone sensitive cancers. Thyroid hormones strongly influence the menstrual cycle, implantation and fertility, suggesting that thyroid hormones also could play a role in the development of endometrial cancer (EC), which is hormone-sensitive. No prospective studies have examined this association.

Methods

We prospectively evaluated the association between self-reported physician-diagnosed hypothyroidism and hyperthyroidism in the Nurses’ Health Study (NHS) from 1976 - 2010. Reports of thyroid dysfunction were based on self-report on biennial follow-up questionnaires, and have shown high validity when compared to medical records. Incident EC cases were confirmed with medical, pathology and death records. Multivariate rate ratios (RRs) for endometrial cancer were calculated, adjusting for major EC risk factors (i.e., body mass index, smoking, parity, age at last birth, oral contraceptive use, family history, age at menarche, age at menopause, postmenopausal hormone use, type 2 diabetes, and height).

Results

During 34 years of follow-up in NHS, we documented 917 cases of incident EC. Compared to those without any reported thyroid dysfunction, the multivariable RR for hypothyroidism was 0.99 (95% CI, 0.75-1.30) and the null associations were consistent across categories of time since hypothyroidism diagnosis. The multivariable RR for hyperthyroidism was 1.38 (95% CI 0.79 - 2.41). An association was observed with longer time since hyperthyroidism diagnosis (median time since hyperthyroidism diagnosis was 6 years in NHS); the RR for EC associated with 6+ years since hyperthyroidism diagnosis was 2.08 (95% CI, 1.02, 4.23).

Conclusions

Overall self-reported hypothyroidism and hyperthyroidism were not associated with risk of EC; however, longer time since hyperthyroidism diagnosis may be associated with an increased risk of EC. Due to the overall small number of cases with hyperthyroidism, on-going analyses will incorporate results from the Nurses’ Health Study II to increase power for these analyses.

Citation Format: Angela Kueck, Jae Hee Kang, Richard Stevens, Gary Curhan, Erik Alexander, Bernard Rosner, Immaculata DeVivo, Shelley Tworoger. Hypothyroidism and hyperthyroidism in relation to endometrial cancer: A prospective study. [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 153. doi:10.1158/1538-7445.AM2013-153
Abstract 1542: Th1 and Th2 cytokines determine how CD40 activation changes human macrophage function in vitro.
Authors: Luheshi Nadia , Davies Gareth C. , Poon Edd , McCourt Matthew , Legg James W.
Published: August 2014

Tumour associated macrophages (TAM) often adopt an M2-like immunosuppressive phenotype, promoting tumour growth and metastasis. However, TAM display phenotypic plasticity, and can be repolarised by agents including CD40 agonists to a Th1-immunostimulatory, tumoricidal M1 phenotype, inhibiting tumour growth. T cell-activating therapies synergise with CD40 agonists to induce more extensive TAM repolarisation and tumour growth inhibition, indicating that T cell-derived cytokines may enhance CD40 agonist effects on TAM. We therefore set out to determine how Th1 and Th2 cytokines shape the functional consequences of CD40 activation on human macrophages in vitro. We report that CD40 ligand (CD40L) activates human monocyte-derived macrophages in a similar, but not identical fashion to bacterial LPS in the absence of Th1 or Th2 cytokines. Under these conditions, CD40L activated macrophages adopt an M2-like phenotype, with low tumoricidal activity and producing high levels of immunosuppressive IL-10. The Th1 cytokine IFNγ enhanced anti-tumour functions of CD40-activated macrophages, promoting a switch from immunosuppressive IL-10 to Th1-immunostimulatory IL-12p70 production, and enhancing direct tumoricidal activity. We found that the Th2 cytokines IL-4 and IL-13 also enhanced IL-12p70 production by CD40L-activated macrophages, but, unlike IFNγ, these cytokines did not inhibit IL-10 production. IL-4 and IL-13 also promoted a smaller enhancement in CD40-induced macrophage tumoricidal activity than IFNγ. Thus the presence of Th1 / 2 T cell-derived cytokines in the tumour microenvironment may shape the outcome of CD40 agonist therapy by modulating CD40 agonist effects on TAM.

Citation Format: Nadia Luheshi, Gareth C. Davies, Edd Poon, Matthew McCourt, James W. Legg. Th1 and Th2 cytokines determine how CD40 activation changes human macrophage function in vitro. [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 1542. doi:10.1158/1538-7445.AM2013-1542
Abstract 1544: Control of tumor-associated macrophage alternative activation by MIF.
Authors: Yaddanapudi Kavitha , Putty Kalyani , Rendon Beatriz , Lamont Gwyneth , Faughn Jonathan , Mitchell Robert
Published: August 2014

Tumor stromal alternatively activated macrophages are important determinants of anti-tumor T lymphocyte responses, intratumoral neovascularization and metastatic dissemination. Our recent efforts to investigate the mechanism of macrophage migration inhibitory factor (MIF) in antagonizing anti-melanoma immune responses reveal that macrophage-derived MIF is necessary for maximal macrophage alternative activation in melanoma-bearing mice. Both peripheral and tumor-associated macrophages (TAMs) isolated from melanoma bearing MIF-deficient mice display elevated pro-inflammatory cytokine expression and reduced anti-inflammatory, immunosuppressive and pro-angiogenic gene products compared to macrophages from tumor bearing MIF wildtype mice. Moreover, TAMs and myeloid-derived suppressor cells (MDSCs) from MIF-deficient mice exhibit reduced T lymphocyte immunosuppressive activities than do those from their wildtype littermates. Corresponding with reduced tumor immunosuppression and neoangiogenic potential by TAMs, MIF-deficiency confers protection against transplantable subcutaneous melanoma outgrowth and melanoma lung metastatic colonization. Finally, we report for the first time that our previously discovered MIF small molecule antagonist, 4-iodo-6-phenylpyrimidine (4-IPP), recapitulates MIF-deficiency in vitro and in vivo and attenuates tumor polarized macrophage alternative activation, immunosuppression, neoangiogenesis and melanoma tumor outgrowth. These studies are the first to describe a functional contribution by MIF to tumor-associated macrophage alternative activation and provide justification for immunotherapeutic targeting of MIF in melanoma patients.

Citation Format: Kavitha Yaddanapudi, Kalyani Putty, Beatriz Rendon, Gwyneth Lamont, Jonathan Faughn, Robert Mitchell. Control of tumor-associated macrophage alternative activation by MIF. [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 1544. doi:10.1158/1538-7445.AM2013-1544
Abstract 1547: Tumor-derived macrophage Migration Inhibitory Factor and interleukin-6 cooperated in hypoxic accumulation of CD11b+Gr-1+ myeloid cells.
Authors: Zhu Guiquan , Tang Yaling , Li Ling , Liang Xinhua
Published: August 2014

Recently, myeloid derived suppressor cells (MDSCs) have gained much attention due to their role in tumor immunity suppression as well as promotion of angiogenesis, tumor cell invasion, and metastases. However, the mechanisms that regulate MDSCs recruitment have not been well clarified. Here, we demonstrated that hypoxic tumor microenvironment stimulate the recruitment of MDSCs through increased production of macrophage migration inhibitory factor (MIF) and interleukin-6 (IL-6) by head and neck squamous cell carcinoma (HNSCC) cells. Knock-down (KD) of HIF-1/2α in HNSCC cells decreased MIF level but did not inhibit the MDSCs migration, because HIF-1/2α KD increased NF-κB activity which further enhanced IL-6 secretion. In vivo administration of neutralizing IL-6 antibody and inhibitors against MIF and NF-κB decreased MDSCs accumulation, accompanied by reduced micro-vessel density (MVD), lymphatic vessel density (LVD), and xenograft tumor growth. The supplementation of MDSCs increased MVD, LVD, and xenograft tumor growth. In conclusion, hypoxic HNSCC cells may stimulate the accumulation of MDSCs through secreting MIF and IL-6. This process is mediated by interactions between NF-κB and HIF-1/2α. Strategies targeting hypoxic tumor microenvironment may represent attractive approaches to HNSCC therapy.

[G Zhu and Y Tang contributed equally to this paper.]

Citation Format: Guiquan Zhu, Yaling Tang, Ling Li, Xinhua Liang. Tumor-derived macrophage Migration Inhibitory Factor and interleukin-6 cooperated in hypoxic accumulation of CD11b+Gr-1+ myeloid cells. [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 1547. doi:10.1158/1538-7445.AM2013-1547
Abstract 1552: Cigarette smoke induces TNF-α secretion from macrophages through a pathway consisting of PPARγ, MUC1, ERK and JNK.
Authors: Xu Xiuling , Padilla Mabel T. , Wells Alexandria , Kim Kwang Chul , Lin yong
Published: August 2014

The glycoprotein Mucin 1 (MUC1 in human and Muc1 in animals) is expressed in the lung and has been shown to be associated with lung carcinogenesis. Recently we have found that cigarette smoke (CS) significantly increased MUC1 expression in mouse lung macrophages and that MUC1 is required for CS-induced tumor necrosis factor-alpha (TNF-α) secretion from macrophages. Because CS potently elicits chronic pulmonary inflammation that is important for the development of lung cancer, we investigated the cell signaling pathways for CS-induced MUC1 expression and MUC1-mediated TNF-α secretion in macrophages using an in vitro model. The results showed that: CS extract (CSE) strongly induced MUC1 expression at both the mRNA and protein levels; Suppression of PPARγ effectively blocked CSE-induced MUC1 expression in THP1 cells, suggesting CSE induces MUC1 expression through PPARγ-mediated transcription; The induction of TNF-α expression was effectively blocked when ERK and JNK were blocked, suggesting these pathways are involved in CSE-induced TNF-α expression in macrophages; Knockdown of MUC1 significantly suppressed CSE-induced ERK and JNK activation; and Suppression of PPARγ that effectively blocked CSE-induced MUC1 expression also inhibited CSE-induced TNF-α secretion from macrophages. Thus, our results suggest a possible involvement of a signaling pathway consisting of PPARγ, MUC1 and ERK/JNK for CSE-induced TNF-α secretion from macrophages that may contribute to inflammation-associated lung cancer development. Note: This work is supported by the NIH/NIEHS grant 1R01ES017328.

Citation Format: Xiuling Xu, Mabel T. Padilla, Alexandria Wells, Kwang Chul Kim, yong Lin. Cigarette smoke induces TNF-α secretion from macrophages through a pathway consisting of PPARγ, MUC1, ERK and JNK. [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 1552. doi:10.1158/1538-7445.AM2013-1552
Abstract 1553: Down-expression of ATM gene and protein associated with canine breast tumors.
Authors: Raposo Talita Mariana Morata , Bueno Renata C. , de Carvalho Márcio , Fonseca-Alves Carlos E. , Tinucci-Costa Mirela , De Nardi Andrigo B. , Rogatto Silvia R. , Laufer-Amorim Renée
Published: August 2014

Background: Ataxia telangiectasia mutated (ATM) gene encodes a protein associated with DNA damage repair that plays an important role in the maintenance of genome integrity. ATM transcript and protein down-regulation have been reported in sporadic breast carcinomas and the absence of ATM protein expression was also significantly associated with distant metastasis in women, but the mechanisms involved in the ATM deregulation are not well established.

Aim: Due to the fact that mammary tumor biological behavior is similar between canine and humans, we evaluate ATM gene and protein expression in canine breast cancer and compared with normal mammary gland from dogs.

Patients and methods: Fresh samples of non-metastatic (n=23) and metastatic mammary carcinomas (n=11) and normal breasts (n=10) were evaluated for ATM gene expression by RT-qPCR and from the same animals, another sample of tumor was collected, formalin-fixed and paraffin-embedded to assess ATM protein expression by immunohistochemistry. Evaluation of ATM protein expression was performed by the distribution of positively-immunostained tumor cells in scores: 1 (<25% cells positive), 2 (26% to 50% cells positive) and 3 (>51% cells positive).

Results: Down expression of ATM was detected in tumors samples (non-metastatic and metastatic mammary carcinomas) when compared to normal breast samples (P= 0,0414). Additionally, scores 1 and 2 of ATM protein expression were exclusively observed in metastatic tumors presenting score 1: 18% (2 out of 11), score 2: 55% (6 out to 11) and non metastatic tumors presenting the same score 1 and 2: 17% (4 out to 23) while all normal breast tissues showed score 3 (P=0,0087).

Conclusions: Down expression of ATM transcript was confirmed by ATM protein expression. Low levels of ATM suggest an involvement of DNA damage repair transcript and protein in tumorigenesis process of canine mammary tumors, as in women, corroborating for the use of dogs as a natural comparative model to study mammary gland tumors in humans.

Citation Format: Talita Mariana Morata Raposo, Renata C. Bueno, Márcio de Carvalho, Carlos E. Fonseca-Alves, Mirela Tinucci-Costa, Andrigo B. De Nardi, Silvia R. Rogatto, Renée Laufer-Amorim. Down-expression of ATM gene and protein associated with canine breast tumors. [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 1553. doi:10.1158/1538-7445.AM2013-1553
Abstract 1554: Inducible B-Raf and c-Src models as tools for drug development.
Authors: Ehlert Jan E. , Mutschler Bettina , Mueller Melanie , Lingnau Andreas , Hoffmann Steffen , Weber Holger , Kubbutat Michael H. G.
Published: August 2014

C-Src and B-Raf are critical kinases in signal transduction often involved in the onset of cancer and therefore representing promising targets for drug development. In order to assess the kinase-specific inhibitory effect in cellular and in-vivo models, we have generated cell lines exogenously expressing either c-Src or the commonly found mutant B-RafV600E in a regulatable manner, either by Doxicyclin-suppression or Hydroxy-Tamoxifen activation, respectively. Despite moderate kinase expression levels we observe significantly increased kinase activity in the activated state which can be monitored by substrate specific phospho-ELISAs as well as by analysis of phenotypic responses such as soft agar growth. Moreover, these cellular models are applicable for in-vivo analyses, showing strong in vivo growth in the activated state while no tumor growth occurs in the unactivated state. The applicability of these models in drug development is exemplified using c-Src inhibitor Dasatinib and B-Raf inhibitor Sorafenib.

Citation Format: Jan E. Ehlert, Bettina Mutschler, Melanie Mueller, Andreas Lingnau, Steffen Hoffmann, Holger Weber, Michael H. G. Kubbutat. Inducible B-Raf and c-Src models as tools for drug development. [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 1554. doi:10.1158/1538-7445.AM2013-1554
Abstract 1558: Imaging of metastatic B16F10 melanoma in mice using tumor-homing stem cells.
Authors: Shrestha Tej B. , Balivada Sivasai , Basel Matthew , Pyle Marla , Bossmann Stefan H. , Troyer Deryl L.
Published: August 2014

Tumors, like wounds, recruit cells from surrounding tissues and bone marrow. Some of these cells provide support and nutrition, while others participate in an inflammatory response. Bone marrow mesenchymal stem cells (MSCs) exhibit this tropism toward wounds and other areas of pathology. Consequently, stem cell technology for diagnostic and therapeutic purposes is an emerging field in cancer biology. Like bone marrow MSCs, mouse neural progenitor cells (NPCs) tend to migrate toward cancer sites. We found that these cells migrate toward B16F10 lung melanomas, making them very good candidates for delivery of chemotherapeutic drugs, genes of interest, and imaging contrast agents. Among imaging techniques, bioluminescent imaging is non-invasive. Gaussia luciferase (Gluc) is particularly useful, as it is non-toxic and has stronger luminescence than other known luciferases. In the presence of its substrate coelenterazine (coel), Gluc luminescence peaks near 470 nm. In this study, we engineered NPCs to secrete Gluc. These Gluc-engineered cells were injected systemically to C57BL/6 mice bearing metastatic B16F10 lung melanoma and mice were given coel intravenously. Imaging of the mice on the Caliper IVIS Lumina II showed bioluminescence in disseminated areas. The distribution of bioluminescence suggested that the engineered NPCs migrated to the tumor sites and secreted Gluc. In confirmation, necropsy revealed black metastatic tumors in the locations emitting bioluminescence. Although IV injection of metastatic B16F10 lung melanoma cells usually generates lung tumors, we found dissemination of melanoma in other parts of the body. The Gluc-expressing NPCs could find the tumors as early as 2 days post tumor-cell injection, long before tumors were palpable. Thus, this system offers promise for improving cancer diagnosis. For improved imaging, this bioluminescence system can be coupled with other fluorescent dyes for red shifted output. Finally, the same sensitivity and tumor tropism suggests that the NPCs could be useful delivery cells for tumor-targeted therapeutics.

Citation Format: Tej B. Shrestha, Sivasai Balivada, Matthew Basel, Marla Pyle, Stefan H. Bossmann, Deryl L. Troyer. Imaging of metastatic B16F10 melanoma in mice using tumor-homing stem cells. [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 1558. doi:10.1158/1538-7445.AM2013-1558
Abstract 1563: Mouse models for cancer research available from The Jackson Laboratory Repository.
Authors: Beckwith Jason , Lutz Cathleen M. , Mills Kevin D. , Rockwood Stephen F. , Sasner Michael , Donahue Leah Rae
Published: August 2014

The Jackson Laboratory Repository serves as a centralized facility for the development, rederivation, distribution and cryopreservation of mouse models. Hundreds of new strains are added annually to one of the largest collections of characterized mouse strains available to the international biomedical research community.

This poster features a detailed description of recently added mouse lines having cancer research applications across multiple therapeutic areas. A wide range of models are ideally suited for studies related to cellular cancers, solid tumors, and conditional/inducible cancer-associated gene expression. To highlight transplantation/engraftment and "humanized" mice research, several mouse lines with improved engraftment and specialized human cell colonization are also described. Such animals complement our NSG (NOD scid IL2rγc-/-) and NRG (NOD Rag1-/- IL2rγc-/-) mice as ideal tools for engraftment with a wide range of malignant or non-malignant human or mouse tissues, as well as a diverse spectrum of cell lines. In addition, newly available Cre-lox, Tet-On/Tet-Off, and fluorescent tool strains are illustrated.

An essential infrastructural component to The Jackson Laboratory Repository is a robust quality control program that surveys allele identity and genetic background, and excludes unwanted alleles. In addition, The Jackson Laboratory's Rare and Orphan Disease Center is a new initiative that focuses on partnering with scientists, foundations, and other experts to enable the development, standardization, optimization, and rapid distribution of preclinical models for drug discovery.

The Jackson Laboratory Repository maintains a searchable online resource to find mice (JAXMice database: www.jaxmice.jax.org/query). Anyone wishing to donate their mouse strains for cryopreservation/distribution may submit them using our online submission form (www.jax.org/donate-a-mouse). The Jackson Laboratory Repository is supported through our NCI designated Cancer Center and other NIH mechanisms, The Howard Hughes Medical Institute, The Ellison Medical Foundation and several private charitable foundations.

Please stop by for a detailed list of recently added mouse lines with cancer research applications across multiple therapeutic areas. Please also visit our cancer-related strains website at: http://jaxmice.jax.org/cancer/index.html

Citation Format: Jason Beckwith, Cathleen M. Lutz, Kevin D. Mills, Stephen F. Rockwood, Michael Sasner, Leah Rae Donahue. Mouse models for cancer research available from The Jackson Laboratory Repository. [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 1563. doi:10.1158/1538-7445.AM2013-1563
Abstract 1588: Canonical Wnt signaling mediates resistance of colorectal cancer cells to radiation therapy.
Authors: Emons Georg , Möller Janneke , Spitzner Melanie , Kendziorra Emil , Gaedcke Jochen , Rave-Fränk Margret , Pukrop Tobias , Ghadimi Michael , Ried Thomas , Grade Marian
Published: August 2014

Background: The clinical response of locally advanced rectal cancers to preoperative chemoradiotherapy is very heterogeneous. We previously identified TCF4, the key downstream effector of the Wnt/β-catenin signaling pathway, as over-expressed in tumors that were resistant to chemoradiotherapy. The aim of this study was to explore the potential functional relevance of Wnt/ß-catenin signaling for mediating treatment resistance.

Methods: Using RNA interference, TCF4 and β-catenin were inhibited in the colorectal cancer cell lines SW837 and SW480. Subsequently, these cell lines were irradiated with varying doses of X-rays. In addition, normal retinal epithelial cells (RPE) were stimulated with Wnt-3a for 28 hours, and also irradiated. Wnt/β-catenin activity was assessed using the TOPFLASH/FOPFLASH reporter assay, and induction of Axin2 was confirmed by Western blotting.

Results: Protein levels of both TCF4 and β-catenin were considerably reduced following RNAi, accompanied by a decreased TOP/FOP reporter activity. Importantly, silencing of both proteins led to a pronounced radiosensitization in SW837 and SW480 cells. Exogenous stimulation of canonical Wnt signaling in RPE cells using Wnt-3a resulted in an increased Axin2 expression and an increased TOP/FOP reporter activity. Importantly, however, this activation of canonical Wnt signaling was associated with significant increase in resistance to radiation therapy.

Conclusion: TCF4 was found to be over-expressed in resistant rectal carcinomas, and its RNAi-mediated silencing caused a significant radiosensitization. To determine whether this effect was dependent on canonical Wnt signaling, we inhibited ß-catenin, another key factor of this pathway and binding partner of TCF4, and observed a similar sensitization to radiation therapy. Follow-up experiments show that external stimulation of canonical Wnt signaling confers radiation resistance in normal cells. These data suggest that targeting canonical Wnt signaling may represent a potential therapeutic strategy for sensitizing tumor cells to radiation. In vivo studies are ongoing to validate these results.

Citation Format: Georg Emons, Janneke Möller, Melanie Spitzner, Emil Kendziorra, Jochen Gaedcke, Margret Rave-Fränk, Tobias Pukrop, Michael Ghadimi, Thomas Ried, Marian Grade. Canonical Wnt signaling mediates resistance of colorectal cancer cells to radiation therapy. [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 1588. doi:10.1158/1538-7445.AM2013-1588