bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 4644: PDGFRα/β and VEGFR2 SNPs incolorectal cancer.
Authors: Estevez-Garcia Purificacion , Castaño A , Martin Ana , Lopez-Rios Fernando , Iglesias J , Muñoz-Galvan Sandra , Lopez-Calderero Iker , Molina-Pinelo Sonia , Pastor Maria D. , Carnero Amancio , Pa-Ares Luis , Garcia-Carbonero Rocio
Published: August 2014

Angiogenesis plays an essential role in tumor growth and metastasis, and is a major target in cancer therapy. VEGFR and PDGFR are key players involved in this process. The purpose of this study was to assess the incidence of genetic variants in these receptors and its potential clinical implications in colorectal cancer (CRC). VEGFR2, PDGFRα and PDGFRβ mutations were evaluated by sequencing their tyrosine kinase domains in 8 CRC cell lines and in 92 samples of patients with CRC. Correlations with clinicopathological features and survival were analyzed. Four SNPs were identified, three in PDGFRα [exon 12 (A12): c.1701A>G; exon13 (A13): c.1809G>A; and exon 17 (A17): c.2439+58C>A) and one in PDGFRβ (exon19 (B19): c.2601A>G) (figure 1). SNP B19 was present in 58% of tumor samples and in 4 cell lines (LS174T, LS180, SW48, COLO205), and this was associated with higher PDGFR and pPDGFR protein levels. 5-year survival was greater for patients with PDGFR B19 wild type tumors (AA) than for those harboring the G-allele genotype (GA or GG) (51%vs17%; P=0.073). Multivariate analysis confirmed SNP B19 (P=0.029) was a significant prognostic factor for survival, independent of age (P=0.060) or TNM stage (P<0.001). In conclusion, PDGFRβ exon 19 c.2601A>G SNP is commonly encountered in CRC patients and is associated with increased pathway activation and poorer survival. Implications regarding its potential influence in response to PDGFR targeted agents remain to be ellucidated.

Citation Format: Purificacion Estevez-Garcia, A Castaño, Ana Martin, Fernando Lopez-Rios, J Iglesias, Sandra Muñoz-Galvan, Iker Lopez-Calderero, Sonia Molina-Pinelo, Maria D. Pastor, Amancio Carnero, Luis Pa-Ares, Rocio Garcia-Carbonero. PDGFRα/β and VEGFR2 SNPs incolorectal 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 4644. doi:10.1158/1538-7445.AM2013-4644
Abstract 4646: Notch and DLL4 expression in bevacizumab-treated colon cancer patients.
Authors: Negri Francesca , Bozzetti Cecilia , Crafa Pellegrino , Lagrasta Costanza , Cavalli Stefano , Pedrazzi Giuseppe , Gardini Giorgio , Tamagnini Ione , Porzio Rosa , Cavanna Luigi , Paties Carlo , Romano Ida , Musolino Antonino , Tomasello Gianluca , Petronini Pier Giorgio , Ardizzoni Andrea
Published: August 2014

Background: In order to investigate mechanisms of resistance to angiogenesis inhibitors, Notch and Delta-like 4 ligand (DLL4) expression was correlated with response and survival in a series of bevacizumab treated advanced colon cancer patients. DLL4-mediated Notch signalling has recently emerged as an attractive target for angiogenesis-based cancer therapies, given that DLL4 is an important component of Notch-mediated stem cell self-renewal and vascular development. DLL4-induced Notch signalling mediates tumour-resistance to anti-VEGF therapy by inducing the formation of large vessels and activating multiple pathways in preclinical models. Method: Notch and DLL4 expression was evaluated by immunohistochemistry (IHC) on 65 primary colon cancer patients enrolled within randomized clinical trials assessing first-line bevacizumab plus chemotherapy. Results: Notch positivity was localized to the cytoplasm of malignant epithelial cells. In all, 10/60 (17%) evaluable primary tumours had a high Notch expression. Five of the 10 cases (50%) with high Notch expression experienced progressive disease compared with 5/49 (10%) Notch negative cases (p=0.008). Median progression-free survival (PFS) was 2.8 months for Notch positive cases compared with 13.07 months for Notch negative cases (p=0.045). Membranous and/or cytoplasmic DLL4 immunoreactivity of tumour vessels was observed in 30/56 (54%) evaluable colon cancers. No correlation was found between DLL4 expression and clinical outcome. Conclusion: Clinical trials investigating the therapeutic efficacy of bevacizumab in colon cancer did not explore the impact of DLL4-Notch pathway on response and clinical outcome. Our results seem to suggest that high Notch expression might be involved in tumour resistance in colon cancer patients treated with bevacizumab.

Citation Format: Francesca Negri, Cecilia Bozzetti, Pellegrino Crafa, Costanza Lagrasta, Stefano Cavalli, Giuseppe Pedrazzi, Giorgio Gardini, Ione Tamagnini, Rosa Porzio, Luigi Cavanna, Carlo Paties, Ida Romano, Antonino Musolino, Gianluca Tomasello, Pier Giorgio Petronini, Andrea Ardizzoni. Notch and DLL4 expression in bevacizumab-treated colon cancer patients. [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 4646. doi:10.1158/1538-7445.AM2013-4646

Note: This abstract was not presented at the AACR Annual Meeting 2013 because the presenter was unable to attend.
Abstract 4654: Genomic and proteomic profiling of signaling pathways targeted by a multi-kinase inhibitor, sorafenib.
Authors: Masuda Mari , Honda Kazufumi , Yamada Tesshi
Published: August 2014

Background: Deregulation of intracellular signaling by various genetic alterations is a hallmark of cancer. Therapeutics targeting such activated signaling pathways have achieved significant progress, but the precise pathway(s) targeted by sorafenib has remained unclear.

Methods: Using high-density reverse-phase protein arrays, we profiled the phosphorylation status of 180 signaling molecules in the 120 pathways registered on the NCI-Nature curated database in 23 hepatocellular carcinoma (HCC) cell lines showing different sensitivities to sorafenib. We also sequenced the entire coding regions of 518 protein kinase genes in 20 overlapping HCC cell lines.

Results: We identified 25 non-synomymous alterations in the 4 mTOR and MAPK pathway genes (RPS6KA1, RPS6KA6, RPS6KB2, and BRAF) in HCC cell lines 1 to 11. We also found that the levels of phosphorylated S6 ribosomal protein and c-Raf were associated with the sensitivity of the HCC cell lines to sorafenib. A mTOR inhibitor, AZD8055, exerted growth-inhibitory activity against sorafenib-resistant HCC cell lines.

Discussion: Although a recent phase III clinical trial has demonstrated the effectiveness of sorafenib in patients with unresectable HCC, only a small proportion of the patients achieved the anticipated therapeutic benefits. The present results indicate cross-talk between the mTOR and MAPK signaling pathways in sorafenib-resistant HCC cell lines. Use of a combination of mTOR and MAPK inhibitors may improve the efficacy of molecular targeting therapy against HCC.

Citation Format: Mari Masuda, Kazufumi Honda, Tesshi Yamada. Genomic and proteomic profiling of signaling pathways targeted by a multi-kinase inhibitor, sorafenib. [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 4654. doi:10.1158/1538-7445.AM2013-4654
Abstract 4659: Phase II study of a weekly liposomal paclitaxel formulation (Genexol®-PM) and gemcitabine® combination chemotherapy in patients with advanced biliary cancer.
Authors: Keon Uk Keon UK , Kim Jin Young , Do Young Rok , Song Hong Suk , Jeon Young June , Ryoo Hun Mo , Bae Sung Hwa , Kim Jong Gwang , Baek Jin Ho , Chae Yee Soo , Kim Min Kyoung , Lee Kyung Hee , Cho Yoon Young
Published: August 2014

OBJECTIVE: Advanced biliary cancer is associated with a poor prognosis due to wide resistance to chemotherapeutic agents. This phase II clinical trial was conducted to determine the efficacy and safety of weekly gemcitabine and liposomal paclitaxel in patients with unresectable of metastatic biliary cancer.

METHODS: The eligibility criteria were patients 1) with pathologically proven biliary cancer, 2) with an ECOG performance status 0 to 2, 3) aged more than 18, 4) with measurable lesions, 5) with adequate hematologic, renal and liver functions, and 6) who provided written informed consent. Each treatment cycle was consisted of gemcitabine 1000 mg/m² and liposomal paclitaxel (Genexol-PM)100 mg/m² on days 1, 8 followed by rest perioid of 14 days. It was repeated until the appearance of disease progression or unacceptable toxicity up to maximal 10 cycles. The primary end point of this study was reponse rate, and secondary end points included toxicity, progression free survival and overall survival.

RESULTS: Forty five patients were enrolled; median age was 63 years; male (n=33) and female (n=12). The median number of cycles administered was 4.0 (range,1-10). Thirty nine patients were assessable for efficacy. Ten partial responses and 18 stable diseases were confirmed. Giving an overall response rate was 25.6% and disease control rate was 71.8% in per-protocol population. The median time to progression and median overall survival was 4.0 (95% CI; 3.17∼4.83) months and 8.13 (95% CI; 4.76∼11.51) months, respectively. MUC4 expression was not significantly correlated with overall survival and progression free survival (p=0.391).

CONCLUSION: Weekly gemcitabine combined with Genexol-PM® appears to be effective against advanced biliary cancers. Further randomized trials are needed to confirm this finding.

Citation Format: Keon Uk Keon Uk, Jin Young Kim, Young Rok Do, Hong Suk Song, Young June Jeon, Hun Mo Ryoo, Sung Hwa Bae, Jong Gwang Kim, Jin Ho Baek, Yee Soo Chae, Min Kyoung Kim, Kyung Hee Lee, Yoon Young Cho. Phase II study of a weekly liposomal paclitaxel formulation (Genexol®-PM) and gemcitabine® combination chemotherapy in patients with advanced biliary 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 4659. doi:10.1158/1538-7445.AM2013-4659
Abstract 466: Tumor-induced fibrocytes mediate immune suppression via Indoleamine Oxidase.
Authors: Zhang Hua , Mackall Crystal L.
Published: August 2014

Fibrocytes are hematopoietic stem cell-derived, fibroblast precursors implicated in chronic inflammation, fibrosis and wound healing, whereas human myeloid derived suppressor cells (MDSC) have been described as CD14-CD11b+CD33+HLADR- cells that exert T cell suppression via ROS, ARG1 and iNOS2. Here we report that patients with metastatic pediatric sarcomas have abnormal expansions of myeloid cells within elutriated mononuclear apheresis samples of pediatric cancer patients. Like myeloid derived suppressor cells, these cells are CD14-CD11b+, but they also cells bear markers of classical fibrocytes, including surface α smooth muscle actin, fibroblast activation protein, collagen I/V, and HLA-DR, but are PM2K negative. They also mediate angiogenesis and are CD33lo/-CD34+CD15+CD16+CD66b+CD68+PDL1+ and IL4Rα+. While classical fibrocytes serve as antigen presenting cells and augment immune reactivity, fibrocytes from cancer patients suppressed anti-CD3 mediated T cell proliferation, primarily via indoleamine oxidase (IDO). Interestingly the degree of fibrocyte expansion observed in individual patients directly correlated with the frequency of circulating GATA3+CD4+ cells (R=0.80) and monocytes from healthy donors cultured with IL-4 differentiated into fibrocytes with the same phenotypic profile and immunosuppressive properties as those observed in patients with cancer. We conclude that fibrocytes are a subset of myeloid derived suppressor cells expanded in cancer patients likely in response to Th2 immune deviation. These cells may contribute to tumor progression via both immune evasion and angiogenesis.

Citation Format: Hua Zhang, Crystal L. Mackall. Tumor-induced fibrocytes mediate immune suppression via Indoleamine Oxidase. [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 466. doi:10.1158/1538-7445.AM2013-466
Abstract 4691: Class III beta tubulin is a strong predictive marker for taxane-based first-line palliative chemotherapy in recurrent or metastatic gastric cancer.
Authors: Hwang Jun-Eul , Kim Dae-Eun , Shim Hyun-Jeong , Bae Woo-Kyun , Oh In-Jae , Cho Sang-Hee , Chung Ik-Joo
Published: August 2014

Backgrounds. Class III β tubulin (TUBB3) is a prognostic marker in various tumors and role of TUBB3 in advanced gastric cancer is not clearly defined yet. We analyzed the significance of TUBB3 expression along with ERCC1 in recurrent or metastatic gastric cancer patients receiving taxane based first-line palliative chemotherapy.

Methods. We reviewed 146 patients with advanced gastric adenocarcinoma who received taxane based first-line palliative chemotherapy between 2004 and 2010 at Chonnam National University Hwasun hospital. Immunohistochemical stain of TUBB3 and ERCC1 was done in paraffin-embedded tumor tissue. We evaluated response to chemotherapy, progression-free survival (PFS), and overall survival (OS).

Results. A total of 146 patients with advanced gastric cancer received docetaxel and cisplatin (n=15), or paclitaxel and cisplatin (n=131). The median PFS was significantly shorter for patients with TUBB3 high expression than patients with TUBB3 low expression (3.63 versus 6.67 months, p=0.001). OS was not associated with TUBB3 expression status (12.9 versus 13.1 months, p=0.769). In multivariate analysis, only TUBB3 was related to shorter PFS (HR 2.74, 95% CI 1.91-3.91, p=0.001). Patients with ERCC1 high expression showed lower response rate than patients with ERCC1 low expression (24% versus 63.2%, p=0.001), however ERCC1 showed no clinical influence on PFS and OS.

Conclusions. TUBB3 is a strong predictive marker in recurrent or metastatic gastric cancer patients receiving taxane based first-line palliative chemotherapy. Clinical impact of ERCC1 is not evident in this setting.

Citation Format: Jun-Eul Hwang, Dae-Eun Kim, Hyun-Jeong Shim, Woo-Kyun Bae, In-Jae Oh, Sang-Hee Cho, Ik-Joo Chung. Class III beta tubulin is a strong predictive marker for taxane-based first-line palliative chemotherapy in recurrent or metastatic gastric 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 4691. doi:10.1158/1538-7445.AM2013-4691
Abstract 4694: Nuclear expression of IL23p19 shows indolent tumor features and favorable outcome in patients with colorectal cancer.
Authors: Zlobec Inti , Helbling Melina , Diamantis-Karamitopoulou Eva , Koelzer Viktor H. , Dawson Heather , Lugli Alessandro
Published: August 2014

Background: The IL23 signaling pathway is involved in chronic inflammation and autoimmune disorders but its role in cancer progression is not fully elucidated. The aim of this study was to characterize the expression of IL23 subunits p40 and p19 as well as IL23R in the normal-adenoma-carcinoma-metastasis cascade of colorectal cancers and investigate the relationship between these proteins and clinicopathological, molecular and clinical outcome data in colorectal cancer patients.

Methods: Immunohistochemistry for IL12p40 (p40 subunit shared between IL12 and IL23), IL23 (polyclonal), IL23p19 (monoclonal) and IL23R was performed on a multiple-punch tissue microarray of 213 colorectal cancers from patients with full clinicopathological and follow-up information. Normal tissues, matched adenomas and lymph node metastases were assessed as well as microsatellite instability (MSI), KRAS and BRAF mutations.

Results: IL12p40, IL23, IL23p19 and IL23R expression were significantly increased in colorectal cancers in comparison to all other tissue types (p<0.001; all). Cytoplasmic and/or membranous expression of all proteins was unrelated to clinicopathological or molecular features. Unexpectedly, 45 cases (23.1%) showed distinct nuclear IL23p19+ expression associated with smaller tumor diameter (p=0.0333), early TNM stage (p=0.0186), absence of venous (p=0.0124) and lymphatic invasion (p=0.0493), lymph node negativity (p=0.0621) and MSI (p=0.0032). IL23p19+ patients had more favorable overall survival (p=0.0634); none had distant metastatic disease (p=0.0146). No relationship with KRAS or BRAF was observed.

Conclusion: Our results of a more indolent, less aggressive cancer expressing nuclear IL23p19 support a more “protective” role of this protein in colorectal cancer progression. These findings add to the current controversy regarding the pro- or anti-tumor effects of IL23 signaling in cancer.

Citation Format: Inti Zlobec, Melina Helbling, Eva Diamantis-Karamitopoulou, Viktor H. Koelzer, Heather Dawson, Alessandro Lugli. Nuclear expression of IL23p19 shows indolent tumor features and favorable outcome in patients with colorectal 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 4694. doi:10.1158/1538-7445.AM2013-4694
Abstract 4701: Cancer-specific genomic rearrangements used to quantify disease burden in plasma from patients with colorectal cancer.
Authors: Schøler Lone V. , Thomsen Rune , Tobiasen Heidi , Vang Søren , Nielsen Hans Jørgen , Laurberg Søren , Ørntoft Torben F. , Andersen Claus L.
Published: August 2014

Tumor-specific chromosomal rearrangements have the potential to serve as highly sensitive biomarkers for tumor detection. Such alterations are not present in normal cells and should be exquisitely specific. It is well recognized that some colorectal cancers (CRC) release small amounts of DNA into plasma. Accordingly, sensitive and specific assays targeting these tumor-specific rearrangements should enable quantification of changes in disease burden if applied to serial plasma samples. Here we report the establishment of efficient procedures for 1) identification of tumor-specific genomic rearrangements, 2) development of sensitive and specific assays to these, 3) quantification of these rearrangements in plasma, and 4) results from applying these procedures to quantify disease burden in serial plasma samples from CRC patients. The tumor-specific genomic rearrangements were identified using a combination of next generation paired-end sequencing of large-insert libraries and SNP array based DNA copy number analysis. Nested real-time PCR assays targeting the rearrangements were applied to plasma samples in order to quantify disease burden. Initially the approach was tested using the CRC cell line SW620 as a tumor model, and serial dilutions of SW620 DNA and normal human plasma DNA as models of patient plasma. Multiple cell line specific rearrangements were identified and nested real-time PCR analysis of these in the serial dilutions revealed an excellent linearity, recovery across three log scales. Also the sensitivity was very good. As little as 6.4 pg cell line DNA could be detected in a background 20 ng of normal plasma DNA, corresponding to DNA from 1-2 tumor cells circulating in 2 ml plasma. We are currently in the process of analyzing 25 CRC patients from which we have more than three years of follow-up and serial plasma samples collected every third month, starting the day before surgery and until month 36. The results from these analyses will be presented. In conclusion: We have shown that the combination of large insert paired end sequencing and copy number analysis readily identifies multiple tumor-specific rearrangements that can be targeted by sensitive and specific nested real-time PCR assays to quantify disease burden in plasma samples.

Citation Format: Lone V. Schøler, Rune Thomsen, Heidi Tobiasen, Søren Vang, Hans Jørgen Nielsen, Søren Laurberg, Torben F. Ørntoft, Claus L. Andersen. Cancer-specific genomic rearrangements used to quantify disease burden in plasma from patients with colorectal 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 4701. doi:10.1158/1538-7445.AM2013-4701
Abstract 4708: Anti-tumor phenotype of tumor-associated macrophages are induced by COX-2 inhibition via the up-regulation of IFN-γ.
Authors: Nakanishi Yuki , Nakatsuji Masato , Seno Hiroshi , Chiba Tsutomu
Published: August 2014

BACKGROUNDS AND AIMS

Macrophages are a major constituent in tumor stroma, and macrophages infiltrating tumors are called tumor-associated macrophages (TAMs). Macrophages have a functional plasticity depending on the stimuli from microenvironment. M1 macrophages are induced by Th1 cytokines, and exerted anti-tumor function. On the contrary, M2 macrophages are induced by Th2 cytokines, and showed pro-tumor phenotype. TAMs are considered to shift from M1 to M2 during tumor progression. However, the mechanism for this M1/M2 switch is poorly understood. Cyclooxygenase-2 (COX-2)/PGE2 pathway is up-regulated in a variety of cancers including colorectal cancers, and promotes the cancer progression. COX-2 inhibition shows anti-tumor effect in familial adenomatous polyposis patients and in Apc knockout mice. Although COX-2 inhibition affects cytokine expression profiles and tumor microenvironments, the relationship between COX-2/PGE2 pathway and TAM phenotypes remains unclear.

The aim of this study is to uncover the effects of COX-2 inhibition on TAM phenotypes and cytokine expression.

METHODS

Cytokine expression and the phenotypes of TAMs infiltrating the stroma of ApcMin/+ mouse polyps were examined by qRT-PCR and immunohistochemistry in terms of M1 and M2 dichotomy. Selective COX-2 inhibitor was administered to ApcMin/+ mice, and the effects of COX-2 inhibition on cytokine expression and TAM phenotypes were evaluated. Furthermore, by using mouse peritoneal macrophages, the direct effects of COX-2 inhibitor on Th1- or Th2-related cytokines on macrophage phenotypes were examined.

RESULTS

Th2 cytokines were predominant in ApcMin/+ mouse polyps, and TAMs were polarized to M2. Selective COX-2 inhibitor skewed TAM phenotypes from M2 to M1, accompanied by tumor regression in ApcMin/+ mice. Concomitantly, the expression of M1-related cytokine IFN-γ was significantly up-regulated by COX-2 inhibition, although that of M2-related cytokines IL-4, IL-13, and IL-10 was not significantly altered. Administration of COX-2 inhibitor alone was insufficient to alter the phenotypes of mouse peritoneal macrophages from M2 to M1. However, IFN-γ treatment polarized the macrophage phenotypes from M2 to M1 even in the presence of M2-related cytokines IL-4, IL-13, and IL10.

CONCLUSIONS

Our results suggest that COX-2 inhibition induced anti-tumor (M1) phenotype of TAMs via the up-regulation of IFN-γ, and subsequently, may suppress intestinal tumor progression in ApcMin/+ mouse polyps.

Citation Format: Yuki Nakanishi, Masato Nakatsuji, Hiroshi Seno, Tsutomu Chiba. Anti-tumor phenotype of tumor-associated macrophages are induced by COX-2 inhibition via the up-regulation of IFN-γ. [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 4708. doi:10.1158/1538-7445.AM2013-4708
Abstract 471: A case of bone-metastatic breast cancer resulted in complete remission 5 years after the comprehensive treatment including autologous formalin-fixed tumor vaccine.
Authors: Kuranishi Fumito , Ohno Tadao
Published: August 2014

Introduction: All cases, so far we have experienced,of bone-metastatic breast carcinoma patients show the downhill course to a tragic ending despite of intensive chemo-radiation therapy. We here report a case of advanced breast cancer that has been additively treated with autologous formalin-fixed tumorvaccine (AFTV) (Clin Cancer Res 10: 1574-9 2004). Case : A 53 years old female was diagnosed bearing a bone metastasis of breast cancer at vertebra Th7 by bone scintigraphy,

MRI and SPECT. The breast was operated, irradiated according to palliative radiation therapy with 36 Gy, vaccinated with AFTV, and then chemotherapy (CEF) was done as the last step of adjuvant therapy. Afterwards aromatase inhibitor and zoledronic acid hydrate was used for maintenance therapy. Five years later, the bonemetastasis has disappeared and she was diagnosed complete remission. Tumor markers have been in normal level . QOL of this patient is good at present.

Conclusion: Complete remission of bone metastasis of breast carcinoma is very rare. In this case, the preceded use of AFTV and radiation to the chemotherapy may have contributed to the cure. Additional use of AFTV should be considered in a larger scale clinicaltrial to improve therapeutic efficacy in bone-metastatic breast carcinoma.

Citation Format: Fumito Kuranishi, Tadao Ohno. A case of bone-metastatic breast cancer resulted in complete remission 5 years after the comprehensive treatment including autologous formalin-fixed tumor vaccine. [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 471. doi:10.1158/1538-7445.AM2013-471
Abstract 4724: No-lyse no-wash immunophenotyping.
Authors: Bradford Jolene A. , Kerndt Rick
Published: August 2014

Immunophenotyping whole blood is a primary application in the study of blood cell populations and their functions. Red blood cells have traditionally been removed during sample preparation by lysis methods or density-gradient selection to enable the study of nucleated cell populations. This processing of whole blood has its draw backs due to the potential loss of cells of interest thru inadvertent lysis and unintentional selection of fragile and rare cells including blast cells, cancer cells, and cancer stem cells. The application of a No-Lyse No-Wash protocol in human whole blood samples benefit from a streamlined protocol that reduces cell loss. Testing of mouse whole blood presents additional challenges for cancer researchers due to the limited sample volume available (≤100 μL/day/animal), particularly in longitudinal studies. Specifically, these small volumes limit the ability to perform immunophenotyping experiments with required controls. This work describes a No-lyse No-wash method which takes advantage of acoustic focusing technology. Immunophenotyping panels are shown for both human and mouse whole blood. A total of 10μL of whole blood is stained in a 50 μL total volume and then diluted 400-fold in PBS. A fluorescence threshold is used to distinguish the white blood cell populations from the more abundant red blood cell population. In the mouse immunophenotyping example, CD45 is used to threshold on white blood cells. For the human immunophenotyping example, a cell permeable nucleic acid stain is used to threshold on all nucleated cells. At this dilution, the coincidence of the target population with red blood cells is reduced sufficiently so scatter signals are reliable for population identification. The significantly higher sample collection rates available with acoustic cytometry allows for the acquisition of high quality data utilizing a No-lyse No-wash protocol in a timely manner. Difficult-to-collect and precious samples like mouse blood can be stained and diluted without washing or performing RBC removal procedures. Loss of cells due to sample preparation is essentially eliminated, and ability to detect the cells of interest is enabled.

Citation Format: Jolene A. Bradford, Rick Kerndt. No-lyse no-wash immunophenotyping. [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 4724. doi:10.1158/1538-7445.AM2013-4724
Abstract 4725: High expression of TLR4 in hepatocellular carcinoma promotes proliferation and drug resistance.
Authors: Tseng Ping-Hui , Liu Yi-Ting , Cheng Yi-Chen
Published: August 2014

Toll-like receptors (TLRs) expressed in hematopoietic cells have emerged as sensors for pathogen-associated molecular patterns (PAMPs) and damage-associated molecular patterns (DAMPs), and are critical for innate immunity. Meanwhile, TLRs, especially TLR4, are also found to be expressed in various tumor cells. In hepatocellular carcinoma (HCC), TLR4 in marcorphages and hepatic stellate cells has been shown to be involved in promoting tumorigenesis. It has also been reported that hepatocytes with hepatitis B or C virus infection, the risk factor for HCC, lead to up-regulation of TLR4. However, the association between TLR4 in hepatocytes and HCC has not been well-studied. Here, the hepatotic cells with different TLR4 expression from human cancer cell lines and DEN-induced mouse HCCs were investigated. The results demonstrated that proliferation and colony formation in cells with higher TLR4, but not in those with low TLR4, was enhanced in response to LPS. Additionally, Akt activation, cyclinD1 induction, IL-6 production and STAT3 activation were detected in LPS-sensitive cancer cells. We also found that HCC with LPS pretreatment was resistant to anti-tumor drug-induced cell death. In xenograft model, the results were also in line with in vitro experiments. Briefly, TLR4 expressed in liver cancer cells provide advantage in growth and survival in the present of PAMPs and DAMPs. Better understanding of TLR activation in HCC might help the development of novel therapeutic approaches.

Citation Format: Ping-Hui Tseng, Yi-Ting Liu, Yi-Chen Cheng. High expression of TLR4 in hepatocellular carcinoma promotes proliferation and drug resistance. [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 4725. doi:10.1158/1538-7445.AM2013-4725
Abstract 473: Long term clinical and biochemical outcomes following immunotherapy with an novel autologous immunomodulated vaccine in patients with prostate cancer.
Authors: Kreutz Fernando T. , Schroeder Gustavo , Berger Milton
Published: August 2014

Background: VAP is an new autologous immunomodulate vaccine, that was previously demonstrate to be safe in an initial phase I trial with local advance prostate cancer patients. The extension of this initial trail into a Phase IIb is now been reported. The primary endpoint of the study reported here is clinical response (PSA and survival) in local advance (T2 and T3) prostate cancer patients, with safety and immunologic responses as secondary endpoints.

Methods: Tumor cells from 107 prostatectomy patients were collected (HCPA - Porto Alegre - Brazil). 48 (45%) patients with T3 or T2 with co-morbidity factors were enrolled. 22 patients received the vaccine and 26 were in the control group. Vaccine was given by intradermal injections (4x weekly, 2x monthly and after 3 months). First two doses also ontained BGC as adjuvant. DTH was measured 48 hours after the vaccine doses that did not contained BCG. PBMCs and were collected via apheresis at baseline and D+54 for Tcell proliferation assay in some patients. Clinical flow up was performed and all standard care was given to all patients.

Results: The overall follow up was 7.4 years. No grade 3 or 4 toxicity attributable to vaccine was noted. Side effects were largely limited to grade 1 or 2 injection site reactions. DTH was positive (= or higher than 5 mm) in 73% of the vaccinated patients. Cancer related mortality was 9% (2/22) on the vaccinated group and 19% (5/26) on the non-vaccinated group. PSA was undetectable (less than 0.04 ng/ml) in 85% (17/20) of the vaccinated patients and in 48% (10/21) of the non-vaccinated patients after 5 years of follow up.In vitro specific T cell proliferation was demonstrated in vaccinatedpatients.

Conclusions: VAP is safe and well tolerated vaccine. There is evidence of clinical activity and immune changes in selected patients. Further studies to confirm these results are warranted. (Financial support from FINEP - Brazilian Innovation Agency)

Citation Format: Fernando T. Kreutz, Gustavo Schroeder, Milton Berger. Long term clinical and biochemical outcomes following immunotherapy with an novel autologous immunomodulated vaccine in patients with prostate 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 473. doi:10.1158/1538-7445.AM2013-473
Abstract 4731: Discovering cancer biomarkers using a novel cytokine screening panel.
Authors: Manimala Joseph , Gupte Prachi , Swaminathan Ashok , Tituana Jorge P. , Hansen Caitlin , Mathew Anu , Stewart David , Oberoi Pankaj , Wohlstadter Jacob
Published: August 2014

Cytokines and chemokines that are produced in the tumor micro-environment play major roles in tumorigenesis, tumor progression, and apoptosis. They are involved in the modulation of angiogenesis and consequently, in tumor growth and metastases. Cytokine/chemokine expression by tumor cells leads to tumor proliferation and enhances tumor cell survival. Profiling these biomarkers may be useful for cancer diagnosis. For example, biomarkers such as IFNγ, TNFα, IL-1β, MCP-1, and IL-8 are overexpressed in breast carcinoma; IL-2, IL-4, IL-6, IL-8, IL-10, IL-16, and TARC are associated with prostate cancer; and increased expression levels of TNFα, IL-6, and IL-1β are implicated in colon cancer.

We have developed and validated multiplexed panels of 30 assays that allow users to simultaneously detect cytokines and chemokines with high precision and accuracy in different human matrices. These panels detect IFNγ, IL-1β, IL-2, IL-4, IL-6, IL-8, IL-10, IL-12p70, IL-13, TNFα, IL-5, IL-7, GM-CSF, IL-1α, IL-17A, IL-15, IL-16, IL-12/IL-23 p40, TNFβ, and VEGF MIP-1α, MIP-1β, Eotaxin, Eotaxin-3, IP-10, TARC, IL-8 (high abundance), MDC, MCP-1, and MCP-4.

The MSD® platform has the ability to quantitate these biomarkers in multiple sample types with superior sensitivity and performance, quantifying many low abundance analytes at levels below 0.5 pg/mL.

A challenge with current multiplex assays is a lack of analytical validation and lot-to-lot reproducibility. These new multiplex panels were developed with highly characterized and qualified reagents, and the assays were developed in accordance with industry guidelines. The panels demonstrated suitable spike recovery and dilution linearity performance in different biological matrices. Diluents, other kit components, and an optimized assay protocol are provided to achieve optimum performance. Each assay in the panel is well characterized and shows low non-specific binding. Each has been evaluated for potential assay and analytical interference. Controls are available for each panel.

These panels are suitable tools for accurately quantitating cytokines and chemokines in various tumor samples. Data are presented for samples from normal subjects as well as samples from benign prostatic hyperplasia and prostate cancer patients (serum), cirrhosis and liver cancer patients (serum), kidney cancer patients (plasma), and colon cancer patients (serum).

Citation Format: Joseph Manimala, Prachi Gupte, Ashok Swaminathan, Jorge P. Tituana, Caitlin Hansen, Anu Mathew, David Stewart, Pankaj Oberoi, Jacob Wohlstadter. Discovering cancer biomarkers using a novel cytokine screening panel. [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 4731. doi:10.1158/1538-7445.AM2013-4731
Abstract 4738: TLR8 stimulation enhances cetuximab-mediated natural killer cell lysis of head and neck cancer cells and dendritic cell cross priming of EGFR-specific CD8+ T cells.
Authors: Stephenson Ryan M. , Lim Chwee Ming , Matthews Maura , Dietsch Gregory , Hershberg Robert , Ferris Robert L.
Published: August 2014

Background: Cetuximab is an anti-epidermal growth factor receptor (EGFR) monoclonal antibody (mAb) that prolongs survival in the treatment of head and neck cancer (HNC), but only in 10-20%. An immunological mechanism of action including natural killer (NK) cell-mediated antibody-dependent cellular cytotoxicity (ADCC) has been suggested. We investigated the effects of activating toll-like receptor (TLR)-8 to enhance activity of cetuximab-stimulated, FcγR bearing cells.

Objective: To determine the capability of TLR8-stimulation to enhance the activation and function of NK cells and dendritic cells (DC) in the presence of cetuximab-coated HNC cells.

Methods: Peripheral blood mononuclear cells (PBMC), NK, DC and CD8+ T cells were isolated and analyzed using 51Cr release ADCC, flow cytometry analysis, cytokine ELISA, and EGFR853-861 tetramer staining.

Results: TLR8 stimulation of unfractionated PBMC led to enhanced cetuximab-mediated ADCC in healthy donors (p=0.01) and HNC patients (p<0.05), which was dependent on NK cells. Secretion of Th1 cytokines TNFα(p<0.0001), IFNγ(p<0.0001), and IL-12p40(p<0.005) was increased. NK cells in TLR8 stimulated PBMC showed higher degranulation marker CD107a expression after incubation with cetuximab-treated HNC cells (p<0.001). TLR8 stimulation enhanced DC maturation markers CD80, CD83, and CD86 in co-culture with NK cells and cetuximab-treated HNC cells. Stimulation of DC using TLR8 agonist plus cetuximab-activated NK cells significantly increased EGFR-specific CD8+ T cell induction, compared to either condition alone (p<0.01).

Discussion: VTX-2337 and cetuximab combination therapy can activate innate and adaptive anti-cancer immune responses. Further investigation in human trials will be important for determining the clinical benefit from this combination, and for determining biomarkers of response.

Citation Format: Ryan M. Stephenson, Chwee Ming Lim, Maura Matthews, Gregory Dietsch, Robert Hershberg, Robert L. Ferris. TLR8 stimulation enhances cetuximab-mediated natural killer cell lysis of head and neck cancer cells and dendritic cell cross priming of EGFR-specific CD8+ T 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 4738. doi:10.1158/1538-7445.AM2013-4738
Abstract 4748: Therapy model for advanced intracerebral B16 mouse melanoma using radiosurgery combined with immunotherapy.
Authors: Smilowitz Henry M. , Sasso Dan , Lee Edward W. , Micca Peggy , Goh Gyuhyeong , Dilmanian F Avrahamn
Published: August 2014

A reproducible therapy model for advanced intracerebral B16 melanoma is reported. Implanted tumors (D0), suppressed by a single 15 Gy radiosurgical dose of 100 kVp X-rays (D8) were further suppressed by a single ip injection of a Treg depleting Mab given two days prior to the initiation (D9) of four weekly then eight bi-monthly sc injections of GMCSF-transfected, mitotically disabled B16 cells. The results of seven independent experiments were similar to the combined result: the Median (Days) [SD/Total N] of survival went from 15[1.09/62] (No Treatment Control) to 35.8[8.8/58] (Radiosurgery-Only) to 52.5[13.5/57] (Radiosurgery Plus Immunotherapy); Radiosurgery-Only and Radiosurgery Plus Immunotherapy groups were highly distinct from Controls as well as from one another (p<0.0001). Within two weeks after immunization, tumors in mice receiving Radiosurgery Plus Immunotherapy were significantly smaller than tumors in mice treated only with Radiosurgery. Splenocytes and lymph node cells from immunized mice showed increased interferon Y production when cultured with syngenic tumor cells. We hypothesize that our model will be useful for the development and testing of novel combination therapies for brain tumors.

Citation Format: Henry M. Smilowitz, Dan Sasso, Edward W. Lee, Peggy Micca, Gyuhyeong Goh, F Avrahamn Dilmanian. Therapy model for advanced intracerebral B16 mouse melanoma using radiosurgery combined with immunotherapy. [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 4748. doi:10.1158/1538-7445.AM2013-4748
Abstract 4750: Maximising the activity of immune checkpoint blockade through rational combination.
Authors: Mullins Stefanie R. , Watkins Amanda , Morrow Michelle , Vallee Theresa La , Robbins Paul , Stewart Ross , Wilkinson Robert
Published: August 2014

Harnessing the power of the immune system to identify and eliminate cancer is a promising area of development in cancer therapy. Increasing T-cell activation, in particular, is now a clinically validated approach to the treatment of cancer. Cytotoxic T-lymphocyte antigen 4 (CTLA-4, CD152) and B7-H1 (PD-L1, CD274) are part of a complex system of receptors and ligands that are involved in limiting T-cell activation through the delivery of inhibitory signals. MedImmune is currently developing tremelimumab (anti-CTLA4) and MEDI4736 (anti-PD-L1), for the treatment of advanced or metastatic solid tumours. Both of these antibodies antagonise T-cell inhibition signals with the aim of enhancing anti-tumor T cell activity, and tremelimumab has demonstrated benefit in a sub-set of patients with melanoma (Ribas, 2010).

Published studies characterising the immunogenic nature of some existing cancer therapeutics provide reason to believe that a combination strategy, utilising checkpoint blockade together with current standard of care, or novel therapeutics, could result in increased activity. We have established and characterised a panel of syngeneic mouse tumor models and employed these to explore the immune effects of a number of existing cancer therapies using pharmacodynamic (PD) and tumor growth inhibition (TGI) endpoints. In addition mouse cross reactive anti-CTLA-4 and PD-L1 antibodies were used to assess the potential for combination with these existing therapeutics and the effect of schedule on PD and TGI activity.

Citation Format: Stefanie R. Mullins, Amanda Watkins, Michelle Morrow, Theresa La Vallee, Paul Robbins, Ross Stewart, Robert Wilkinson. Maximising the activity of immune checkpoint blockade through rational combination. [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 4750. doi:10.1158/1538-7445.AM2013-4750
Abstract 4760: Evaluation of the progesterone receptor status using three different antibodies: A comparison by Allred score system.
Authors: Madeira Klesia P. , Daltoe Renata , Carvalho Alex , Rezende Lucas , Silva Ian , Rangel Leticia
Published: August 2014

Abstract:

Breast cancer (BC) hormonal receptors (progesterone, PgR and estrogen) status is assessed by immunohistochemistry (IHC), a specific, sensitive, and accessible method that guide BC treatment. We evaluated PgR expression in 53 BC cases using 3 anti-PgR antibodies (AB): monoclonal (SP42 and RP636) and polyclonal ab62621.

Methods:

Primary BC cases (with signed informed consent) were used to generate tissue microarray platforms, where PgR expression was accessed by IHC and evaluated by the Allred score. Categorical and quantitative data are shown in percentage and mean, respectively. Concordance (CON) and correlation among ABs were analyzed by Kappa factor (K), Spearman's correlation coefficient (ρ) or intraclass correlation coefficient. Staining patterns of each AB were compared by paired T-Test.

Findings:

We noted poor CON and K between ab62621 vs SP42 (CON=64.1%; K=0.247), and ab62621 vs RP636 (CON=62.3%; K=0.204), but higher CON between SP42 vs RP636 (CON 90.6%; K =0.738). Data were corroborated by Mc Nemar statistical test (p=0.019, p=0.014 and p>0.05, respectively). Regarding staining intensity among PgR+ samples, we found higher proportion of weak staining and lower IS for ab62621 (48.3%; mean IS=1.6), when compared to SP42 (20.0%, mean IS=2.1, T-test p<0.01) and RP636 (2.3, 21.9%, T-test p<0.01). Within the entire sample, similar results were observed following ρ: SP42 vs RP636 (ρ=0.8103); ab62621 vs SP42 (ρ=0.3524); ab62621 vs RP636 (ρ=0.4075). As for proportion of stained cells and PS, among PgR+ samples, the mean values for ab62621 (75.4%; 4.8) were significantly higher than those of SP42 (56.3%, 4.3; T-test p<0.01) and RP636 (60.1%; 4.2; T-test p<0.01). Similar data were found after analyzing PS for the entire sample: SP42 vs RP636 (ρ=0.8588); SP42 vs ab62621(ρ=0.4832); RP636 vs ab62621(ρ=0.4050).

Interpretation:

Our data indicate that anti-PgR monoclonal ABs, RP636 and SP42, are, unlike ab62621, equally suitable to test BC PgR status by IHC due to their higher accuracy. Therefore, we suggest their clinical use during BC diagnosis; thus, enabling more precise therapeutic decisions to treat BC.

Citation Format: Klesia P. Madeira, Renata Daltoe, Alex Carvalho, Lucas Rezende, Ian Silva, Leticia Rangel. Evaluation of the progesterone receptor status using three different antibodies: A comparison by Allred score system. [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 4760. doi:10.1158/1538-7445.AM2013-4760

Note: This abstract was not presented at the AACR Annual Meeting 2013 because the presenter was unable to attend.